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Adult T-cell leukemia/lymphoma (ATLL) is a type of blood cancer related to human T-cell lymphotropic virus type 1 (HTLV-1). The principal aim of this study was to investigate cellular processes related to innate immune response, intracellular protein transport, and translational initiation regulation in individuals afflicted with ATLL and Acute lymphoblastic leukemia (ALL). Whole blood samples and peripheral blood mononuclear cells were collected from 10 viral ATLL patients and 10 ALL subjects. Real-time quantitative PCR was then performed to quantify mRNA expression levels of SMC6, FANCM, EIF4H, WDR7, RAB3GAP2, and IFN α/β. The study revealed some distinctions between ATLL and ALL patients. Particularly, RAB3GAP2 level (P = 0.028) was found to be elevated in ATLL patients compared to ALL. Conversely, expression levels of IFN-β (P = 0.31), SMC6 (P = 0.68), WDR7 (P = 0.43), EIF4H (P = 0.38), and FANCM (P = 0.57) were diminished in ATLL patients in contrast to ALL. These proteins play a pivotal role in both translation and immune activation, suggesting a potential correlation between the observed disparities and the virus-mediated progression of cancer. However, it is worth noting that the expression differences in FANCM, EIF4H, SMC6, and WDR7 between ATLL and ALL were minimal. This proposes that the underlying molecular mechanisms governing ATLL and ALL may largely overlap concerning these cellular processes. However, considerable increased expression of RAB3GAP2 was observed in ATLL compared to ALL., Competing Interests: Declarations. Ethics approval and consent to participate: informed consent to participate was obtained from all of the participants in the study. In addition, this study was approved at Tehran University of Medical Sciences by the Ethics Committees of Medical Sciences Research (IR.TUMS.SPH.REC.1399.112). This study adhered to the Declaration of Helsinki. Consent for publication: N/A. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

The heterodimeric Rab3GAP complex is a guanine nucleotide exchange factor (GEF) for the Rab18 GTPase that regulates lipid droplet metabolism, ER-to-Golgi trafficking, secretion, and autophagy. Why both subunits of Rab3GAP are required for Rab18 GEF activity and the molecular basis of how Rab3GAP engages and activates its cognate substrate are unknown. Here we show that human Rab3GAP is conformationally flexible and potentially autoinhibited by the C-terminal domain of its Rab3GAP2 subunit. Our high-resolution structure of the catalytic core of Rab3GAP, determined by cryo-EM, shows that the Rab3GAP2 N-terminal domain binds Rab3GAP1 via an extensive interface. AlphaFold3 modelling analysis together with targeted mutagenesis and in vitro activity assay reveal that Rab3GAP likely engages its substrate Rab18 through an interface away from the switch and interswitch regions. Lastly, we find that three Warburg Micro Syndrome-associated missense mutations do not affect the overall architecture of Rab3GAP but instead likely interfere with substrate binding., Competing Interests: Competing interests: J.E.B. reports personal fees from Scorpion Therapeutics and Reactive Therapeutics and research contracts from Novartis and Calico Life Sciences. All other authors declare no competing interests., (© 2025. The Author(s).)

Kinesin-3 KIF1A (UNC-104 in C. elegans) is the major axonal transporter of synaptic vesicles and mutations in this molecular motor are linked to KIF1A-associated neurological disorders (KAND), encompassing Charcot-Marie-Tooth disease, amyotrophic lateral sclerosis and hereditary spastic paraplegia. UNC-104 binds to lipid bilayers of synaptic vesicles via its C-terminal PH (pleckstrin homology) domain. Since this interaction is relatively weak and non-specific, we hypothesize that other, more specific, interaction schemes exist. From the literature, it is evident that UNC-104 regulator SYD-2 interacts with UNC-10 and that UNC-10 itself interacts with RAB-3 bound to synaptic vesicles. RT-PCR and Western blot experiments expose genetic relationships between unc-10 and syd-2, but not between unc-10 and rab-3. Also, neither unc-10 nor rab-3 affects UNC-104 expression. However, co-immunoprecipitation and bimolecular fluorescence complementation (BiFC) assays reveal functional interactions between UNC-104, SYD-2, UNC-10 and RAB-3. Though both SNB-1 and RAB-3 are actively transported by UNC-104, motility of RAB-3 is facilitated in the presence of SYD-2 and UNC-10. Deletion of UNC-104's PH domain did not affect UNC-104/RAB-3 colocalization, but significantly affected UNC-104/SNB-1 colocalization. Similarly, motility of RAB-3-labeled vesicles is only slightly altered in nematodes carrying a point mutation in the PH domain, whereas movement of SNB-1 is significantly reduced in this mutant. Western blots from purified fractions of synaptic vesicles reveal strong reduction of UNC-104 in rab-3/unc-10 double mutants. Our findings suggest that the UNC-10/SYD-2 complex acts as a functional linker to connect UNC-104 to RAB-3-containing vesicles. Thus, this linker complex contributes to the specificity of motor/cargo interactions., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Some subtypes of hereditary spastic paraplegia (HSP), especially with autosomal recessive inheritance (AR-HSP), have been reported rarely. In this study, we report the clinical features and molecular results of three unrelated Iranian patients with rare subtypes of HSP, including SPG76, SPG56, and SPG69; thereafter, we compare them to other reported cases. Three patients who were clinically diagnosed with HSP and born to consanguineous parents underwent molecular assessment by whole-exome sequencing (WES), followed by Sanger sequencing and co-segregation analysis. Two patients carried biallelic pathogenic variants in CAPN1, or CYP2U1, resulting in SPG76, and SPG56, respectively. Additionally, another patient presented with a variant of uncertain significance (VUS) in the gene associated with SPG69, known as RAB3GAP2. Variants of CAPN1 and RAB3GAP2 are novel while the CYP2U1 variant has been previously reported. The patient with the RAB3GAP2 variant is the second reported SPG69 case. Our findings emphasize that the rare forms of AR-HSP may be more prevalent in communities with a high rate of consanguineous marriages, and WES can be a highly effective tool for identifying pathogenic variants in these communities. Also, the CYP2U1 variant seems to be a founder mutation because it was previously reported in 8 patients of three families from the Middle East. These results expand the variant spectrum of the CAPN1 and RAB3GAP2 genes. Also, given the association of variants in CAPN1 and RAB3GAP2 with a diverse array of phenotypes, we propose the use of the terms "CAPN1-related disorders" and "RAB3GAP2-related disorders" as alternatives to HSP76 and HSP69, respectively., Competing Interests: Declarations. Competing interests: The authors declare no competing interests. Ethics approval: This research was conducted in accordance with the principles outlined in the Declaration of Helsinki and received approval from the ethics board of the University of Social Welfare and Rehabilitation Sciences (IR.USWR.REC.1400.053) in Iran. Consent to participate: Informed consent was obtained from the patients or their guardians to participate in the project. Consent to publication: The authors affirm that patients or their guardians provided signed informed consent to publish the data of the project., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The intricate molecular and structural sequences guiding the formation and consolidation of memories within neuronal circuits remain largely elusive. In this study, we investigate the roles of two pivotal presynaptic regulators, the small GTPase Rab3, enriched at synaptic vesicles, and the cell adhesion protein Neurexin-1, in the formation of distinct memory phases within the Drosophila mushroom body Kenyon cells. Our findings suggest that both proteins play crucial roles in memory-supporting processes within the presynaptic terminal, operating within distinct plasticity modules. These modules likely encompass remodeling and maturation of existing active zones (AZs), as well as the formation of new AZs., (© 2024 Turrel et al.; Published by Cold Spring Harbor Laboratory Press.)

