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Purpose: Radiation therapy is a vital adjuvant treatment for liver cancer, although the challenge of radiation-induced liver diseases (RILDs) limits its implementation. Kupffer cells (KCs) are a crucial cell population of the hepatic immune system, and their biologic function can be modulated by multiple epigenetic RNA modifications, including N 6 -methyladenosine (m 6 A) methylation. However, the mechanism for m 6 A methylation in KC-induced inflammatory responses in RILD remains unclear. The present study investigated the function of m 6 A modification in KCs contributing to RILD., Methods and Materials: Methylated RNA-immunoprecipitation sequencing and RNA transcriptome sequencing were used to explore the m 6 A methylation profile of primary KCs isolated from mice after irradiation with 3 × 8 Gy. Western blotting and quantitative real-time PCR were used to evaluate gene expression. DNA pulldown and chromatin immunoprecipitation assays were performed to verify target gene binding and identify binding sites., Results: Methylated RNA-immunoprecipitation sequencing revealed significantly increased m 6 A modification levels in human KCs after irradiation, suggesting the potential role of upregulated m 6 A in RILD. In addition, the study results corroborated that methyltransferase-like 3 (METTL3) acts as a main modulator to promote the methylation and gene expression of TEAD1, leading to STING-NLRP3 signaling activation. Importantly, it was shown that IGF2BP2 functions as an m 6 A "reader" to recognize methylated TEAD1 mRNA and promote its stability. METTL3/TEAD1 knockdown abolished the activation of STING-NLRP3 signaling, protected against RILD, and suppressed inflammatory cytokines and hepatocyte apoptosis. Moreover, clinical human normal liver tissue samples collected after irradiation showed increased expression of STING and interleukin-1β in KCs compared with nonirradiated samples. Notably, STING pharmacologic inhibition alleviated irradiation-induced liver injury in mice, indicating its potential therapeutic role in RILD., Conclusions: The results of our study reveal that TEAD1-STING-NLRP3 signaling activation contributes to RILD via METTL3-dependent m 6 A modification., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Complex behaviors depend on the precise developmental specification of neuronal circuits, but the relationship between genetic programs for neural development, circuit structure, and behavioral output is often unclear. The central complex (CX) is a conserved sensory-motor integration center in insects, which governs many higher-order behaviors and largely derives from a small number of type II neural stem cells (NSCs). Here, we show that Imp, a conserved IGF-II mRNA-binding protein expressed in type II NSCs, plays a role in specifying essential components of CX olfactory navigation circuitry. We show the following: (1) that multiple components of olfactory navigation circuitry arise from type II NSCs. (2) Manipulating Imp expression in type II NSCs alters the number and morphology of many of these circuit elements, with the most potent effects on neurons targeting the ventral layers of the fan-shaped body (FB). (3) Imp regulates the specification of Tachykinin-expressing ventral FB input neurons. (4) Imp is required in type II NSCs for establishing proper morphology of the CX neuropil structures. (5) Loss of Imp in type II NSCs abolishes upwind orientation to attractive odor while leaving locomotion and odor-evoked regulation of movement intact. Taken together, our findings establish that a temporally expressed gene can regulate the expression of a complex behavior by developmentally regulating the specification of multiple circuit components and provides a first step toward a developmental dissection of the CX and its roles in behavior., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Hepatocellular carcinoma (HCC) is one of the most common tumors worldwide, with high incidence and mortality rate. There is an urgent need to identify effective diagnostic and prognostic biomarkers for HCC. Members of the acidic leucine-rich nucleophosphoprotein 32 (ANP32) family, which mainly includes ANP32A , ANP32B , and ANP32E , are abnormally expressed and have prognostic value in certain cancers. However, the diagnostic, prognostic, and therapeutic value of ANP32 family members in HCC has not yet been fully studied. In this study, we identified the diagnostic and prognostic value of ANP32 family members in HCC. Transcriptome data from public databases, such as the Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx) databases, suggested that ANP32A , ANP32B , and ANP32E were upregulated in HCC tissues, and high expression of ANP32 family members was associated with advanced pathologic stage and histologic grade. Our immunohistochemistry and western blot results further verified the differential expression of ANP32 family members. ANP32A , ANP32B , and ANP32E had an outstanding diagnostic potential. Survival analysis of HCC patients in TCGA databases demonstrated that ANP32A , ANP32B , and ANP32E were associated with poor overall survival (OS) and disease-specific survival (DSS). Univariate and multivariate Cox analyses suggested the capability of ANP32B and ANP32E to independently predict the OS and DSS of HCC patients. Gene set enrichment analysis (GSEA) showed that ANP32 family members were associated with immune response, epidermal cell differentiation, and stem cell proliferation. Expression of ANP32 family members was associated with immune cell infiltration and immune status in the tumor microenvironment of HCC, and patients with high ANP32 family expression had poor sensitivity to immunotherapy. Finally, we identified potential chemotherapy drugs for HCC patients with high ANP32 family expression by CellMiner database. This study suggested the diagnostic, prognostic, and therapeutic roles of the ANP32 family in HCC patients, providing potential therapeutic targets for HCC., Competing Interests: The authors have declared that no competing interests exist., (Copyright © 2022 Xuxu Liu et al.)

Introduction: Triggering receptor expressed on myeloid cells-1 (TREM-1) has important implications in sepsis and inflammation and is a novel receptor for extracellular cold-inducible RNA-binding protein (eCIRP). We hypothesize that the inhibition of TREM-1 via its interaction with eCIRP by novel peptide inhibitor M3 or knockout gene will attenuate the inflammation and injury associated with severe hepatic ischemia/reperfusion (I/R)., Methods: Wild-type (WT) C57BL/6 and TREM-1-/- mice underwent 60 min of 70% hepatic ischemia, with 24 h of reperfusion. Additionally, WT mice underwent hepatic I/R and were treated with M3 (10 mg/kg body weight) or vehicle (normal saline) at the start of reperfusion. Blood and ischemic liver tissues were collected, and analysis was performed using enzymatic assays, enzyme-linked immunosorbent assay, reverse-transcription quantitative polymerase chain reaction, and pathohistology techniques. For survival surgery, mice additionally underwent resection of non-ischemic lobes of the liver and survival was monitored for 10 days., Results: There was an increase in serum levels of tissue markers including aspartate aminotransferase, alanine aminotransferase, and lactate dehydrogenase as well as cytokine levels (IL-6) and histological scoring of hematoxylin and eosin sections in WT I/R mice. These markers decreased substantially in TREM-1-/- mice. Additionally, neutrophil infiltration markers and markers of local inflammation (myeloperoxidase, macrophage inflammatory protein-2, cyclooxygenase-2) were attenuated in TREM-1-/- mice. Similarly, we show a significant decrease in injury and inflammation markers with M3 treatment. Additionally, we demonstrate decreased apoptosis with TREM-1 inhibition. Finally, M3 treatment improved the survival rate from 42% to 75% after hepatic I/R., Conclusion: TREM-1 is an important eCIRP receptor in the inflammatory response of hepatic I/R, and deficiency of TREM-1 via knockout gene or peptide inhibition attenuated liver injury and inflammation, and improved survival. Inhibition of the TREM-1 and eCIRP interaction in hepatic I/R may have important therapeutic potential., Competing Interests: The authors report no conflicts of interest., (Copyright © 2021 by the Shock Society.)

The eukaryotic translation initiation factor 5A (eIF5A) is an evolutionarily conserved protein that binds ribosomes to facilitate the translation of peptide motifs with consecutive prolines or combinations of prolines with glycine and charged amino acids. It has also been linked to other molecular functions and cellular processes, such as nuclear mRNA export and mRNA decay, proliferation, differentiation, autophagy, and apoptosis. The growing interest in eIF5A relates to its association with the pathogenesis of several diseases, including cancer, viral infection, and diabetes. It has also been proposed as an anti-aging factor: its levels decay in aged cells, whereas increasing levels of active eIF5A result in the rejuvenation of the immune and vascular systems and improved brain cognition. Recent data have linked the role of eIF5A in some pathologies with its function in maintaining healthy mitochondria. The eukaryotic translation initiation factor 5A is upregulated under respiratory metabolism and its deficiency reduces oxygen consumption, ATP production, and the levels of several mitochondrial metabolic enzymes, as well as altering mitochondria dynamics. However, although all the accumulated data strongly link eIF5A to mitochondrial function, the precise molecular role and mechanisms involved are still unknown. In this review, we discuss the findings linking eIF5A and mitochondria, speculate about its role in regulating mitochondrial homeostasis, and highlight its potential as a target in diseases related to energy metabolism.