Current study aims to investigate the biological function of circular RNA (circRNA, circ_0000337) in cervical cancer (CC). Bioinformatic analyses were used to predict targets for circ_0000337 and miR-155-5p, and analyze the gene expression differences between cervical squamous cell carcinoma and endocervical adenocarcinoma (CESC) tissues and normal tissues. Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot were applied to assess mRNA and protein expressions of circ_0000337, microRNA-155-5p (miR-155-5p) and member RAS oncogene family (RAB3B), respectively. Following the establishment of gain/loss-of-function models, CCK-8 was performed to evaluate cell proliferation. Bioinformatics analysis, dual-luciferase reporter assay and RNA immunoprecipitation (RIP) were used to identify the interaction in circ_0000337, miR-155-5p, and RAB3B. Circ_0000337 and RAB3B were upregulated, while miR-155-5p was downregulated in CC tissues and cell lines. circ_0000337 overexpression promoted cell proliferation, circ_0000337 knock down inhibited cell proliferation by sponging miR-155-5p. RAB3B was a target of miR-155-5p which was positively regulated by circ_0000337. In the collected CC tissues, there was a negative correlation between miR-155-5p and circ_0000337 or RAB3B, and a positive correlation between circ_0000337 and RAB3B. miR-155-5p was positively, while RAB3B was negatively correlated with OS in patients with CC, and they were negatively correlated. In conclusion, circ_0000337 upregulates RAB3B by sponging miR-155-5p to promote CC cell proliferation., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

MAPK activating death domain (MADD) is a multifunctional protein regulating small GTPases RAB3 and RAB27, MAPK signaling, and cell survival. Polymorphisms in the MADD locus are associated with glycemic traits, but patients with biallelic variants in MADD manifest a complex syndrome affecting nervous, endocrine, exocrine, and hematological systems. We identified a homozygous splice site variant in MADD in 2 siblings with developmental delay, diabetes, congenital hypogonadotropic hypogonadism, and growth hormone deficiency. This variant led to skipping of exon 30 and in-frame deletion of 36 amino acids. To elucidate how this mutation causes pleiotropic endocrine phenotypes, we generated relevant cellular models with deletion of MADD exon 30 (dex30). We observed reduced numbers of β cells, decreased insulin content, and increased proinsulin-to-insulin ratio in dex30 human embryonic stem cell-derived pancreatic islets. Concordantly, dex30 led to decreased insulin expression in human β cell line EndoC-βH1. Furthermore, dex30 resulted in decreased luteinizing hormone expression in mouse pituitary gonadotrope cell line LβT2 but did not affect ontogeny of stem cell-derived GnRH neurons. Protein-protein interactions of wild-type and dex30 MADD revealed changes affecting multiple signaling pathways, while the GDP/GTP exchange activity of dex30 MADD remained intact. Our results suggest MADD-specific processes regulate hormone expression in pancreatic β cells and pituitary gonadotropes.

RAB3B is essential for the transportation and secretion within cells. Its increased expression is linked to the development and progression of various malignancies. However, understanding of RAB3B's involvement in carcinogenesis is mostly limited to specific cancer subtypes. Hence, exploring RAB3B's regulatory roles and molecular mechanisms through comprehensive cancer datasets might offer innovative approaches for managing clinical cancer. To examine the potential involvement of RAB3B in the development of cancer, we analyzed data from various sources including The Cancer Genome Atlas (TCGA), Genotype-Tissue Expression Project (GTEx), cBioPortal, HPA, UALCAN, and tissue microarray (TAM). Using bioinformatics techniques, we examined the correlation between RAB3B expression and prognosis, tumor heterogeneity, methylation modifications, and immune microenvironment across different cancer types. Our findings indicate that elevated RAB3B expression can independently predict prognosis in many tumors and has moderate accuracy for diagnosing most cancers. In most cancer types, we identified RAB3B mutations that showed a significant correlation with tumor mutational burden (TMB), mutant-allele tumor heterogeneity (MATH), and microsatellite instability (MSI). Abnormal DNA methylation patterns were also observed in most cancers compared to normal tissues. Additionally, we found significant correlations between RAB3B expression, immune cell infiltration, and immune scores across various cancers. Through pan-cancer analysis, we observed significant differences in RAB3B expression levels between tumors and normal tissues, making it a potential primary factor for cancer diagnosis and prognosis. The IHC results revealed that the expression of RAB3B in six types of tumors was consistent with the results of the pan-cancer analysis of the database. Furthermore, RAB3B showed potential associations with tumor heterogeneity and immunity. Thus, RAB3B can be utilized as an auxiliary diagnostic marker for early tumor detection and a prognostic biomarker for various tumor types., (© 2024. The Author(s).)

RAB3GAP1 is GTPase activating protein localized to the ER and Golgi compartments. In humans, mutations in RAB3GAP1 are the most common cause of Warburg Micro syndrome, a neurodevelopmental disorder associated with intellectual disability, microcephaly, and agenesis of the corpus callosum. We found that downregulation of RAB3GAP1 leads to a reduction in neurite outgrowth and complexity in human stem cell derived neurons. To further define the cellular function of RAB3GAP1, we sought to identify novel interacting proteins. We used a combination of mass spectrometry, co-immunoprecipitation and colocalization analysis and identified two novel interactors of RAB3GAP1: the axon elongation factor Dedicator of cytokinesis 7 (DOCK7) and the TATA modulatory factor 1 (TMF1) a modulator of Endoplasmic Reticulum (ER) to Golgi trafficking. To define the relationship between RAB3GAP1 and its two novel interactors, we analyzed their localization to different subcellular compartments in neuronal and non-neuronal cells with loss of RAB3GAP1. We find that RAB3GAP1 is important for the sub-cellular localization of TMF1 and DOCK7 across different compartments of the Golgi and endoplasmic reticulum. In addition, we find that loss of function mutations in RAB3GAP1 lead to dysregulation of pathways that are activated in response to the cellular stress like ATF6, MAPK, and PI3-AKT signaling. In summary, our findings suggest a novel role for RAB3GAP1 in neurite outgrowth that could encompass the regulation of proteins that control axon elongation, ER-Golgi trafficking, as well as pathways implicated in response to cellular stress., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Warburg Micro (WARBM) syndrome is a rare heterogeneous recessive genetic disorder characterized by ocular, neurological, and endocrine problems. To date, disease-causing variants in four genes have been identified to cause this syndrome; of these, RAB3GAP1 variants are the most frequent. Very little is known about WARBM syndrome in rural populations., Objectives: This study aims to investigate the genetics underpinnings of WARBM syndrome in a Pashtun family with two patients from Pakistan. The patients presented with spastic diplegia, severe intellectual disability, microphthalmia, microcornea, congenital cataracts, optic atrophy, and hypogonadism., Methods: Magnetic resonance imaging (MRI) analysis revealed pronounced cerebral atrophy including corpus callosum hypoplasia and polymicrogyria. Exome sequencing and subsequent filtering identified a novel homozygous missense variant NM_001172435: c.2891A>G, p.Gln964Arg in the RAB3GAP1 gene. The variant was validated, and its segregation confirmed, by Sanger sequencing., Results: Multiple prediction tools assess this variant to be damaging, and structural analysis of the protein shows that the mutant amino acid residue affects polar contact with the neighboring atoms. It is extremely rare and is absent in all the public databases. Taken together, these observations suggest that this variant underlies Micro syndrome in our family and is extremely important for management and family planning., Conclusions: Identification of this extremely rare variant extends the mutations spectrum of Micro syndrome. Screening more families, especially in underrepresented populations, will help unveil the mutation spectrum underlying this syndrome., (© 2023 International Society for Developmental Neuroscience.)

Rab GTPase 3C (RAB3C) is a peripheral membrane protein that is involved in membrane trafficking (vesicle formation) and cell movement. Recently, researchers have noted the exocytosis of RAB proteins, and their dysregulation is correlated with drug resistance and the altered tumor microenvironment in tumorigenesis. However, the molecular mechanisms of exocytotic RABs in the carcinogenicity of colorectal cancer (CRC) remain unknown. Researchers have used various in silico datasets to evaluate the expression profiles of RAB family members. We confirmed that RAB3C plays a key role in CRC progression. Its overexpression promotes exocytosis and is related to the resistance to several chemotherapeutic drugs. We established a proteomic dataset based on RAB3C, and found that dystrophin is one of the proteins that is upregulated with the overexpression of RAB3C. According to our results, RAB3C-induced dystrophin expression promotes vesicle formation and packaging. A connectivity map predicted that the cannabinoid receptor 2 (CB2) agonists reverse RAB3C-associated drug resistance, and that these agonists have synergistic effects when combined with standard chemotherapy regimens. Moreover, we found high dystrophin expression levels in CRC patients with poor survival outcomes. A combination of the dystrophin and RAB3C expression profiles can serve as an independent prognostic factor in CRC and is associated with several clinicopathological parameters. In addition, the RAB3C-dystrophin axis is positively correlated with the phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha isoform (PIK3CA) genetic alterations in CRC patients. These findings can be used to provide novel combined therapeutic options for the treatment of CRC., (© 2023 The Authors. Molecular Oncology published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Objective: To investigate the inhibitory effect of miR-125b-5p on proliferation and migration of osteosarcoma and the role of RAB3D in mediating this effect., Methods: The expression level of miR-125b-5p was detected by qRT-PCR in a normal bone cell line (hFOB1.19) and in two osteosarcoma OS cell lines (MG63 and HOS). A miR-125b-5p mimic or inhibitor was transfected in the osteosarcoma cell lines via liposome and the changes in cell proliferation and migration were detected with EDU and Transwell experiments. Bioinformatic analysis was conducted for predicting the target gene of miR-125b-5p, and the expression level of RAB3D in hFOB1.19, MG63, and HOS cells was detected by Western blotting. In the two osteosarcoma cell lines transfected with miR-125b-5p mimic or inhibitor, the expression levels of RAB3D mRNA and protein in osteosarcoma cells were examined with qRT-PCR and Western blotting. The effects of RAB3D overexpression, RAB3D knockdown, or overexpression of both miR-125b-5p and RAB3D on the proliferation and migration of cells were assessed using EDU and Transwell experiments., Results: The two osteosarcoma cell lines had significantly lower expression levels of miR-125b-5p ( P < 0.05). Bioinformatic analysis predicted that RAB3D was a possible target gene regulated by miR-125b-5p. In osteosarcoma cells, overexpression of miR-125b-5p significantly lowered the expression of RAB3D protein ( P < 0.05); inhibiting miR-125b-5p expression significantly decreased RAB3D expression only at the protein level ( P < 0.05) without obviously affecting its mRNA level. Modulation of miR-125b-5p and RAB3D levels produced opposite effects on proliferation and migration of osteosarcoma cells, and in cells with overexpression of both miR-125b-5p and RAB3D, the effect of RAB3D on cell proliferation and migration was blocked by miR-125b-5p overexpression ( P < 0.05)., Conclusion: Overexpression of miR-125b-5p inhibits the proliferation and migration of osteosarcoma cells by regulating the expression of RAB3D at the post-transcriptional level.

Warburg Micro syndrome (WARBM) is an autosomal recessive neuro-ophthalmologic syndrome characterized by microcephaly, microphthalmia, congenital cataracts, cortical dysplasia, corpus callosum hypoplasia, spasticity, and hypogonadism. WARBM is divided into four subtypes according to the causative genes, of which RAB3GAP1 (OMIM# 602536) accounts for the highest proportion. We collected detailed medical records and performed whole-exome sequencing (WES) for a congenital cataract patient. A novel heterozygous frameshift RAB3GAP1 variant was detected in a boy with a rare ocular phenotype of bilateral membranous cataracts accompanied by a persistent papillary membrane. Further copy number variation (CNV) analysis identified a novel deletion on chromosome 2q21.3 that removed 4 of the 24 exons of RAB3GAP1 . The patient was diagnosed with WARBM following genetic testing. The present study expands the genotypic and phenotypic spectrum of WARBM. It suggests applying whole exome sequencing (WES) and CNV analysis for the early diagnosis of syndromic diseases in children with congenital cataracts.

Many patients with prostate cancer (PCa) cannot be diagnosed until an advanced stage, which make PCa become a large threat to human health. It is an urgent need to explore novel biomarkers for accurate diagnosis and targets for the effective treatment of PCa. This study aimed to investigate the effects of RAB3D (which belongs to the secretory Rab GTPases) on the progression of PCa. The results showed that RAB3D was highly expressed in PCa tissues compared to normal tissues according to the gene expression omnibus dataset. Consistent with the bioinformatics results, RAB3D exhibited a higher expression in PCa cells. Overexpression of RAB3D promoted the proliferation, migration, and invasion of PCa cells, whereas the knockdown of RAB3D led to the opposite results. The procancer effects of RAB3D were further confirmed by the in vivo growth of xenograft model. Subsequently, RAB3D upregulated the PI3K/AKT signaling pathway both in vivo and in vitro. LY294002 (a PI3K inhibitor) rescued the RAB3D upregulation-induced promotion of malignant phenotypes of PCa cells. Furthermore, the transcription activity of RAB3D was found to be enhanced by aryl hydrocarbon receptor (AhR; a transcription factor). The AhR silencing-induced inhibition of the proliferation and migration of PCa cells was reversed by the overexpression of RAB3D. Taken together, RAB3D, upregulated by AhR, promotes the PCa progression by activating the PI3K/AKT signaling pathway., (© 2022 International Federation for Cell Biology.)

Background: The metastatic mechanisms of axillary lymph nodes (ALNs) in triple-negative breast cancer (TNBC) remain unclear. We aimed to identify the potential circRNA regulatory network in ALN metastasis., Methods: We performed whole transcriptome sequencing (WTS) to determine the expression profiles of RNAs and screen out differentially expressed messenger RNAs (DEMs), microRNAs (DEMis), and circRNAs (DECs) between ALN-positive and ALN-negative TNBC patients. Functional enrichment analysis and Kaplan-Meier survival analysis were utilized to unearth the potential regulatory mechanisms of the DEMs. A competing endogenous RNA (ceRNA) network was constructed using computational biology. The expression levels of DECs in cell lines were confirmed by real-time polymerase chain reaction (RT‒PCR)., Results: Following WTS and differential expression analysis, 739 DEMs, 110 DEMis, and 206 DECs were identified between ALN-positive and ALN-negative TNBC patients. Functional analysis indicated that the DEMs mainly functioned in carcinogenesis and tumor progression-related pathways. ceRNA networks containing eight circRNAs, six miRNAs, and eighteen mRNAs were developed. In the ceRNA network, two mRNAs (RAB3D and EDARADD) that were significantly associated with better overall survival and one mRNA (GSR) that predicted favorable recurrence-free survival in TNBC patients were chosen for further analysis. Then, a survival-related ceRNA network containing two DECs (hsa&#95;circ&#95;0061260 and hsa&#95;circ&#95;0060876), two DEMis (hsa-miR-5000-3p and hsa-miR-4792), and three mRNAs (GSR, RAB3D, and EDARADD) was identified. Then, two candidate DECs were validated by real-time PCR., Conclusion: Our research constructed a ceRNA network that provides novel insights into the molecular mechanism of ALN metastasis and potential therapeutic targets in TNBC., (© 2022. The Author(s).)

Little is known about the role of hsa&#95;circ&#95;0103232 in melanoma. This study researched the role of hsa&#95;circ&#95;0103232 in melanoma progression. Hsa&#95;circ&#95;0103232 expression in clinical tissues of melanoma patients and melanoma cells was detected by qRT-PCR. Hsa&#95;circ&#95;0103232 localization in melanoma cells was visualized by fluorescence in situ hybridization. Hsa&#95;circ&#95;0103232 effect on melanoma cells viability, proliferation, migration, and invasion was explored by cell counting kit-8 (CCK-8) assay, Edu experiment, wound healing assay, and Transwell experiment. RNA pull-down assay and dual-luciferase reporter gene assay were performed to verify the binding of hsa&#95;circ&#95;0103232 with miR-661, and the binding of miR-661 and RAB3D. Xenograft tumor models were constructed. Western blot and immunohistochemistry were used for protein expression detection. Hsa&#95;circ&#95;0103232 expression was increased in melanoma patients, indicating lower overall survival. Hsa&#95;circ&#95;0103232 was mainly expressed in the cytoplasm of melanoma cells. Silencing hsa&#95;circ&#95;0103232 suppressed melanoma cell viability, proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) ( P < 0.01). Hsa&#95;circ&#95;0103232 functioned as a sponge of miR-661 to increase RAB3D expression. miR-661 overexpression partially reversed hsa&#95;circ&#95;0103232 promoting effect on melanoma cells viability, proliferation, migration, invasion, and EMT ( P < 0.01). In melanoma patients, hsa&#95;circ&#95;0103232 expression was negatively correlated with miR-661 and positively correlated with RAB3D. Silencing hsa&#95;circ&#95;0103232 suppressed melanoma cell growth in vivo and Ki67 and RAB3D expression in xenograft tumors ( P < 0.01). Hsa&#95;circ&#95;0103232 is a tumor promoter in melanoma to enhance malignant phenotype and growth in vivo via sponging miR-661/RAB3D. Hsa&#95;circ&#95;0103232 may be a novel target for melanoma treatment.

We describe a sibling pair of Mennonite origin born from consanguineous parentage with a likely new phenotype of limb-girdle muscular dystrophy, short stature, ptosis, and tracheomalacia. Exome sequencing in the affected subjects identified a novel homozygous RAB3GAP2 missense variant as the potential causal variant. As RAB3GAP2 has been recently shown to be involved in the autophagy process, we analyzed patient-derived fibroblasts by fluorescence microscopy and demonstrated defective autophagic flux under rapamycin and serum starvation conditions when compared with wild-type cells. The phenotype in the siblings described here is distinct from Martsolf and Warburg's micro syndromes, the currently known diseases arising from RAB3GAP2 pathogenic variants. Thus, this work describes a potentially novel recessive phenotype associated with a RAB3GAP2 defect and manifesting as a muscular dystrophy-short stature disorder with no ocular anomalies. Functional analyses indicated defective autophagy in patient-derived fibroblasts, supporting the involvement of RAB3GAP2 in the etiology of this disorder. Our results contribute to a better characterization of the Martsolf/micro spectrum phenotype., (© 2022 Wiley Periodicals LLC.)