Oncogene-induced senescence (OIS) is a form of stable cell-cycle arrest arising in response to oncogenic stimulation. OIS must be bypassed for transformation, but the mechanisms of OIS establishment and bypass remain poorly understood, especially at the post-transcriptional level. Here, we show that the RNA-binding protein UNR/CSDE1 enables OIS in primary mouse keratinocytes. Depletion of CSDE1 leads to senescence bypass, cell immortalization, and tumor formation, indicating that CSDE1 behaves as a tumor suppressor. Unbiased high-throughput analyses uncovered that CSDE1 promotes OIS by two independent molecular mechanisms: enhancement of the stability of senescence-associated secretory phenotype (SASP) factor mRNAs and repression of Ybx1 mRNA translation. Importantly, depletion of YBX1 from immortal keratinocytes rescues senescence and uncouples proliferation arrest from the SASP, revealing multilayered mechanisms exerted by CSDE1 to coordinate senescence. Our data highlight the relevance of post-transcriptional control in the regulation of senescence., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Despite recent advances in therapeutic approaches, patients with MLL-rearranged leukemia still have poor outcomes. Here, we find that the RNA-binding protein IGF2BP3, which is overexpressed in MLL-translocated leukemia, strongly amplifies MLL-Af4-mediated leukemogenesis. Deletion of Igf2bp3 significantly increases the survival of mice with MLL-Af4-driven leukemia and greatly attenuates disease, with a minimal impact on baseline hematopoiesis. At the cellular level, MLL-Af4 leukemia-initiating cells require Igf2bp3 for their function in leukemogenesis. At the molecular level, IGF2BP3 regulates a complex posttranscriptional operon governing leukemia cell survival and proliferation. IGF2BP3-targeted mRNA transcripts include important MLL-Af4-induced genes, such as those in the Hoxa locus, and the Ras signaling pathway. Targeting of transcripts by IGF2BP3 regulates both steady-state mRNA levels and, unexpectedly, pre-mRNA splicing. Together, our findings show that IGF2BP3 represents an attractive therapeutic target in this disease, providing important insights into mechanisms of posttranscriptional regulation in leukemia., (© 2021. The Author(s).)

Development of the ocular lens - a transparent tissue capable of sustaining frequent shape changes for optimal focusing power - pushes the boundaries of what cells can achieve using the molecular toolkit encoded by their genomes. The mammalian lens contains broadly two types of cells, the anteriorly located monolayer of epithelial cells which, at the equatorial region of the lens, initiate differentiation into fiber cells that contribute to the bulk of the tissue. This differentiation program involves massive upregulation of select fiber cell-expressed RNAs and their subsequent translation into high amounts of proteins, such as crystallins. But intriguingly, fiber cells achieve this while also simultaneously undergoing significant morphological changes such as elongation - involving about 1000-fold length-wise increase - and migration, which requires modulation of cytoskeletal and cell adhesion factors. Adding further to the challenges, these molecular and cellular events have to be coordinated as fiber cells progress toward loss of their nuclei and organelles, which irreversibly compromises their potential for harnessing genetically hardwired information. A long-standing question is how processes downstream of signaling and transcription, which may also participate in feedback regulation, contribute toward orchestrating these cellular differentiation events in the lens. It is now becoming clear from findings over the past decade that post-transcriptional gene expression regulatory mechanisms are critical in controlling cellular proteomes and coordinating key processes in lens development and fiber cell differentiation. Indeed, RNA-binding proteins (RBPs) such as Caprin2, Celf1, Rbm24 and Tdrd7 have now been described in mediating post-transcriptional control over key factors (e.g. Actn2, Cdkn1a (p21 Cip1 ), Cdkn1b (p27 Kip1 ), various crystallins, Dnase2b, Hspb1, Pax6, Prox1, Sox2) that are variously involved in cell cycle, transcription, cytoskeleton maintenance and differentiation in the lens. Furthermore, deficiencies of these RBPs have been shown to result in various eye and lens defects and/or cataract. Because fiber cell differentiation in the lens occurs throughout life, the underlying regulatory mechanisms operational in development are expected to also be recruited for the maintenance of transparency in aged lenses. Indeed, in support of this, TDRD7 and CAPRIN2 loci have been linked to age-related cataract in humans. Here, I will review the role of key RBPs in the lens and their importance in understanding the pathology of lens defects. I will discuss advances in RBP-based gene expression control, in general, and the important challenges that need to be addressed in the lens to define the mechanisms that determine the epithelial and fiber cell proteome. Finally, I will also discuss in detail several key future directions including the application of bioinformatics approaches such as iSyTE to study RBP-based post-transcriptional gene expression control in the aging lens and in the context of age-related cataract., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Chromatin is a highly dynamic biological entity that allows for both the control of gene expression and the stabilization of chromosomal domains. Given the high degree of plasticity observed in model and non-model organisms, it is not surprising that new chromatin components are frequently described. In this work, we tested the hypothesis that the remnants of the Doc5 transposable element, which retains a heterochromatin insertion pattern in the melanogaster species complex, can be bound by chromatin proteins, and thus be involved in the organization of heterochromatic domains. Using the Yeast One Hybrid approach, we found Rpl22 as a potential interacting protein of Doc5 . We further tested in vitro the observed interaction through Electrophoretic Mobility Shift Assay, uncovering that the N- terminal portion of the protein is sufficient to interact with Doc5 . However, in situ localization of the native protein failed to detect Rpl22 association with chromatin. The results obtained are discussed in the light of the current knowledge on the extra-ribosomal role of ribosomal protein in eukaryotes, which suggests a possible role of Rpl22 in the determination of the heterochromatin in Drosophila .

Carbon Storage Regulator A (CsrA) is a well-characterized post-transcriptional global regulator that plays a critical role in response to environmental changes in many bacteria. CsrA has been reported to regulate several metabolic pathways, motility, biofilm formation, and virulence-associated genes. The role of csrA in Leptospira spp., which are able to survive in different environmental niches and infect a wide variety of reservoir hosts, has not been characterized. To investigate the role of csrA as a gene regulator in Leptospira, we generated a L. biflexa csrA deletion mutant (ΔcsrA) and csrA overexpressing Leptospira strains. The ΔcsrA L. biflexa displayed poor growth under starvation conditions. RNA sequencing revealed that in rich medium only a few genes, including the gene encoding the flagellar filament protein FlaB3, were differentially expressed in the ΔcsrA mutant. In contrast, 575 transcripts were differentially expressed when csrA was overexpressed in L. biflexa. Electrophoretic mobility shift assay (EMSA) confirmed the RNA-seq data in the ΔcsrA mutant, showing direct binding of recombinant CsrA to flaB3 mRNA. In the pathogen L. interrogans, we were not able to generate a csrA mutant. We therefore decided to overexpress csrA in L. interrogans. In contrast to the overexpressing strain of L. biflexa, the overexpressing L. interrogans strain had poor motility on soft agar. The overexpressing strain of L. interrogans also showed significant upregulation of the flagellin flaB1, flaB2, and flaB4. The interaction of L. interrogans rCsrA and flaB4 was confirmed by EMSA. Our results demonstrated that CsrA may function as a global regulator in Leptospira spp. under certain conditions that cause csrA overexpression. Interestingly, the mechanisms of action and gene targets of CsrA may be different between non-pathogenic and pathogenic Leptospira strains., Competing Interests: The authors have declared that no competing interests exist.