Background: Guanine nucleotide exchange factors (GEFs) play pivotal roles in neuronal cell functions by exchanging GDP to GTP nucleotide and activation of GTPases. We aimed to determine the genotype and phenotype spectrum of GEF mutations by collecting data from a large Iranian cohort with intellectual disability (ID) and/or developmental delay (DD)., Methods: We collected data from nine families with 20 patients extracted from Iranian cohort of 640 families with ID and/or DD. Next-generation sequencing (NGS) was used to identify the causing variants in recruited families. We also compared our clinical and molecular findings with previously reported patients carrying mutations in these GEF genes in the literature published until mid-2021., Results: We identified disease-causing variants in eight GEF genes including ALS2, IQSEC2, MADD, RAB3GAP1, RAB3GAP2, TRIO, ITSN1, and DENND2A. The major clinical manifestations in 203 previously reported cases along with our 20 patients with disease causing variants in eight GEF genes were as follow; speech disorder (85.2%), ID (81.6%), DD (81.1%), inability to walk (71.3%), facial dysmorphisms features (52.4%), abnormalities in skull morphology (55.6%), hypotonia and muscle weakness (47%), and brain MRI abnormalities (43.4%)., Conclusion: Our study provides new insights into the genotype and phenotype spectrum of mutations in GEF genes., (© 2022 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals LLC.)

Neurotransmitter release is stabilized by homeostatic plasticity. Presynaptic homeostatic potentiation (PHP) operates on timescales ranging from minute- to life-long adaptations and likely involves reorganization of presynaptic active zones (AZs). At Drosophila melanogaster neuromuscular junctions, earlier work ascribed AZ enlargement by incorporating more Bruchpilot (Brp) scaffold protein a role in PHP. We use localization microscopy (direct stochastic optical reconstruction microscopy [dSTORM]) and hierarchical density-based spatial clustering of applications with noise (HDBSCAN) to study AZ plasticity during PHP at the synaptic mesoscale. We find compaction of individual AZs in acute philanthotoxin-induced and chronic genetically induced PHP but unchanged copy numbers of AZ proteins. Compaction even occurs at the level of Brp subclusters, which move toward AZ centers, and in Rab3 interacting molecule (RIM)-binding protein (RBP) subclusters. Furthermore, correlative confocal and dSTORM imaging reveals how AZ compaction in PHP translates into apparent increases in AZ area and Brp protein content, as implied earlier., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021. Published by Elsevier Inc.)

Rab proteins are GTPases involved in all stages of vesicular transport and membrane fusion in mammalian cells. Individual Rab proteins localize to specific cellular organelles and regulate a specific membrane trafficking pathway. Recent studies suggest an important role for Rab proteins in cancer. Rab3 isoforms (Rab3A, Rab3B, Rab3C, and Rab3D) are expressed almost exclusively in neurons and secretory cells. In this review, the role of Rab3 isoforms in a variety of tumor types is discussed. Of the four Rab3 isoforms, Rab3D has been studied most extensively in cancer cells and this isoform appears to play an oncogenic role in breast, colon, esophageal, skin, and brain tumors. Overexpression of Rab3A and Rab3C was observed in gliomas and colon cancers, respectively. Increased expression of the Rab3 isoforms is related to increased proliferation, migration, and invasiveness. Moreover, high Rab3 isoform levels are often associated with decreased survival and advanced pathological stage in clinical samples. Rab3 isoform-dependent activation of the AKT pathway has been observed in several studies. Although the effects of Rab3 isoforms on cancer cell growth and function have been examined in many tumor types, a number of important questions remain. Are the Rab3-positive vesicles in cancer cells actually secretory in nature? If so, are the contents of these vesicles secreted in a regulated or constitutive manner? How does Rab3-regulated secretion affect cellular signaling and tumor growth? Finally, can Rab3 isoforms be therapeutically manipulated in cancer cells? The fact that knockout of a single Rab3 isoform does not affect viability, at least in mouse models, suggests that targeting of these proteins may be a safe and effective treatment strategy for tumor cells expressing any of the Rab3 isoforms., (© 2021 Wiley Periodicals LLC.)

Little is known about the relationship between stimulation of lacrimal gland (LG) tear protein secretion by parasympathetic versus sympathetic nerves, particularly whether the spectrum of tear proteins evoked through each innervation pathway varies. We have previously shown that activity and abundance of cathepsin S (CTSS), a cysteine protease, is greatly increased in tears of Sjögren's syndrome (SS) patients and in tears from the male NOD mouse of autoimmune dacryoadenitis that recapitulates SS-associated dry eye disease. Beyond the increased synthesis of CTSS detected in the diseased NOD mouse LG, increased tear CTSS secretion in NOD mouse tears was recently linked to increased exocytosis from a novel endolysosomal secretory pathway. Here, we have compared secretion and trafficking of CTSS in healthy mouse LG acinar cells stimulated with either the parasympathetic acetylcholine receptor agonist, carbachol (CCh), or the sympathetic α 1 -adrenergic agonist, phenylephrine (PE). In situ secretion studies show that PE significantly increases CTSS activity and protein in tears relative to CCh stimulation by 1.2-fold (***, p = 0.0009) and ∼5-fold (*, p-0.0319), respectively. A similar significant increase in CTSS activity with PE relative to CCh is observed when cultured LGAC are stimulated in vitro. CCh stimulation significantly elevates intracellular [Ca 2+ ], an effect associated with increases in the size of Rab3D-enriched vesicles consistent with compound fusion, and subsequently decreases in their intensity of labeling consistent with their exocytosis. PE stimulation induces a lower [Ca 2+ ] response and has minimal effects on Rab3D-enriched SV diameter or the intensity of Rab3D-enriched SV labeling. LG deficient in Rab3D exhibit a higher sensitivity to PE stimulation, and secrete more CTSS activity. Significant increases in the colocalization of endolysosomal vesicle markers (Lamp1, Lamp2, Rab7) with the subapical actin suggestive of fusion of endolysosomal vesicles at the apical membrane occur both with CCh and PE stimulation, but PE demonstrates increased colocalization. In conclusion, the α 1 -adrenergic agonist, PE, increases CTSS secretion into tears through a pathway independent of the exocytosis of Rab3D-enriched mature SV, possibly representing an alternative endolysosomal secretory pathway., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Mounting studies have demonstrated that RAB3GAP1 expression is modified in brain diseases with multiple neurobiological functions and processes and acts as a potentially significant target. However, the cellular and molecular events arising from RAB3GAP1 dysexpression are still incompletely understood. In this work, underexpression and overexpression of RAB3GAP1 were first induced into cultured mouse cortical neurons by transfection with lentivirus plasmids. Then we globally explored the effects of RAB3GAP1 dysexpression on the proteome of the neurons through the use of isobaric tag for relative and absolute quantitation (iTRAQ)-based quantitative proteomics with bioinformatics. A total of 364 proteins in the RAB3GAP1-underexpression group and 314 proteins in the RAB3GAP1-overexpression group were identified to be differentially expressed. Subsequent bioinformatics analysis indicated that the proteome functional expression profiles induced by RAB3GAP1 underexpression and overexpression were different, suggesting the potential differences in biological processes and cellular effects. Subsequent intergroup cross-comparison revealed some candidate target proteins regulated directly by RAB3GAP1. Further parallel reaction monitoring (PRM) analysis illustrated that Sub1, Ssrp1, and Top1 proteins might serve as new potentially important linkers in the RAB3GAP1-mediated autophagy pathway in the cortical neurons. Collectively, the current proteomics data furnished new valuable insights to better understand the regulatory molecular mechanism of neuronal RAB3GAP1., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Sertoli cells provide protection and nutrition for developing sperm. Each stage of sperm development occurs on the surface of Sertoli cells. MicroRNA (MiR)-125a-5p is involved in male reproduction. The current research aimed to probe the role of miR-125a-5p in Sertoli cell function. Functionally, miR-125a-5p knockdown facilitated Sertoli cell proliferation, while miR-125a-5p overexpression suppressed Sertoli cell proliferation, as evidenced by 5-ethynyl-20-deoxyuridine incorporation assay. Additionally, miR-125a-5p knockdown inhibited Sertoli cell apoptosis, while miR-125a-5p upregulation facilitated Sertoli cell apoptosis, as evidenced by flow cytometry analysis. Computationally, we identified four predicted mRNA targets of miR-125a-5p. Based on the results of luciferase reporter assay, miR-125a-5p was confirmed to bind to the predicted sequence in the Ras-related protein Rab-3D (RAB3D) 3'UTR. Rescue experiments showed that miR-125a-5p suppressed the proliferative ability of TM4 Sertoli cells and facilitated their apoptosis by targeting RAB3D. Finally, our data confirmed that miR-125a-5p and RAB3D modulated activation of the PI3K/AKT pathway. In conclusion, our data showed that miR-125a-5p regulated Sertoli cell proliferation and apoptosis by targeting RAB3D and regulating the PI3K/AKT pathway., (© The Author(s) 2021. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Warburg-Micro syndrome (WARBM) is a rare autosomal recessively inherited neuro-ophthalmologic syndrome. Although WARBM shows genetic heterogeneity, the pathogenic variants in RAB3GAP1 were the most common cause of WARBM. In this study, we aimed to evaluate the detailed clinical and dysmorphic features of seven WARBM1 patients and overview the variant spectrum of RAB3GAP1 in comparison with the literature who were referred due to congenital cataracts. A previously reported homozygous variant (c.2187&#95;2188delGAinsCT) was identified in three of these patients, while the other four had three novel variants (c.251&#95;258delAGAA, c.2606+1G>A, and c.2861&#95;2862dupGC). Congenital cataract and corpus callosum hypo/agenesia are pathognomonic for WARBM, which could be distinguished from other similar syndromes with additional typical dysmorphic facial features. Although there is no known phenotype and genotype correlation in any type of WARBM, RAB3GAP1 gene analysis should be previously requested as the first step of genetic diagnosis in clinically suspicious patients when it is not possible to request a multi-gene panel., (© 2021 Wiley Periodicals LLC.)

Osteosarcoma (OSA), the malignant bone tumor, predominantly affecting children and adolescents, threatens the life and life quality of the patients. An increasing number of studies have indicated the role of long non-coding RNA (lncRNA) dysregulation in cancer biology. Herein, the study was aimed to explore the role of FGD5 antisense RNA 1 (FGD5-AS1), a lncRNA, in OSA. Expression levels of FGD5-AS1, miR-506-3p and RAB3D mRNA were quantified utilizing qRT-PCR. The expression of RAB3D protein was examined employing Western blot. A series of functional experiments including CCK-8 assay, BrdU assay, wound healing assay, Transwell assay were performed for studying the effects of FGD5-AS1 on the malignancy of OSA cell lines 143B and HOS. The binding site between miR-506-3p and FGD5-AS1 was identified and validated by luciferase reporter assay and RNA immunoprecipitation assay. It was demonstrated that the expression of FGD5-AS1 was up-regulated in OSA tissues and cell lines, and its high expression is associated with higher Enneking stage and poorer histological differentiation. Gain-of-function and loss-of-function studies suggested that FGD5-AS1 facilitated OSA cells proliferation and migration. The promoting effects of FGD5-AS1 overexpression on OSA cell proliferation and migration could be counteracted by miR-506-3p. Moreover, FGD5-AS1 competitively adsorbed miR-506-3p to repress its expression so as to up-regulate the expression of RAB3D. These results indicate that FGD5-AS1 is capable of expediting OSA cell proliferation and migration via sponging miR-506-3p to up-regulate RAB3D.

Background: The present study aimed to determine the underlying genetic factors causing the possible Warburg micro syndrome (WARBM) phenotype in two Iranian patients., Case Presentation: A 5-year-old female and a 4.5-year-old male were referred due to microcephaly, global developmental delay, and dysmorphic features. After doing neuroimaging and clinical examinations, due to the heterogeneity of neurodevelopmental disorders, we subjected 7 family members to whole-exome sequencing. Three candidate variants were confirmed by Sanger sequencing and allele frequency of each variant was also determined in 300 healthy ethnically matched people using the tetra-primer amplification refractory mutation system-PCR and PCR-restriction fragment length polymorphism. To show the splicing effects, reverse transcription-PCR (RT-PCR) and RT-qPCR were performed, followed by Sanger sequencing. A novel homozygous variant-NM&#95;012233.2: c.151-5 T > G; p.(Gly51IlefsTer15)-in the RAB3GAP1 gene was identified as the most likely disease-causing variant. RT-PCR/RT-qPCR showed that this variant can activate a cryptic site of splicing in intron 3, changing the splicing and gene expression processes. We also identified some novel manifestations in association with WARBM type 1 to touch upon abnormal philtrum, prominent antitragus, downturned corners of the mouth, malaligned teeth, scrotal hypoplasia, low anterior hairline, hypertrichosis of upper back, spastic diplegia to quadriplegia, and cerebral white matter signal changes., Conclusions: Due to the common phenotypes between WARBMs and Martsolf syndrome (MIM: 212720), we suggest using the "RABopathies" term that can in turn cover a broad range of manifestations. This study can per se increase the genotype-phenotype spectrum of WARBM type 1.

Abstract: Single-cell RNA-seq has become a powerful tool to understand tumor cell heterogenicity. This study tried to screen prognosis-related genes in basal-like breast tumors and evaluate their correlations with cellular states at the single-cell level.Bulk RNA-seq data of basal-like tumor cases from The Cancer Genome Atlas-Breast Cancer (TCGA-BRCA) and single-cell RNA-seq from GSE75688 were retrospectively reviewed. Kaplan-Meier survival curves, univariate and multivariate analysis based on Cox regression model were conducted for survival analysis. Gene set enrichment analysis (GSEA) and single-cell cellular functional state analysis were performed.Twenty thousand five hundred thirty genes with bulk RNA-seq data in TCGA were subjected to screening. Preliminary screening identified 10 candidate progression-related genes, including CDH19, AQP5, SDR16C5, NCAN, TTYH1, XAGE2, RIMS2, GZMB, LY6D, and FAM3B. By checking their profiles using single-cell RNA-seq data, only CDH19, SDR16C5, TTYH1, and RIMS2 had expression in primary triple-negative breast cancer (TNBC) cells. Prognostic analysis only confirmed that RIMS2 expression was an independent prognostic indicator of favorable progression free survival (PFS) (HR: 0.78, 95%: 0.64-0.95, P  = .015). GSEA analysis showed that low RIMS2 group expression had genes significantly enriched in DNA Repair, and MYC Targets V2. Among the 89 basal-like cells, RIMS2 expression was negatively correlated with DNA repair and epithelial-to-mesenchymal transition (EMT).RIMS2 expression was negatively associated with DNA repair capability of basal-like breast tumor cells and might serve as an independent indicator of favorable PFS., Competing Interests: The authors have no conflict of interest to disclose., (Copyright © 2021 the Author(s). Published by Wolters Kluwer Health, Inc.)

Rationale: Warburg Micro syndrome is a rare, autosomal recessive disorder characterized by multiple organ abnormalities involving the ocular, nervous, and genital systems. This case report describes a novel mutation in the RAB3GAP1 gene associated with Warburg Micro syndrome., Patient Concerns: A 6-month-old female infant with bilateral congenital cataracts and developmental delay was referred to our department for further assessment. She presented with facial dysmorphic features, including a prominent forehead, microphthalmia, wide nasal bridge, relatively narrow mouth, large anteverted ears, and micrognathia., Diagnoses: The patient was diagnosed with Warburg Micro syndrome based on clinical manifestations, as well as a novel homozygous mutation in RAB3GAP1: c.75-2A>C. Both parents were identified as heterozygotic carriers of this mutation., Interventions: Bilateral cataract extraction and anterior vitrectomy were performed at age 6 months, followed by physical rehabilitation. Convex lenses were used to protect the eyes postoperatively until intraocular lens implantation., Outcomes: Although the patient received physical rehabilitation, she suffered global developmental delay., Lessons: The c.75-2A>C mutation in RAB3GAP1 expands the spectrum of known mutations in this gene, and it may be associated with Warburg Micro syndrome. Genetic counselors may wish to take this finding into consideration, especially given the poor prognosis associated with the disease., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2021 the Author(s). Published by Wolters Kluwer Health, Inc.)

Warburg micro syndrome (WMS) is a hereditary autosomal neuromuscular disorder in humans caused by mutations in Rab18, Rab3GAP1, or Rab3GAP2 genes. Rab3GAP1/2 forms a heterodimeric complex, which acts as a guanosine nucleotide exchange factor and activates Rab18. Although the genetic causes of WMS are known, it is still unclear whether loss of the Rab3GAP-Rab18 module affects neuronal or muscle cell physiology or both, and how. In this work, we characterize a Rab3GAP2 mutant Drosophila line to establish a novel animal model for WMS. Similarly to symptoms of WMS, loss of Rab3GAP2 leads to highly decreased motility in Drosophila that becomes more serious with age. We demonstrate that these mutant flies are defective for autophagic degradation in multiple tissues including fat cells and muscles. Loss of Rab3GAP-Rab18 module members leads to perturbed autolysosome morphology due to destabilization of Rab7-positive autophagosomal and late endosomal compartments and perturbation of lysosomal biosynthetic transport. Importantly, overexpression of UVRAG or loss of Atg14, two alternative subunits of the Vps34/PI3K (vacuole protein sorting 34/phosphatidylinositol 3-kinase) complexes in fat cells, mimics the autophagic phenotype of Rab3GAP-Rab18 module loss. We find that GTP-bound Rab18 binds to Atg6/Beclin1, a permanent subunit of Vps34 complexes. Finally, we show that Rab3GAP2 and Rab18 are present on autophagosomal and autolysosomal membranes and colocalize with Vps34 Complex I subunits. Our data suggest that the Rab3GAP-Rab18 module regulates autolysosomal maturation through its interaction with the Vps34 Complex I, and perturbed autophagy due to loss of the Rab3GAP-Rab18 module may contribute to the development of WMS., (© 2020 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