Sam68 is an RNA-binding protein with an adaptor role in signal transduction. Our previous work identified critical proinflammatory and apoptotic functions for Sam68, downstream of the TNF/TNFR1 and TLR2/3/4 pathways. Recent studies have shown elevated Sam68 in inflamed tissues from rheumatoid arthritis and ulcerative colitis (UC) patients, suggesting that Sam68 contributes to chronic inflammatory diseases. Here, we hypothesized that deletion of Sam68 is protective against experimental colitis in vivo, via reductions in TNF-associated inflammatory signaling. We used Sam68 knockout (KO) mice to study the role of Sam68 in experimental colitis, including its contributions to TNF-induced inflammatory gene expression in three-dimensional intestinal organoid cultures. We also studied the expression of Sam68 and inflammatory genes in colon tissues of UC patients. Sam68 KO mice treated with an acute course of DSS exhibited significantly less weight loss and histopathological inflammation compared to wild-type controls, suggesting that Sam68 contributes to experimental colitis. Bone marrow transplants showed no pathologic role for hematopoietic cell-specific Sam68, suggesting that non-hematopoietic Sam68 drives intestinal inflammation. Gene expression analyses showed that Sam68 deficiency reduced the expression of proinflammatory genes in colon tissues from DSS-treated mice, as well as TNF-treated three-dimensional colonic organoids. We also found that inflammatory genes, such as TNF, CCR2, CSF2, IL33 and CXCL10, as well as Sam68 protein, were upregulated in inflamed colon tissues of UC patients. This report identifies Sam68 as an important inflammatory driver in response to intestinal epithelial damage, suggesting that targeting Sam68 may hold promise to treat UC patients., (© 2021. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Vitiligo is a common depigmentation disease characterized by melanocyte death, which is attributed to various mechanisms such as apoptosis and autoimmune destruction. However, whether necroptosis, a newly discovered way of cell death, plays a key role in the pathogenesis of vitiligo is still elusive and has not been well-studied. In this study, we found that necroptosis markers, including phosphorylated RIP3 and phosphorylated-MLKL, were positive in melanocytes from vitiligo perilesional skin, which supported the existence of necroptosis in vitiligo. Furthermore, the expression of RIP1 was remarkably upregulated in melanocytes treated with hydrogen peroxide. Then, RIP1 intervention suppression and MLKL deficiency could significantly enhance the resistance of melanocytes to hydrogen peroxide‒induced necroptosis. Mechanistically, we confirmed that RIP1 and RIP3 could form necrosomes under oxidative stress and further trigger phosphorylated MLKL translocation to the cell membrane, which led to the destruction of melanocytes. Finally, we showed that RIP1-mediated generation of mitochondrial ROS contributed to necrosome formation in melanocytes. Collectively, our study confirms that necroptosis significantly facilitates oxidative stress‒induced melanocyte death through the RIP1 signaling pathway, offering insight into vitiligo., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) is a condition with an imbalanced inflammatory response and delayed resolution of inflammation. Macrophage polarization plays an important role in inflammation and resolution. However, the mechanism of macrophage polarization in ALI/ARDS is not fully understood. We found that mice with lipopolysaccharide administration developed lung injury with the accumulation of extracellular cold-inducible RNA-binding protein (eCIRP) in the lungs. eCIRP, as a damage-associated molecular pattern (DAMP), inhibited M2 macrophage polarization, thereby tipping the balance toward inflammation rather than resolution. Anti-CIRP antibodies reversed such phenotypes. The levels of macrophage erythropoietin (EPO) receptor (EPOR) were reduced after eCIRP treatment. Myeloid-specific EPOR-deficient mice displayed restrained M2 macrophage polarization and impaired inflammation resolution. Mechanistically, eCIRP impaired Rab26, a member of Ras superfamilies of small G proteins, and reduced the transportation of surface EPOR, which resulted in macrophage polarization toward the M1 phenotype. Moreover, EPO treatment hardly promotes M2 polarization in Rab26 knockout (KO) macrophages through EPOR. Collectively, macrophage EPOR signaling is impaired by eCIRP through Rab26 during ALI/ARDS, leading to the restrained M2 macrophage polarization and delayed inflammation resolution. These findings identify a mechanism of persistent inflammation and a potential therapy during ALI/ARDS., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zhang, Wang, Li, Xu, Wang, Wu, Gao, Qian, You, Zhang, He and Wang.)

Intracranial aneurysm (IA) rupture leads to subarachnoid hemorrhage, a sudden-onset disease that often causes death or severe disability. Although genome-wide association studies have identified common genetic variants that increase IA risk moderately, the contribution of variants with large effect remains poorly defined. Using whole-exome sequencing, we identified significant enrichment of rare, deleterious mutations in PPIL4, encoding peptidyl-prolyl cis-trans isomerase-like 4, in both familial and index IA cases. Ppil4 depletion in vertebrate models causes intracerebral hemorrhage, defects in cerebrovascular morphology and impaired Wnt signaling. Wild-type, but not IA-mutant, PPIL4 potentiates Wnt signaling by binding JMJD6, a known angiogenesis regulator and Wnt activator. These findings identify a novel PPIL4-dependent Wnt signaling mechanism involved in brain-specific angiogenesis and maintenance of cerebrovascular integrity and implicate PPIL4 gene mutations in the pathogenesis of IA., (© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Cardiomyocyte apoptosis in response to inflammation is a primary cause of myocardial ischemia-reperfusion injury (IRI). Nuclear factor erythroid 2 like 2 (Nrf2) reportedly plays an important role in myocardial IRI, but the underlying mechanism remains obscure. Expression data from the normal heart tissues of mice or heart tissues treated with reperfusion for 6 h after ischemia (IR6h) were acquired from the GEO database; changes in biological function and infiltrating immune cells were analyzed. The binding between the molecules was verified by chromatin immunoprecipitation sequencing. Based on confirmation that early myocardial ischemia-reperfusion (myocardial ischemia/reperfusion for 6 hours, IR6h) promoted myocardial apoptosis and inflammatory response, we found that Nrf2, cooperating with Programmed Cell Death 4, promoted transcription initiation of C-C Motif Chemokine Ligand 3 (Ccl3) in myocardial tissues of mice treated with IR6h. Moreover, Ccl3 contributed to the high signature score of C-C motif chemokine receptor 1 (Ccr1)-positive macrophages. The high signature score of Ccr1-positive macrophages leads to the release of pro-inflammatory factors interleukin 1 beta and interleukin 6. This study is the first to elucidate the damaging effect of Nrf2 via remodeling of the immune microenvironment in early myocardial ischemia-reperfusion, which provides us with new perspectives and treatment strategies for myocardial ischemia-reperfusion., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zhang, Liu, Cao, Wang, Cui, Yang, Wang and Shi.)

Inhibition of the 14q32 microRNAs, miR-329-3p and miR-495-3p, improves post-ischemic neovascularization. Cold-inducible RNA-binding protein ( CIRBP ) facilitates maturation of these microRNAs. We hypothesized that CIRBP deficiency improves post-ischemic angiogenesis via downregulation of 14q32 microRNA expression. We investigated these regulatory mechanisms both in vitro and in vivo. We induced hindlimb ischemia in Cirp -/- and C57Bl/6-J mice, monitored blood flow recovery with laser Doppler perfusion imaging, and assessed neovascularization via immunohistochemistry. Post-ischemic angiogenesis was enhanced in Cirp -/- mice by 34.3% with no effects on arteriogenesis. In vivo at day 7, miR-329-3p and miR-495-3p expression were downregulated in Cirp -/- mice by 40.6% and 36.2%. In HUVECs, CIRBP expression was upregulated under hypothermia, while miR-329-3p and miR-495-3p expression remained unaffected. siRNA-mediated CIRBP knockdown led to the downregulation of CIRBP -splice-variant-1 ( CIRBP-SV1 ), CIRBP antisense long noncoding RNA (lncRNA- CIRBP -AS1), and miR-495-3p with no effects on the expression of CIRBP-SV2-4 or miR-329-3p. siRNA-mediated CIRBP knockdown improved HUVEC migration and tube formation. SiRNA-mediated lncRNA- CIRBP -AS1 knockdown had similar long-term effects. After short incubation times, however, only CIRBP knockdown affected angiogenesis, indicating that the effects of lncRNA- CIRBP -AS1 knockdown were secondary to CIRBP-SV1 downregulation. CIRBP is a negative regulator of angiogenesis in vitro and in vivo and acts, at least in part, through the regulation of miR-329-3p and miR-495-3p.