The various malformations of the aerodigestive tract collectively known as esophageal atresia/tracheoesophageal fistula (EA/TEF) constitute a rare group of birth defects of largely unknown etiology. Previous studies have identified a small number of rare genetic variants causing syndromes associated with EA/TEF. We performed a pilot exome sequencing study of 45 unrelated simplex trios (probands and parents) with EA/TEF. Thirteen had isolated and 32 had nonisolated EA/TEF; none had a family history of EA/TEF. We identified de novo variants in protein-coding regions, including 19 missense variants predicted to be deleterious (D-mis) and 3 likely gene-disrupting (LGD) variants. Consistent with previous studies of structural birth defects, there is a trend of increased burden of de novo D-mis in cases (1.57-fold increase over the background mutation rate), and the burden is greater in constrained genes (2.55-fold, p = 0.003). There is a frameshift de novo variant in EFTUD2, a known EA/TEF risk gene involved in mRNA splicing. Strikingly, 15 out of 19 de novo D-mis variants are located in genes that are putative target genes of EFTUD2 or SOX2 (another known EA/TEF gene), much greater than expected by chance (3.34-fold, p value = 7.20e-5). We estimated that 33% of patients can be attributed to de novo deleterious variants in known and novel genes. We identified APC2, AMER3, PCDH1, GTF3C1, POLR2B, RAB3GAP2, and ITSN1 as plausible candidate genes in the etiology of EA/TEF. We conclude that further genomic analysis to identify de novo variants will likely identify previously undescribed genetic causes of EA/TEF.

Objective: To explore the clinical and genetic characteristics of a child featuring developmental delay., Methods: The child was subjected to whole exome sequencing. Candidate variant was verified by Sanger sequencing., Results: Whole genome sequencing revealed that the child has carried compound heterozygous variants c.2607-1G>C and c.899 + 2dupT of the RAB3GAP1 gene, which were respectively derived from her mother and father., Conclusion: A rare case of Warburg micro syndrome type 1 was diagnosed. The phenotype of the child was consistent with the literature, in addition with dysplasia of palatine arch, prominent high palatal arch and tooth dysplasia. Above finding has provided a basis for genetic counseling and prenatal diagnosis for the family.

Mixing genomes of different species by hybridization can disrupt species-specific genetic interactions that were adapted and fixed within each species population. Such disruption can predispose the hybrids to abnormalities and disease that decrease the overall fitness of the hybrids and is therefore named as hybrid incompatibility. Interspecies hybridization between southern platyfish and green swordtails leads to lethal melanocyte tumorigenesis. This occurs in hybrids with tumor incidence following progeny ratio that is consistent with two-locus interaction, suggesting melanoma development is a result of negative epistasis. Such observations make Xiphophorus one of the only two vertebrate hybrid incompatibility examples in which interacting genes have been identified. One of the two interacting loci has been characterized as a mutant epidermal growth factor receptor. However, the other locus has not been identified despite over five decades of active research. Here we report the localization of the melanoma regulatory locus to a single gene, rab3d , which shows all expected features of the long-sought oncogene interacting locus. Our findings provide insights into the role of egfr regulation in regard to cancer etiology. Finally, they provide a molecular explainable example of hybrid incompatibility., Competing Interests: The authors declare no competing interest., (Copyright © 2020 the Author(s). Published by PNAS.)

Micro and Martsolf syndromes are rare clinically and genetically overlapping disorders caused by mutations in RAB3GAP1, RAB3GAP2, RAB18 and TBC1D20 genes. We describe 34 new patients, 27 with Micro and seven with Martsolf. Patients presented with the characteristic clinical manifestations of the two syndromes, including postnatal microcephaly, congenital cataracts, microphthalmia, optic atrophy, spasticity and intellectual disability. Brain imaging showed in the majority of cases polymicrogyria, thin corpus callosum, cortical atrophy, and white matter dysmyelination. Unusual additional findings were pectus excavatum (four patients), pectus carinatum (three patients), congenital heart disease (three patients) and bilateral calcification in basal ganglia (one patient). Mutational analysis of RAB3GAP1 and RAB3GAP2 revealed 21 mutations, including 14 novel variants. RAB3GAP1 mutations were identified in 22 patients with Micro, including a deletion of the entire gene in one patient. On the other hand, RAB3GAP2 mutations were identified in two patients with Micro and all Martsolf patients. Moreover, exome sequencing unraveled a TBC1D20 mutation in an additional family with Micro syndrome. Our results expand the phenotypic and mutational spectrum associated with Micro and Martsolf syndromes. Due to the overlapped severities and genetic basis of both syndromes, we suggest to be comprehended as one entity "Micro/Martsolf spectrum" or "RAB18 deficiency.", (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

At presynaptic active zones, arrays of large conserved scaffold proteins mediate fast and temporally precise release of synaptic vesicles (SVs). SV release sites could be identified by clusters of Munc13, which allow SVs to dock in defined nanoscale relation to Ca2+ channels. We here show in Drosophila that RIM-binding protein (RIM-BP) connects release sites physically and functionally to the ELKS family Bruchpilot (BRP)-based scaffold engaged in SV recruitment. The RIM-BP N-terminal domain, while dispensable for SV release site organization, was crucial for proper nanoscale patterning of the BRP scaffold and needed for SV recruitment of SVs under strong stimulation. Structural analysis further showed that the RIM-BP fibronectin domains form a "hinge" in the protein center, while the C-terminal SH3 domain tandem binds RIM, Munc13, and Ca2+ channels release machinery collectively. RIM-BPs' conserved domain architecture seemingly provides a relay to guide SVs from membrane far scaffolds into membrane close release sites., (© 2020 Petzoldt et al.)

Biallelic pathogenic variants in RAB3GAP2 cause Warburg Micro syndrome (WARBM) and Martsolf syndrome (MS), two rare, phenotypically overlapping disorders characterized by congenital cataracts, intellectual disability, and hypogonadism. Although the initial report documented hypergonadotropic hypogonadism (implying a gonadal defect), an adolescent girl with WARBM/MS was subsequently reported to have hypogonadotropic hypogonadism (implying a central defect in either the hypothalamus or anterior pituitary). However, in adult MS, hypogonadotropism has not been convincingly demonstrated. Additionally, the correlation between the pathogenic severity of variants in RAB3GAP2 and the phenotypic severity also remains unclear. Here we present a clinical report of a woman with congenital cataracts, apparent intellectual disability, and pubertal failure who underwent exome sequencing (ES) to determine a precise molecular diagnosis. Reproductive phenotypes reported previously in individuals with MS and the genotypic spectrum of previous RAB3GAP2 variants were also reviewed. The ES identified pathogenic compound heterozygous RAB3GAP2 variants (c.387-2A > G; p.(Arg428Glu)) combined with her phenotypic features, which enabled a unifying molecular diagnosis of MS. Reproductive evaluation confirmed a normosmic idiopathic hypogonadotropic hypogonadism. Review of the RAB3GAP2 allelic spectrum in WARBM/MS suggests that although variants resulting in complete abrogation of RAB3GAP2 protein function cause severe WARBM, variants associated with partially preserved RAB3GAP2 function cause milder MS. This report expands the genotypic and phenotypic spectrum of MS and demonstrates hypogonadotropic hypogonadism as a key pathophysiologic abnormality in MS. Genotype-phenotype associations of previously reported RAB3GAP2 variants indicate that variants that fully abolish RAB3GAP2 function result in WARBM, whereas MS is associated with variants of lesser severity with residual RAB3GAP2 function., (© 2020 Xu et al.; Published by Cold Spring Harbor Laboratory Press.)

Rod photoreceptor synapses use large, ribbon-type active zones for continuous synaptic transmission during light and dark. Since ribbons are physically connected to the active zones, we asked whether illumination-dependent changes of ribbons influence Cav1.4/RIM2 protein clusters at the active zone and whether these illumination-dependent effects at the active zone require the presence of the synaptic ribbon. We found that synaptic ribbon length and the length of presynaptic Cav1.4/RIM2 clusters are tightly correlated. Dark-adaptation did not change the number of ribbons and active zone puncta. However, mean ribbon length and length of presynaptic Cav1.4/RIM2 clusters increased significantly during dark-adaptation when tonic exocytosis is highest. In the present study, we identified by the analyses of synaptic ribbon-deficient RIBEYE knockout mice that synaptic ribbons are (1) needed to stabilize Cav1.4/RIM2 at rod photoreceptor active zones and (2) are required for the darkness-induced active zone enrichment of Cav1.4/RIM2. These data propose a role of the ribbon in active zone stabilization and suggest a homeostatic function of the ribbon in illumination-dependent active zone remodeling.

Autophagy is dependent on appropriate lipid supply for autophagosome formation. The regulation of lipid acquisition and the autophagy network response to lipid-limiting conditions are mostly elusive. Here, we show that the knockout of the RAB GTPase RAB18 interferes with lipid droplet catabolism, causing an impaired fatty acid release. The resulting reduced lipid-droplet-derived lipid availability influences autophagy and provokes adaptive modifications of the autophagy network. These adjustments include increased expression and phosphorylation of ATG2B as well as augmented formation of the ATG12-ATG5 conjugate. Moreover, ATG9A shows an enhanced phosphorylation at amino acid residues tyrosine 8 and serine 14, resulting in an increased ATG9A trafficking. Via pharmacological inhibition of Y8 phosphorylation, we demonstrate that this ATG9A modification is important to maintain basal autophagy under RAB18 knockout conditions. However, while the network adaptations are sufficient to maintain basal autophagic activity, they are incapable of ensuring autophagy induction upon starvation, which is characterized by an enhanced lipid demand. Thus, here, we define the molecular role of RAB18 in connecting lipid droplets and autophagy, emphasize the significance of lipid droplets as lipid sources for the degradative pathway, and uncover a remarkable autophagy network plasticity, including phosphorylation-dependent ATG9A activation, to compensate reduced lipid availability in order to rescue basal autophagic activity., (Copyright © 2019 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Renal cell carcinoma (RCC) is the most common type of kidney cancer with rising incidence. Long noncoding RNA (lncRNA) LINC01133 is a novel lncRNA that is involved in the development of several types of cancers. However, the role of LINC01133 in RCC has not been reported. Thus, in this study, we investigated the functions of LINC01133 in RCC. The qualitative real-time polymerase chain reaction analysis was performed to examine the levels of LINC01133 in RCC tissues and adjacent tissues, as well as RCC cell lines. The results showed that LINC01133 was highly expressed in RCC tissue specimens and cell lines. Downregulation of LINC01133 significantly inhibited the proliferation, migration, and invasion of RCC cells. Further mechanistic investigations proved that LINC01133 directly interacted with microRNA (miR)-30b-5p and regulated the miR-30b-5p expression in RCC cell lines. Moreover, miR-30b-5p exhibited tumor-suppressive activity in RCC cell lines, which was mediated by targeting Ras-related protein Rab-3D (Rab3D). In vivo study showed that LINC01133 knockdown suppressed tumor growth in the nude mice. Taken together, these findings indicated that LINC01133 might be an oncogene in RCC through regulation of the miR-30b-5p/Rab3D axis. Thus, LINC01133 might serve as a potential therapeutic target for the treatment of RCC.

Warburg micro syndrome (WARBM) is a rare autosomal recessive disorder characterized by microcephaly, cortical dysplasia, intellectual disability, ocular abnormalities, spastic diplegia, and microgenitalia. WARBM has 4 subtypes arising from pathogenic variants in 4 genes (RAB18, RAB3GAP1, RAB3GAP2, and TBC1D20). Here, we report on a patient with a homozygous pathogenic c.665delC (p.Pro222HisfsTer30) variant in the RAB3GAP1 gene identified by whole-exome sequencing (WES) analyses. Only his father was a heterozygous carrier, and homozygosity mapping analysis of the WES data revealed large loss-of-heterozygosity regions in both arms of chromosome 2, interpreted as uniparental isodisomy. This uniparental disomy pattern could be due to paternal meiosis I nondisjunction because of the preserved heterozygosity in the pericentromeric region. This report provides novel insights, including a rare form of UPD, usage of homozygosity mapping analysis for the evaluation of isodisomy, and the first reported case of WARBM1 as a result of uniparental isodisomy., (© 2020 S. Karger AG, Basel.)

Vici syndrome is a severe and progressive multisystem disease caused by mutations in the EPG5 gene. In patient tissues and animal models, loss of EPG5 function is associated with defective autophagy caused by accumulation of non-degradative autolysosomes, but very little is known about the mechanism underlying this cellular phenotype. Here, we demonstrate that loss of function of the RBG-1-RBG-2 complex ameliorates the autophagy defect in C. elegans epg-5 mutants. The suppression effect is independent of the complex's activity as a RAB-3 GAP and a RAB-18 GEF. Loss of rbg-1 activity promotes lysosomal biogenesis and function, and also suppresses the accumulation of non-functional autolysosomes in epg-5 mutants. The mobility of late endosome- and lysosome-associated RAB-7 is reduced in epg-5 mutants, and this defect is rescued by simultaneous loss of function of rbg-1 Expression of the GDP-bound form of RAB-7 also promotes lysosomal biogenesis and suppresses the autophagy defect in epg-5 mutants. Our study reveals that the RBG-1-RBG-2 complex acts by modulating the dynamics of membrane-associated RAB-7 to regulate lysosomal biogenesis, and provides insights into the pathogenesis of Vici syndrome., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)

Estrogen increases bone formation by promoting mineralization and prolonging the lifespan of osteoblasts. To understand the underlying molecular mechanism/s, we identified estrogen-regulated proteins at different stages of human osteoblast differentiation using differential proteomics approach. Among the identified proteins, we observed that estrogen upregulated RAB3GAP1 on day 1 and 5 of differentiation. RAB3GAP1 is critically involved in the process of autophagy, a eukaryotic degradative pathway essential for cell survival. We, therefore, investigated the effect of estrogen on autophagy in differentiating human osteoblasts and their precursors, the mesenchymal stem cells (MSCs). MSCs exhibited high autophagic flux which declined during osteoblast differentiation, resulting in high basal apoptosis in osteoblasts. Estrogen reduced apoptosis in differentiating osteoblasts by promoting autophagy, thus contributing towards their longer lifespan. Further, MSCs were resistant against starvation-induced apoptosis, whereas, differentiating osteoblasts showed significant susceptibility towards it. Estrogen, in addition to promoting mineralization, protected differentiating osteoblasts from starvation-induced apoptosis by increasing autophagic flux. Autophagic flux in RAB3GAP1 knockdown osteoblasts appeared diminished, and showed increased apoptosis even in nutrient-rich conditions, and exhibited significantly impaired mineralization. However, irrespective of the presence of estrogen, starvation further enhanced apoptosis in these cells. Furthermore, estrogen failed to promote mineralization in these osteoblasts. Our study illustrates that autophagy is essential for human osteoblast survival and mineralization, and osteoblasts are susceptible to apoptosis due to reduced autophagy during differentiation. Estrogen, via upregulation of RAB3GAP1, promotes autophagy in osteoblasts during differentiation thereby increasing their survival and mineralization capacity. Our study demonstrates the positive role of autophagy in bone homeostasis., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: Macroautophagy (or autophagy) is a conserved degradative pathway that breaks down sequestered cytoplasmic proteins and organelles in specialized double-membrane compartments called autophagosomes that fuse with lysosomes. Several proteins orchestrate this process, specifically Rab GTPases that are master regulators of molecular trafficking. RAB18 GTPase, a known mediator of stellate cell activation, is known to modulate autophagic flux in fibroblasts. However, its role in autophagy is unexplored in hepatic stellate cells., Objective: The aim of this study was to investigate the role of RAB18 in modulating autophagy in hepatic stellate cells., Methods: Role of RAB18 was determined by genetic depletion, pharmacologic inhibition, and overexpression studies to monitor autophagy flux and proteostasis in human LX2 stellate cell line., Results: RAB18 knockdown increases autophagy flux and regulates proteostasis. LX2 cells stimulated with transforming growth factor-beta robustly increases expression of profibrotic genes such as COL1A1 and ACTA2 along with RAB18 and its guanine nucleotide exchange factor, RAB3GAP1., Conclusion: The study elucidates a role for RAB18 in autophagy and regulation of proteostasis in human stellate cells. Molecular insights into this process can provide therapeutic opportunities for intervention in liver fibrosis., (Copyright © 2019 National Lipid Association. Published by Elsevier Inc. All rights reserved.)

In many species, sperm can remain viable in the reproductive tract of a female well beyond the typical interval to remating. This creates an opportunity for sperm from different males to compete for oocyte fertilization inside the female's reproductive tract. In Drosophila melanogaster , sperm characteristics and seminal fluid content affect male success in sperm competition. On the other hand, although genome-wide association studies (GWAS) have demonstrated that female genotype plays a role in sperm competition outcome as well, the biochemical, sensory, and physiological processes by which females detect and selectively use sperm from different males remain elusive. Here, we functionally tested 26 candidate genes implicated via a GWAS for their contribution to the female's role in sperm competition, measured as changes in the relative success of the first male to mate (P1). Of these 26 candidates, we identified eight genes that affect P1 when knocked down in females, and showed that five of them do so when knocked down in the female nervous system. In particular, Rim knockdown in sensory pickpocket ( ppk) + neurons lowered P1, confirming previously published results, and a novel candidate, caup , lowered P1 when knocked down in octopaminergic Tdc2 + neurons. These results demonstrate that specific neurons in the female's nervous system play a functional role in sperm competition and expand our understanding of the genetic, neuronal, and mechanistic basis of female responses to multiple matings. We propose that these neurons in females are used to sense, and integrate, signals from courtship or ejaculates, to modulate sperm competition outcome accordingly., (Copyright © 2019 by the Genetics Society of America.)