Cellular senescence is associated with pleiotropic physiopathological processes, including aging and age-related diseases. The persistent DNA damage is a major stress leading to senescence, but the underlying molecular link remains elusive. Here, we identify La Ribonucleoprotein 7 (LARP7), a 7SK RNA binding protein, as an aging antagonist. DNA damage-mediated Ataxia Telangiectasia Mutated (ATM) activation triggers the extracellular shuttling and downregulation of LARP7, which dampens SIRT1 deacetylase activity, enhances p53 and NF-κB (p65) transcriptional activity by augmenting their acetylation, and thereby accelerates cellular senescence. Deletion of LARP7 leads to senescent cell accumulation and premature aging in rodent model. Furthermore, we show this ATM-LARP7-SIRT1-p53/p65 senescence axis is active in vascular senescence and atherogenesis, and preventing its activation substantially alleviates senescence and atherogenesis. Together, this study identifies LARP7 as a gatekeeper of senescence, and the altered ATM-LARP7-SIRT1-p53/p65 pathway plays an important role in DNA damage response (DDR)-mediated cellular senescence and atherosclerosis., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

FRG1 has a role in tumorigenesis and angiogenesis. Our preliminary analysis showed that FRG1 mRNA expression is associated with overall survival (OS) in certain cancers, but the effect varies. In cervix and gastric cancers, we found a clear difference in the OS between the low and high FRG1 mRNA expression groups, but the difference was not prominent in breast, lung, and liver cancers. We hypothesized that FRG1 expression level could affect the functionality of the correlated genes or vice versa, which might mask the effect of a single gene on the OS analysis in cancer patients. We used the multivariate Cox regression, risk score, and Kaplan Meier analyses to determine OS in a multigene model. STRING, Cytoscape, HIPPIE, Gene Ontology, and DAVID (KEGG) were used to deduce FRG1 associated pathways. In breast, lung, and liver cancers, we found a distinct difference in the OS between the low and high FRG1 mRNA expression groups in the multigene model, suggesting an independent role of FRG1 in survival. Risk scores were calculated based upon regression coefficients in the multigene model. Low and high-risk score groups showed a significant difference in the FRG1 mRNA expression level and OS. HPF1, RPL34, and EXOSC9 were the most common genes present in FRG1 associated pathways across the cancer types. Validation of the effect of FRG1 mRNA expression level on these genes by qRT-PCR supports that FRG1 might be an upstream regulator of their expression. These genes may have multiple regulators, which also affect their expression, leading to the masking effect in the survival analysis. In conclusion, our study highlights the role of FRG1 in the survivability of cancer patients in tissue-specific manner and the use of multigene models in prognosis., (© 2021. The Author(s).)

Ferroptosis, an iron-dependent nonapoptotic cell death, is a highly regulated tumor suppressing process. However, functions and mechanisms of RNA-binding proteins in regulation of evasion of ferroptosis during lung cancer progression are still largely unknown. Here, we report that the RNA-binding protein RBMS1 participates in lung cancer development via mediating ferroptosis evasion. Through an shRNA-mediated systematic screen, we discovered that RBMS1 is a key ferroptosis regulator. Clinically, RBMS1 was elevated in lung cancer and its high expression was associated with reduced patient survival. Conversely, depletion of RBMS1 inhibited lung cancer progression both in vivo and in vitro. Mechanistically, RBMS1 interacted with the translation initiation factor eIF3d directly to bridge the 3'- and 5'-UTR of SLC7A11. RBMS1 ablation inhibited the translation of SLC7A11, reduced SLC7A11-mediated cystine uptake, and promoted ferroptosis. In a drug screen that targeted RBMS1, we further uncovered that nortriptyline hydrochloride decreased the level of RBMS1, thereby promoting ferroptosis. Importantly, RBMS1 depletion or inhibition by nortriptyline hydrochloride sensitized radioresistant lung cancer cells to radiotherapy. Our findings established RBMS1 as a translational regulator of ferroptosis and a prognostic factor with therapeutic potential and clinical value.

High expression of LIN28B is associated with aggressive malignancy and poor survival. Here, probing MYCN-amplified neuroblastoma as a model system, we showed that LIN28B expression was associated with enhanced cell migration in vitro and invasive and metastatic behavior in murine xenografts. Sequence analysis of the polyribosome fraction of LIN28B-expressing neuroblastoma cells revealed let-7-independent enrichment of transcripts encoding components of the translational and ribosomal apparatus and depletion of transcripts of neuronal developmental programs. We further observed that LIN28B utilizes both its cold shock and zinc finger RNA binding domains to preferentially interact with MYCN-induced transcripts of the ribosomal complex, enhancing their translation. These data demonstrated that LIN28B couples the MYCN-driven transcriptional program to enhanced ribosomal translation, thereby implicating LIN28B as a posttranscriptional driver of the metastatic phenotype.

Background: LIN28B is functionally driving malignant transformation and relevance to the worse disease outcomes by promoting cancer aggressiveness. However, a typical role of LIN28B in cholangiocarcinoma (CCA) is primarily unknown. In this study, the tumorigenic potential of LIN28B in the cholangiocyte context was investigated., Methods: Stable LIN28B expression in MMNK-1 cells was generated by infecting with retrovirus-containing LIN28B gene. LIN28B-overexpressing cells were further validated the amount of released cytokines by using human cytokine arrays. After treatment of chemo-drugs, cell viability was subsequently measured using MTT assay. Aldehyde dehydrogenase (ALDH) activity was determined using ALDEFLUOR assay Kit and analyzed by flow cytometry. The mRNA and protein expression levels were respectively assayed by RT-qPCR and western blot., Results: Cytokine release results showed that numerous inflammatory cytokines-chemokines related to cancer initiation and development, such as IL-8, IL-6, VEGF, MCP1, TNF-α were significantly increased in LIN28B-overexpressing MMNK-1 cells. Drug sensitivity test showed that LIN28B-overexpressing MMNK-1 treated cells had a high percentage of cell viability compared to MMNK-1-control treated cells. Activity and expression level of a cancer stem cell marker, ALDH was significantly elevated in LIN28B-overexpressing MMNK-1 cells. Moreover, the activation of an oncogenic signaling pathway, signal transducer and activator of transcription 3 (STAT3) was enhanced in LIN28B-overexpressing MMNK-1 cells. Whereas, growth capacity of LIN28B-overexpressing MMNK-1 cells was found to be reduced in STAT3 inhibition., Conclusion: LIN28B can regulate the inflammatory response and resistance to chemotherapy of cholangiocytes through modulation of STAT3 signaling pathway.A recent study suggests that activated cholangiocytes can be induced by regulation of LIN28B/STAT3 pathway and this may partially contribute to the initiating CCA. Here, LIN28B and its downstream signaling could be considered as an attractive therapeutic target in patients with CCA.

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are two clinically distinct classes of neurodegenerative disorders. Yet, they share a range of genetic, cellular, and molecular features. Hexanucleotide repeat expansions (HREs) in the C9orf72 gene and the accumulation of toxic protein aggregates in the nervous systems of the affected individuals are among such common features. Though the mechanisms by which HREs cause toxicity is not clear, the toxic gain of function due to transcribed HRE RNA or dipeptide repeat proteins (DPRs) produced by repeat-associated non-AUG translation together with a reduction in C9orf72 expression are proposed as the contributing factors for disease pathogenesis in ALS and FTD. In addition, several recent studies point toward alterations in protein homeostasis as one of the root causes of the disease pathogenesis. In this review, we discuss the effects of the C9orf72 HRE in the autophagy-lysosome pathway based on various recent findings. We suggest that dysfunction of the autophagy-lysosome pathway synergizes with toxicity from C9orf72 repeat RNA and DPRs to drive disease pathogenesis. Abbreviation: ALP: autophagy-lysosome pathway; ALS: amyotrophic lateral sclerosis; AMPK: AMP-activated protein kinase; ATG: autophagy-related; ASO: antisense oligonucleotide; C9orf72: C9orf72-SMCR8 complex subunit; DENN: differentially expressed in normal and neoplastic cells; DPR: dipeptide repeat protein; EIF2A/eIF2α: eukaryotic translation initiation factor 2A; ER: endoplasmic reticulum; FTD: frontotemporal dementia; GAP: GTPase-activating protein; GEF: guanine nucleotide exchange factor; HRE: hexanucleotide repeat expansion; iPSC: induced pluripotent stem cell; ISR: integrated stress response; M6PR: mannose-6-phosphate receptor, cation dependent; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MN: motor neuron; MTORC1: mechanistic target of rapamycin kinase complex 1; ND: neurodegenerative disorder; RAN: repeat-associated non-ATG; RB1CC1/FIP200: RB1 inducible coiled-coil 1; SLC66A1/PQLC2: solute carrier family 66 member 1; SMCR8: SMCR8-C9orf72 complex subunit; SQSTM1/p62: sequestosome 1; STX17: syntaxin 17; TARDBP/TDP-43: TAR DNA binding protein; TBK1: TANK binding kinase 1; TFEB: transcription factor EB; ULK1: unc-51 like autophagy activating kinase 1; UPS: ubiquitin-proteasome system; WDR41: WD repeat domain 41.