Synaptic terminals grow and retract throughout life, yet synaptic strength is maintained within stable physiological ranges. To study this process, we investigated Drosophila endophilin ( endo ) mutants. Although active zone (AZ) number is doubled in endo mutants, a compensatory reduction in their size homeostatically adjusts global neurotransmitter output to maintain synaptic strength. We find an inverse adaptation in rab3 mutants. Additional analyses using confocal, STED, and electron microscopy reveal a stoichiometric tuning of AZ scaffolds and nanoarchitecture. Axonal transport of synaptic cargo via the lysosomal kinesin adapter Arl8 regulates AZ abundance to modulate global synaptic output and sustain the homeostatic potentiation of neurotransmission. Finally, we find that this AZ scaling can interface with two independent homeostats, depression and potentiation, to remodel AZ structure and function, demonstrating a robust balancing of separate homeostatic adaptations. Thus, AZs are pliable substrates with elastic and modular nanostructures that can be dynamically sculpted to stabilize and tune both local and global synaptic strength., (© 2019 Goel et al.)

Purpose: Martsolf (MS) and Warburg micro syndromes (WARBM) are rare autosomal recessive inherited allelic disorders, which share similar clinical features including microcephaly, intellectual disability, brain malformations, ocular abnormalities, and spasticity. Here, we revealed the functions of novel mutations in RAB3GAP1 in a Turkish female patient with MS and two siblings with WARBM. We also present a review of MS patients as well as all reported RAB3GAP1 pathogenic mutations in the literature., Methods: We present a female with MS phenotype and two siblings with WARBM having more severe phenotypes. We utilized whole-exome sequencing to identify the molecular basis of these syndromes and confirmed suspected variants by Sanger sequencing. Quantitative (q) RT-PCR analysis was carried out to reveal the functions of novel splice site mutation detected in MS patient., Results: We found a novel homozygous c.2607-1G>C splice site mutation in intron 22 of RAB3GAP1 in MS patient and a novel homozygous c.2187&#95;2188delinsCT, p.(Met729&#95;Lys730delinsIleTer) mutation in exon 19 of RAB3GAP1 in the WARBM patients. We showed exon skipping in MS patient by Sanger sequencing and gel electrophoresis. qRT-PCR analysis demonstrated the reduced expression of RAB3GAP1 in the patient with the c.2607-1G>C splice site mutation compared to a healthy control individual., Conclusion: Here, we have studied two novel RAB3GAP1 mutations in two different phenotypes; a MS associated novel splice site mutation, and a WARBM1 associated novel deletion-insertion mutation. Our findings suggest that this splice site mutation is responsible for milder phenotype and the deletion-insertion mutation presented here is associated with severe phenotype., (© 2019 Wiley Periodicals, Inc.)

In order to explore the differentially expressed long non-coding RNAs (lncRNAs) in laryngeal squamous cell carcinoma (LSCC), the GSE84957 lncRNA expression profile was included in the present study through data mining in the National Center for Biotechnology Information/Gene Expression Omnibus (NCBI/GEO). Then, the differentially expressed genes (DEGs) of LSCC (1646 lncRNAs and 2713 mRNAs, fold change ≥ 2, P ≤ 0.05) were identified from the GSE84957 dataset using bioinformatics analysis. Of the 10 selected differentially expressed lncRNAs, the expression of 7 lncRNAs were verified by qRT-PCR method. Then, LINC00668, a potential carcinogenic lncRNA, was screened out by narrowing down the screening criteria (fold change ≥ 4, P ≤ 0.01). Furthermore, correlation analysis demonstrated that expression levels of LINC00668 were associated with age, pathological differentiation degree, T stage, clinical stage and cervical lymph node metastasis. Moreover, a series of bioinformatics tools and in vitro experiments proved that knockdown of LINC00668 inhibited the proliferation, migration and invasion ability of LSCC cells. The present study identified the lncRNAs landscape of LSCC through data mining and bioinformatics analysis, and verified oncogenic LINC00668, which may play important roles in promoting LSCC cells proliferation, migration and invasion., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Background: Warburg micro syndrome is a very rare autosomal recessive disorder characterized by a mutation in the RAB3GAP1, RAB3GAP2, RAB18, and TBC1D20 genes. Warburg Micro syndrome 2 and Martsolf syndrome are clinically overlapping conditions characterized by variable clinical signs counting postnatal growth retardation, cataract, intellectual deficiency, contractures, and central nervous system abnormalities due to RAB3GAP2 gene mutations. The RAB3GAP2 gene encodes a member of the Rab3 protein family, which is involved in regulated exocytosis of neurotransmitters and hormones., Case Presentation: We describe four siblings from healthy consanguineous Turkish parents with developmental delay, congenital cataract, and speech delay. In this study, we performed whole exom sequencing (WES) in a index patient. WES analyses in proposita showed a homozygous c.1998 + 1 G > A mutation in RAB3GAP2 gene. After the Sanger confirmation, the same mutation was detected in the other three siblings., Conclusion: The four siblings had a novel splice site mutation in RAB3GAP2. This report compares the symptoms and features of the our patients with clinical summary of Warburg Micro syndrome 2 and Martsolf syndrome. Further reports will make possible knowing of the genetic and clinical backgrounds of this orphan diseases. Abbreviation: MRI: Magnetic resonance imaging.

Neuron subtypes of the mature nervous system differ in the expression of characteristic marker genes while they share the expression of generic neuronal genes. The regulatory logic that maintains subtype-specific and pan-neuronal genes is not well understood. To begin to address this issue, we analyze RNA sequencing results from whole sympathetic ganglia and single sympathetic neurons in the mouse. We focus on gene products involved in the neuronal cytoskeleton, neurotransmitter synthesis and storage, transmitter release and reception and electrical information processing. We find a particular high correlation in the expression of stathmin 2 and several members of the tubulin beta family, classical pan-neuronal markers. Noradrenergic transmitter-synthesizing enzymes and transporters are also well correlated in their cellular transcript levels. In addition, noradrenergic marker transcript levels correlate well with selected pan-neuronal markers. Such a correlation in transcript levels is also seen between a number of selected ion channel, receptor and synaptic protein genes. These results provide the foundation for the analyses of the coordinated expression of downstream target genes in nerve cells.

Contextual memory is an intricate process involving synaptic plasticity and network rearrangement. Both are governed by many molecular processes including phosphorylation and modulation of protein expression. However, little is known about the molecules involved in it. Here, we exploited the advantages of a quantitative proteomic approach to identify a great number of molecules in the rat dentate gyrus after a contextual fear conditioning session. Our results allowed us to highlight protein expression patterns, not only related to neuroplasticity, but also to myelin structure, such as myelin basic protein and myelin proteolipid protein showing a decrease in expression. Validation of the modification in protein expression reveals a dynamic profile during the 48 h following the fear conditioning session. The expression of proteins involved in neurite outgrowth, such as BASP-1 and calcineurin B1, and in synaptic structure and function, VAMP2 and RAB3C, was increased in the dentate gyrus of rats submitted to fear conditioning compared to controls. We showed that the increase in BASP-1 protein was specific to fear conditioning learning as it was not present in immediate-shock rats, neither in rats exposed to a novel environment without being shocked. As myelin is known to stabilise synaptic network, the decrease in myelin proteins suggests a neuroglia interactive process taking place in the dentate gyrus in the 24 h following contextual fear learning, which has never been demonstrated before. These results therefore open the way to the study of new plasticity mechanisms underlying learning and memory., (© 2017 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Rab3D is a member of the ras-related GTP-binding protein Rab family and was found up-regulated in several types of cancer. However, little is known about the role of Rab3D in carcinogenesis and progression of esophageal squamous cell carcinoma (ESCC). Thus, in this study, we investigated the expression patterns and functional roles of Rab3D in human ESCC. We demonstrated that Rab3D was highly expressed in human ESCC cell lines. In addition, knockdown of Rab3D significantly inhibited the proliferation of ESCC cells and reduced the tumorigenesis in vivo. Moreover, knockdown of Rab3D significantly suppressed ESCC cell migration/invasion and accordingly alerted EMT related markers, which including up-regulated E-cadherin and down-regulated N-cadherin in ESCC cells. Finally, knockdown of Rab3D inhibited the levels of p-PI3K and p-Akt in ECA-109 cells. In conclusion, our data demonstrated that Rab3D functions as an oncogene in ESCC and knockdown of Rab3D suppressed ESCC cell proliferation and invasion, potentially through the PI3K/Akt signaling pathway. Overall, these findings suggest that targeting the Rab3D may be a potential therapeutic target for treatment of ESCC., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)