Somatic cell nuclear transfer (SCNT) can reprogram differentiated somatic cells to produce individual animals, thus having advantages in animal breeding and chromatin reprogramming. Interspecies SCNT (iSCNT) provides extreme cases of reprogramming failure that can be used to understand the basic biological mechanism of genome reprogramming. It is important to understand the possible mechanisms for the failure of zygotic genome activation (ZGA) in iSCNT embryos in order to improve the efficiency of SCNT embryos. In the present study, we compared the development of bovine-bovine (B-B), ovine-ovine (O-O) SCNT, and ovine-bovine (O-B) iSCNT embryos and found that a developmental block existed in the 8-cell stage in O-B iSCNT embryos. RNA sequencing and q-PCR analysis revealed that the large ribosomal subunit genes (RPL) or the small ribosomal subunit genes (RPS) were expressed at lower levels in the O-B iSCNT embryos. The nucleolin (C23) gene that regulates the ribosomal subunit generation was transcribed at a lower level during embryonic development in O-B iSCNT embryos. In addition, the nucleolin exhibited a clear circular-ring structure in B-B 8-cell stage embryos, whereas this was shell-like or dot-like in the O-B embryos. Furthermore, overexpression of C23 could increase the blastocyst rate of both SCNT and iSCNT embryos and partly rectify the ring-like nucleolin structure and the expression of ribosomal subunit related genes were upregulation, while knockdown of C23 increased the shell-like nucleolin-structure in B-B cloned embryos and downregulated the expression of ribosomal subunit related genes. These results implied that abnormal C23 and ribosome subunit gene expression would lead to the developmental block of iSCNT embryos and ZGA failure. Overexpression of the C23 gene could partly improve the blastocyst development and facilitate the nucleolin structure in bovine preimplantation SCNT embryos., (© 2021 Federation of American Societies for Experimental Biology.)

Circ-RBM4 (mm9_circ_013935) has been revealed to have low expression in the renal tissues of diabetic nephropathy (DN) mice, and its underlying regulatory mechanism remains unexplored. The high glucose (HG) - treated mouse podocytes were used to establish DN cell models. A cell counting kit-8 assay was used to examine the viability of mouse podocytes. The expression of proteins related to fibrosis (collagen I, collagen III, fibronectin) was detected using Western blot. The concentration of inflammation cytokines (tumor necrosis factor α, interleukin 1β (IL-1β), IL-8) in mouse podocytes was assessed by ELISA. The interaction between genes was explored by luciferase reporter assays. HG treatment decreased the viability and elevated the expression of fibrosis and inflammation factors in mouse podocytes. Circ-RBM4 expression was downregulated in HG-treated mouse podocytes. Circ-RBM4 overexpression reversed HG-induced increase in levels of proteins related to fibrosis and the concentration of inflammation factors. The miR-153-3p was revealed to bind with circ-RBM4 and directly targeted nuclear factor I/C (NFIC) in mouse podocytes. Rescue assays indicated that circ-RBM4 attenuated HG-induced fibrosis and inflammation response in mouse podocytes by inhibiting miR-153-3p expression or upregulating NFIC expression. Circ-RBM4 alleviated the renal inflammation and renal fibrosis in DN by targeting the miR-153-3p/NFIC axis.

Several genes in innate immunity have been implicated in Alzheimer's disease (AD). However, the effect of innate immunity on amyloid β (Aβ) production, which makes amyloid plaques in AD brains, was previously not known. Recently, the antiviral protein interferon-induced transmembrane protein 3 (IFITM3) has been identified as a novel γ-secretase modulatory protein for Aβ production. In this review, the mechanisms of how innate immunity modulates Aβ production via IFITM3-γ-secretase complexes and contributes to AD pathogenesis are discussed.

Cold‑inducible RNA‑binding protein (CIRBP) is a cold‑shock protein comprised of an RNA‑binding motif that is induced by several stressors, such as cold shock, UV radiation, nutrient deprivation, reactive oxygen species and hypoxia. CIRBP can modulate post‑transcriptional regulation of target mRNA, which is required to control DNA repair, circadian rhythms, cell growth, telomere integrity and cardiac physiology. In addition, the crucial function of CIRBP in various human diseases, including cancers and inflammatory disease, has been reported. Although CIRBP is primarily considered to be an oncogene, it may also serve a role in tumor suppression. In the present study, the controversial roles of CIRBP in various human cancers is summarized, with a focus on the interconnectivity between CIRBP and its target mRNAs involved in tumorigenesis. CIRBP may represent an important prognostic marker and therapeutic target for cancer therapy.

Sex steroids play critical roles in sex differentiation and gonadal development in teleosts. Steroidogenic acute regulatory protein (StAR), transporting cholesterol (the substrate for steroidogenesis) from the outer mitochondrial membrane to the inner membrane, is the first rate-limiting factor of steroidogenesis. Interestingly, two StAR genes (named as StAR1 and StAR2) have been isolated from non-mammalian vertebrates. To characterize the functions of the novel StAR2 gene in the gonadal differentiation and fertility, we generated a StAR2 homozygous mutant line in Nile tilapia (Oreochromis niloticus). StAR2 gene knockout in male tilapia impeded meiotic initiation, associate with the down-regulation of meiosis related gene expressions of vasa, sycp3 and dazl at 90 days after hatching (dah). Meanwhile, cyp11b2 expression and serum 11-KT production significantly declined in StAR2 -/- XY fish at 90 dah. From 120-300 dah, spermatogenesis gradually recovered, and so did the expressions of vasa, sycp3 and dazl in StAR2 -/- XY fish testes. However, seminiferous lobules arranged disorderly in StAR2 -/- XY fish testes at 300 dah. The number of Leydig cells and expressions of downstream steroidogenesis enzymes including cyp11a1, 3β-HSD-I, 3β-HSD-II, cyp17a1 and cyp17a2 decreased in StAR2 -/- XY fish testes at 300 dah. Serum testosterone and 11-KT levels were significantly lower in StAR2 -/- XY fish than that of their control counterparts. Furthermore, significantly elevated ar, fsh and lh expressions in StAR2-deficient XY fish testes and pituitaries were found when compared with the control XY fish. Testes degeneration and spermatogenic cell apoptosis were observed, while no sperm were squeezed out in StAR2 -/- XY fish testes at 540 dah. Taken together, our results suggest that StAR2 has a role in testicular development, spermatogenesis and spermiation by regulating androgen production in tilapia, but may not be essential and could be compensated., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The expression of microRNA‑383 (miR‑383) is downregulated in a variety of tumor tissues, and it exhibits antiproliferative activity in non‑small cell lung cancer cells. In the present study, an association between the downregulation of miR‑383 expression and the deletion of chr8p22 in patients with lung adenocarcinoma was identified. The promoting effect of miR‑383 on cisplatin sensitivity was verified both in vivo and in vitro . Additionally, it was revealed that the expression of RNA binding motif protein 24 (RBM24) protein was regulated by and negatively correlated with miR‑383 expression. Ectopic expression of RBM24 or inhibition of miR‑383 decreased the chemosensitivity of parental A549 cells, whereas knockdown of RBM24 in cisplatin‑resistant A549 cells increased chemosensitivity. Mechanistically, miR‑383 interfered with the activation of nuclear factor κB (NF‑κB) signaling through repression of RBM24‑mediated phosphorylation of Rel‑like domain‑containing protein A and inhibitor α of NF‑κB. Taken together, the downregulation of miR‑383 induced RBM24 expression, which was mediated through the activation of NF‑κB signaling, to contribute to chemotherapy resistance in lung adenocarcinoma cells. The results of the present study highlight potential therapeutic targets for the clinical reversal of the chemotherapy resistance in lung adenocarcinoma.

PGR3 is a P-class pentatricopeptide repeat (PPR) protein required for the stabilization of petL operon RNA and the translation of the petL gene in plastids. Irrespective of its important roles in plastids, key questions have remained unanswered, including how PGR3 protein promotes translation and which plastid mRNA PGR3 activates the translation. Here, we show that PGR3 facilitates the translation from ndhG, in addition to petL, through binding to their 5' untranslated regions (UTRs). Ribosome profiling and RNA sequencing in pgr3 mutants revealed that translation from petL and ndhG was specifically suppressed. Harnessing small RNA fragments protected by PPR proteins in vivo, we probed the PGR3 recruitment to the 5' UTRs of petL and ndhG. The putative PGR3-bound RNA segments per se repress the translation possibly with a strong secondary structure and thereby block ribosomes' access. However, the PGR3 binding antagonizes the effects and facilitates the protein synthesis from petL and ndhG in vitro. The prediction of the 3-dimensional structure of PGR3 suggests that the 26th PPR motif plays important roles in target RNA binding. Our data show the specificity of a plastidic RNA-binding protein and provide a mechanistic insight into translational control., (© The Author(s) 2021. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Adenosine deaminase acting on RNA 1 (ADAR1) is an enzyme responsible for double-stranded RNA (dsRNA)-specific adenosine-to-inosine RNA editing, which is estimated to occur at over 100 million sites in humans. ADAR1 is composed of two isoforms transcribed from different promoters: p150 and N-terminal truncated p110. Deletion of ADAR1 p150 in mice activates melanoma differentiation-associated protein 5 (MDA5)-sensing pathway, which recognizes endogenous unedited RNA as non-self. In contrast, we have recently demonstrated that ADAR1 p110-mediated RNA editing does not contribute to this function, implying that a unique Z-DNA/RNA-binding domain α (Zα) in the N terminus of ADAR1 p150 provides specific RNA editing, which is critical for preventing MDA5 activation. In addition, a mutation in the Zα domain is identified in patients with Aicardi-Goutières syndrome (AGS), an inherited encephalopathy characterized by overproduction of type I interferon. Accordingly, we and other groups have recently demonstrated that Adar1 Zα-mutated mice show MDA5-dependent type I interferon responses. Furthermore, one such mutant mouse carrying a W197A point mutation in the Zα domain, which inhibits Z-RNA binding, manifests AGS-like encephalopathy. These findings collectively suggest that Z-RNA binding by ADAR1 p150 is essential for proper RNA editing at certain sites, preventing aberrant MDA5 activation.

MicroRNAs (miRNAs) are ∼22 nt small noncoding RNAs that control gene expression at the posttranscriptional level through translational inhibition and destabilization of their target mRNAs. The biogenesis of miRNAs involves a series of processing steps beginning with cropping of the primary miRNA transcript by the Microprocessor complex, which is composed of Drosha and DGCR8. Here we report a novel regulatory interaction between the Microprocessor components and coilin, the Cajal body (CB) marker protein. Coilin knockdown causes alterations in the level of primary and mature miRNAs, let-7a and miR-34a, and their miRNA targets, HMGA2 and Notch1, respectively. We also found that coilin knockdown affects the levels of DGCR8 and Drosha in cells with (HeLa) and without (WI-38) CBs. To further explore the role of coilin in miRNA biogenesis, we conducted a series of coimmunoprecipitation experiments using coilin and DGCR8 constructs, which revealed that coilin and DGCR8 can form a complex. Additionally, our results indicate that phosphorylation of DGCR8, which has been shown to increase protein stability, is impacted by coilin knockdown. Collectively, our results implicate coilin as a member of the regulatory network governing miRNA biogenesis.

To explore the effect of insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2) on colorectal cancer (CRC) by recognizing the m6A modification of YAP mRNA thus activating ErbB2 expression. High expressions of IGF2BP2, YAP, and ErbB2 promoted the proliferation, migration and invasion of CRC cells and reduced their apoptosis. IGF2BP2 recognized the m6A on YAP mRNA and promoted the translation of mRNA. YAP regulated ErbB2 expression by promoting TEAD4 enrichment in ErbB2 promoter region. Therefore, IGF2BP2 promoted the expression of ErbB2 to enhance the proliferation, invasion and migration of CRC cells, to repress cell apoptosis, and to promote solid tumor formation in nude mice. IGF2BP2 activates the expression of ErbB2 by recognizing the m6A of YAP, thus affecting the cell cycle of CRC, inhibiting cell apoptosis, and promoting proliferation., (© 2021 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Purpose: Radiation therapy, which is vital for the treatment of primary liver cancer, comes with unavoidable liver injury, which limits its implementation. N6-methyladenosine (m6A) methylation is involved in many molecular functions. However, its role in radiation-induced liver diseases (RILD) remains unknown. Herein, we investigate the role of m6A methylation in RILD., Methods and Materials: Methylated RNA-immunoprecipitation sequencing and RNA transcriptome sequencing were used to reveal the methylation pattern of human hepatic stellate cells (HSCs) exposed to irradiation. C3H/HeN mice and stimulator of interferon genes (STING)-deficient mice underwent x-ray irradiation of 24 Gy in 3 fractions. The m6A methylation of the high-mobility group box 1 (HMGB1) transcript was validated using methylated RNA immunoprecipitation, RNA immunoprecipitation, luciferase assays, and a messenger RNA decay assay., Results: Human hepatic stellate cells showed significant differences in methylation patterns after 8 Gy of x-ray irradiation. Irradiation recruited AlkB homolog 5 (ALKBH5) to demethylate m6A residues in the 3' untranslated region of HMGB1, which resulted in the activation of STING-interferon regulatory factor 3 signaling. Changes in the transcription of the 3' untranslated region of HMGB1 occurred after the knockdown of ALKBH5, which were eliminated after m6A residue mutation. Strikingly, ALKBH5 deficiency or HMGB1 silencing both attenuated type I interferon production and decreased hepatocyte apoptosis. In vivo depletion of ALKBH5 abolished the upregulation of HMGB1-mediated STING signaling and decreased liver inflammation, which was consistent with STING -/- mice treated with irradiation. Notably, YTHDF2 (m6A reader protein) directly bound to HMGB1 m6A-modified sites and promoted its degradation., Conclusions: ALKBH5-dependent HMGB1 expression mediates STING-interferon regulatory factor 3 innate immune response in RILD., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Among low molecular weight substances, polyamines (spermidine, spermine and their precursor putrescine) are present in eukaryotic cells at the mM level together with ATP and glutathione. It is expected therefore that polyamines play important roles in cell proliferation and viability. Polyamines mainly exist as a polyamine-RNA complex and regulate protein synthesis. It was found that polyamines enhance translation from inefficient mRNAs. The detailed mechanisms of polyamine stimulation of specific kinds of protein syntheses and the physiological functions of these proteins are described in this review. Spermine is metabolized into acrolein (CH 2  = CH-CHO) and hydrogen peroxide (H 2 O 2 ) by spermine oxidase. Although it is thought that cell damage is mainly caused by reactive oxygen species (O 2 - , H 2 O 2 , and •OH), it was found that acrolein is much more toxic than H 2 O 2 . Accordingly, the level of acrolein produced becomes a useful biomarker for several tissue-damage diseases like brain stroke. Thus, the mechanisms of cell toxicity caused by acrolein are described in this review., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Homozygous mutation of the RNA kinase CLP1 (cleavage factor polyribonucleotide kinase subunit 1) causes pontocerebellar hypoplasia type 10 (PCH10), a pediatric neurodegenerative disease. CLP1 is associated with the transfer RNA (tRNA) splicing endonuclease complex and the cleavage and polyadenylation machinery, but its function remains unclear. We generated two mouse models of PCH10: one homozygous for the disease-associated Clp1 mutation, R140H, and one heterozygous for this mutation and a null allele. Both models exhibit loss of lower motor neurons and neurons of the deep cerebellar nuclei. To explore whether Clp1 mutation impacts tRNA splicing, we profiled the products of intron-containing tRNA genes. While mature tRNAs were expressed at normal levels in mutant mice, numerous other products of intron-containing tRNA genes were dysregulated, with pre-tRNAs, introns, and certain tRNA fragments up-regulated, and other fragments down-regulated. However, the spatiotemporal patterns of dysregulation do not correlate with pathogenicity for most altered tRNA products. To elucidate the effect of Clp1 mutation on precursor messenger RNA (pre-mRNA) cleavage, we analyzed poly(A) site (PAS) usage and gene expression in Clp1 R140H/- spinal cord. PAS usage was shifted from proximal to distal sites in the mutant mouse, particularly in short and closely spaced genes. Many such genes were also expressed at lower levels in the Clp1 R140H/- mouse, possibly as a result of impaired transcript maturation. These findings are consistent with the hypothesis that select genes are particularly dependent upon CLP1 for proper pre-mRNA cleavage, suggesting that impaired mRNA 3' processing may contribute to pathogenesis in PCH10., Competing Interests: The authors declare no competing interest., (Copyright © 2021 the Author(s). Published by PNAS.)

Gene deletion and gene expression alteration can lead to growth defects that are amplified or reduced when a second mutation is present in the same cells. We performed 154 genetic interaction mapping (GIM) screens with query mutants related with RNA metabolism and estimated the growth rates of about 700 000 double mutant Saccharomyces cerevisiae strains. The tested targets included the gene deletion collection and 900 strains in which essential genes were affected by mRNA destabilization (DAmP). To analyze the results, we developed RECAP, a strategy that validates genetic interaction profiles by comparison with gene co-citation frequency, and identified links between 1471 genes and 117 biological processes. In addition to these large-scale results, we validated both enhancement and suppression of slow growth measured for specific RNA-related pathways. Thus, negative genetic interactions identified a role for the OCA inositol polyphosphate hydrolase complex in mRNA translation initiation. By analysis of suppressors, we found that Puf4, a Pumilio family RNA binding protein, inhibits ribosomal protein Rpl9 function, by acting on a conserved UGUAcauUA motif located downstream the stop codon of the RPL9B mRNA. Altogether, the results and their analysis should represent a useful resource for discovery of gene function in yeast., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Glioma ranks first among the aggressive brain tumors all over the world. LncRNA LINC00689 has been confirmed to play key roles in the progression of cancers, and LINC00689 was upregulated in glioma. However, the biological function of LINC00689 in glioma is unclear. qRT-PCR was applied to detect the expressions of LINC00689 and miR-526b-3p in glioma cells. Dual-luciferase report was performed to examine the relation among LINC00689, miR-526b-3p, and insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1). Then, the growth, migration, and invasion of glioma cells were detected by colony formation, flow cytometry, and transwell assay, respectively. The expressions of p21, cleaved caspase 3, and MAPK signaling-related proteins in glioma cells were tested by western blotting. Finally, xenograft mice model was established to detect the effect of LINC00689 on tumor growth of glioma in vivo. LINC00689 was upregulated in glioma cells, while miR-526b-3p was downregulated. In addition, LINC00689 bound to miR-526b-3p, and IGFBP1 was targeted by miR-526b-3p. Moreover, LINC00689 knockdown or upregulation of miR-526b-3p inhibited the proliferation of glioma cells and induced the apoptosis. Consistently, the migration and invasion of glioma cells were notably reduced by LINC00689 shRNA/miR-526-3p mimics. miR-526b-3p inhibitor or IGF2BP1 upregulation could reverse the effect of LINC00689 knockdown or miR-526b-3p mimics. Finally, knockdown of LINC00689 inhibited the tumor growth of glioma in vivo through regulating miR-526b-3p/IGF2BP1/MAPK axis. In conclusion, silencing of LINC00689 could inhibit the tumorigenesis of glioma via mediation of miR-526b-3p/IGF2BP1 axis. LINC00689 may serve as a new target for the treatment of glioma., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC part of Springer Nature.)

DNA damage, induced by either chemical carcinogens or environmental pollutants, plays an important role in the initiation of colorectal cancer. DNA repair processes, however, are involved in both protecting against cancer formation, and also contributing to cancer development, by ensuring genomic integrity and promoting the efficient DNA repair in tumor cells, respectively. Although DNA repair pathways have been well exploited in the treatment of breast and ovarian cancers, the role of DNA repair processes and their therapeutic efficacy in colorectal cancer is yet to be appreciably explored. To understand the role of DNA repair, especially homologous recombination (HR), in chemical carcinogen-induced colorectal cancer growth, we unraveled the role of RAD51AP1 (RAD51-associated protein 1), a protein involved in HR, in genotoxic carcinogen (azoxymethane, AOM)-induced colorectal cancer. Although AOM treatment alone significantly increased RAD51AP1 expression, the combination of AOM and dextran sulfate sodium (DSS) treatment dramatically increased by several folds. RAD51AP1 expression is found in mouse colonic crypt and proliferating cells. RAD51AP1 expression is significantly increased in majority of human colorectal cancer tissues, including BRAF/KRAS mutant colorectal cancer, and associated with reduced treatment response and poor prognosis. Rad51ap1 -deficient mice were protected against AOM/DSS-induced colorectal cancer. These observations were recapitulated in a genetically engineered mouse model of colorectal cancer ( Apc Min /+ ). Furthermore, chemotherapy-resistant colorectal cancer is associated with increased RAD51AP1 expression. This phenomenon is associated with reduced cell proliferation and colorectal cancer stem cell (CRCSC) self-renewal. Overall, our studies provide evidence that RAD51AP1 could be a novel diagnostic marker for colorectal cancer and a potential therapeutic target for colorectal cancer prevention and treatment. IMPLICATIONS: This study provides first in vivo evidence that RAD51AP1 plays a critical role in colorectal cancer growth and drug resistance by regulating CRCSC self-renewal., (©2021 American Association for Cancer Research.)

Objective: To examine the role of YTHDF1 knock-down macrophages on the immunity of severe sepsis rats with ECMO., Methods: 15 SD rats were randomly allocated into 3 groups: mild sepsis (I), severe sepsis with ECMO (II), and YTHDF1 knock-down macrophages treatment groups (III). Blood biochemical indexes, different immune factors and brain changes were detected by RT-PCR, ELISA, ELISPOT and HE staining. Isolated macrophages subtypes and signal proteins were detected by flow cytometry, western blot and m6A RNA methylation test., Results: The levels of HMGB1, RAGE, YTHDF1 and IL-17 in peripheral blood were significantly higher (p < 0.01), while the level of CXCL9 and TNF-α, and LPS-specific CD8 + CTL function were significantly decreased in group II compared with group I (p < 0.01). The ratio of CD63 + macrophages (p < 0.05) and CD64 + macrophages (p< 0.05) decreased and the level of elastase (p < 0.01) and CCR2 high CX3CR1 low /CCR2 low CX3CR1 high (p < 0.01) of macrophages increased in group II. The above were consistent with the severity of biochemical indicators, the increasing endothelial injury factor (Ang2/Ang1), lower endothelial protective factor (sTie2), severer brain injury in group II. After YTHDF1 knock-down macrophages treatment, the above indexes' changes were opposite when Group III versus Group II through the down-regulation of m6A RNA methylation of JAK2/STAT3 (p < 0.01) and protein expression of P JAK2/ P STAT3 (p < 0.05) in isolated macrophages., Conclusions: YTHDF1 knock-down macrophages improved the immune paralysis of macrophages, Th1/Th17 and CTL and reduced the entry of macrophages into the brain to cause endothelial damage of severe sepsis rats with ECMO through the inhibition of HMGB1/RAGE and YTHDF1, m6A RNA methylation of JAK2/STAT3 and P JAK2/ P STAT3 proteins expression in macrophages., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Extracellular Cold-inducible RNA-binding protein (eCIRP), a damage-associated molecular pattern, is released from cells upon hypoxia and cold-stress. The overall absence of extra- and intracellular CIRP is associated with increased angiogenesis, most likely induced through influencing leukocyte accumulation. The aim of the present study was to specifically characterize the role of eCIRP in ischemia-induced angiogenesis together with the associated leukocyte recruitment. For analyzing eCIRPs impact, we induced muscle ischemia via femoral artery ligation (FAL) in mice in the presence or absence of an anti-CIRP antibody and isolated the gastrocnemius muscle for immunohistological analyses. Upon eCIRP-depletion, mice showed increased capillary/muscle fiber ratio and numbers of proliferating endothelial cells (CD31 + /CD45 - /BrdU + ). This was accompanied by a reduction of total leukocyte count (CD45 + ), neutrophils (MPO + ), neutrophil extracellular traps (NETs) (MPO + CitH3 + ), apoptotic area (ascertained via TUNEL assay), and pro-inflammatory M1-like polarized macrophages (CD68 + /MRC1 - ) in ischemic muscle tissue. Conversely, the number of regenerative M2-like polarized macrophages (CD68 + /MRC1 + ) was elevated. Altogether, we observed that eCIRP depletion similarly affected angiogenesis and leukocyte recruitment as described for the overall absence of CIRP. Thus, we propose that eCIRP is mainly responsible for modulating angiogenesis via promoting pro-angiogenic microenvironmental conditions in muscle ischemia.

Background: Cold-inducible RNA binding protein (CIRP) is a newly discovered proto-oncogene. In this study, we investigated the role of CIRP in the progression of non-small cell lung cancer (NSCLC) using patient tissue samples, cultured cell lines and animal lung cancer models., Methods: Tissue arrays, IHC and HE staining, immunoblotting, and qRT-PCR were used to detect the indicated gene expression; plasmid and siRNA transfections as well as viral infection were used to manipulate gene expression; cell proliferation assay, cell cycle analysis, cell migration and invasion analysis, soft agar colony formation assay, tail intravenous injection and subcutaneous inoculation of animal models were performed to study the role of CIRP in NSCLC cells; Gene expression microarray was used to select the underlying pathways; and RNA immunoprecipitation assay, biotin pull-down assay, immunopurification assay, mRNA decay analyses and luciferase reporter assay were performed to elucidate the mechanisms. The log-rank (Mantel-Cox) test, independent sample T-test, nonparametric Mann-Whitney test, Spearman rank test and two-tailed independent sample T-test were used accordingly in our study., Results: Our data showed that CIRP was highly expressed in NSCLC tissue, and its level was negatively correlated with the prognosis of NSCLC patients. By manipulating CIRP expression in A549, H460, H1299, and H1650 cell lines, we demonstrated that CIRP overexpression promoted the transition of G1/G0 phase to S phase and the formation of an enhanced malignant phenotype of NSCLC, reflected by increased proliferation, enhanced invasion/metastasis and greater tumorigenic capabilities both in vitro and in vivo. Transcriptome sequencing further demonstrated that CIRP acted on the cell cycle, DNA replication and Wnt signaling pathway to exert its pro-oncogenic action. Mechanistically, CIRP directly bound to the 3'- and 5'-UTRs of CTNNB1 mRNA, leading to enhanced stability and translation of CTNNB1 mRNA and promoting IRES-mediated protein synthesis, respectively. Eventually, the increased CTNNB1 protein levels mediated excessive activation of the Wnt/β-catenin signaling pathway and its downstream targets C-myc, COX-2, CCND1, MMP7, VEGFA and CD44., Conclusion: Our results support CIRP as a candidate oncogene in NSCLC and a potential target for NSCLC therapy., (© 2021. The Author(s).)

YTH domain family 2 (YTHDF2) is an N6-methyladenosine (m 6 A) binding protein promoting mRNA degradation in various biological processes. Despite its essential roles, the role of YTHDF2 in determining cell fates has not been fully elucidated. Notch signaling plays a vital role in determining cell fates, such as proliferation, differentiation, and apoptosis. We investigated the effect of YTHDF2 on Notch signaling. Our results show that YTHDF2 inhibits Notch signaling by downregulating the Notch1 , HES1 , and HES5 mRNA levels. Analyzing YTHDF2 deletion mutants indicates that the YTH domain is critical in regulating the Notch signal by directly binding m 6 A of Notch1 mRNA. Recently, YTHDF2 nuclear translocation was reported under heat shock conditions, but its physiological function is unknown. In our study, the YTH domain is required for YTHDF2 nuclear translocation. In addition, under heat shock stress, the Notch signal was significantly restored due to the increased expression of the Notch1 targets. These results suggest that YTHDF2 in the cytoplasm may act as an intrinsic suppressor in Notch signaling by promoting Notch1 mRNA degradation under normal cellular conditions. Conversely, upon the extracellular stress such as heat shock, YTHDF2 nuclear translocation resulting in reduced Notch1 mRNA decay may contribute to the increasing of Notch intracellular domain (NICD) regulating the survival-related target genes., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Background: Germ cell tumors (GCTs) are developmental cancers, tightly linked to embryogenesis and germ cell development. The recent and expanding field of RNA modifications is being increasingly implicated in such molecular events, as well as in tumor progression and resistance to therapy, but still rarely explored in GCTs. In this work, and as a follow-up of our recent study on this topic in TGCT tissue samples, we aim to investigate the role of N6-methyladenosine (m 6 A), the most abundant of such modifications in mRNA, in in vitro and in vivo models representative of such tumors., Methods: Four cell lines representative of GCTs (three testicular and one mediastinal), including an isogenic cisplatin resistant subline, were used. CRISPR/Cas9-mediated knockdown of VIRMA was established and the chorioallantoic membrane assay was used to study its phenotypic effect in vivo., Results: We demonstrated the differential expression of the various m 6 A writers, readers and erasers in GCT cell lines representative of the major classes of these tumors, seminomas and non-seminomas, and we evidenced changes occurring upon differentiation with all-trans retinoic acid treatment. We showed differential expression also among cells sensitive and resistant to cisplatin treatment, implicating these players in acquisition of cisplatin resistant phenotype. Knockdown of VIRMA led to disruption of the remaining methyltransferase complex and decrease in m 6 A abundance, as well as overall reduced tumor aggressiveness (with decreased cell viability, tumor cell proliferation, migration, and invasion) and increased sensitivity to cisplatin treatment, both in vitro and confirmed in vivo. Enhanced response to cisplatin after VIRMA knockdown was related to significant increase in DNA damage (with higher γH2AX and GADD45B levels) and downregulation of XLF and MRE11., Conclusions: VIRMA has an oncogenic role in GCTs confirming our previous tissue-based study and is further involved in response to cisplatin by interfering with DNA repair. These data contribute to our better understanding of the emergence of cisplatin resistance in GCTs and support recent attempts to therapeutically target elements of the m 6 A writer complex., (© 2021. The Author(s).)

Hypobaric hypoxia (HH) is a typical characteristic of high altitude environment and causes a spectrum of pathophysiological effects, including headaches, gliovascular dysfunction and cognitive retardation. Here, we sought to understand the mechanisms underlying cognitive deficits under HH exposure. Our results showed that hypobaric hypoxia exposure impaired cognitive function and suppressed dendritic spine density accompanied with increased neck length in both basal and apical hippocampal CA1 region neurons in mice. The expression of PSD95, a vital synaptic scaffolding molecule, is down-regulated by hypobaric hypoxia exposure and post-transcriptionally regulated by cold-inducible RNA-binding protein (Cirbp) through 3'-UTR region binding. PSD95 expressing alleviates hypoxia-induced dendritic spine morphology changes of hippocampal neurons and memory deterioration. Moreover, overexpressed Cirbp in hippocampus rescues HH-induced abnormal expression of PSD95 and attenuates hypoxia-induced dendritic spine injury and cognitive retardation. Thus, our findings reveal a novel mechanism that Cirbp-PSD-95 axis appears to play an essential role in HH-induced cognitive dysfunction in mice., (© 2021. The Author(s).)

ErbB3-binding protein 1 (EBP1) is a multifunctional protein associated with neural development. Loss of Ebp1 leads to upregulation of the gene silencing unit suppressor of variegation 3-9 homolog 1 (Suv39H1)/DNA (cytosine 5)-methyltransferase (DNMT1). EBP1 directly binds to the promoter region of DNMT1, repressing DNA methylation, and hence, promoting neural development. In the current study, we showed that EBP1 suppresses histone methyltransferase activity of Suv39H1 by promoting ubiquitin-proteasome system (UPS)-dependent degradation of Suv39H1. In addition, we showed that EBP1 directly interacts with Suv39H1, and this interaction is required for recruiting the E3 ligase MDM2 for Suv39H1 degradation. Thus, our findings suggest that EBP1 regulates UPS-dependent degradation of Suv39H1 to govern proper heterochromatin assembly during neural development. [BMB Reports 2021; 54(8): 413-418].

Circular RNAs (circRNAs) are highly expressed in the brain and their expression increases during neuronal differentiation. The factors regulating circRNAs in the developing mouse brain are unknown. NOVA1 and NOVA2 are neural-enriched RNA-binding proteins with well-characterized roles in alternative splicing. Profiling of circRNAs from RNA-seq data revealed that global circRNA levels were reduced in embryonic cortex of Nova2 but not Nova1 knockout mice. Analysis of isolated inhibitory and excitatory cortical neurons lacking NOVA2 revealed an even more dramatic reduction of circRNAs and establishes a widespread role for NOVA2 in enhancing circRNA biogenesis. To investigate the cis-elements controlling NOVA2-regulation of circRNA biogenesis, we generated a backsplicing reporter based on the Efnb2 gene. We found that NOVA2-mediated backsplicing of circEfnb2 was impaired when YCAY clusters located in flanking introns were mutagenized. CLIP (cross-linking and immunoprecipitation) and additional reporter analyses demonstrated the importance of NOVA2 binding sites located in both flanking introns of circRNA loci. NOVA2 is the first RNA-binding protein identified to globally promote circRNA biogenesis in the developing brain., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Reproductive transition of grasses is characterized by switching the pattern of lateral branches, featuring the suppression of outgrowth of the subtending leaves (bracts) and rapid formation of higher-order branches in the inflorescence (panicle). However, the molecular mechanisms underlying such changes remain largely unknown. Here, we show that bract suppression is required for the reproductive branching in rice. We identified a pathway involving the intrinsic time ruler microRNA156/529, their targets SQUAMOSA PROMOTER BINDING PROTEIN LIKE (SPL) genes, NECK LEAF1 (NL1), and PLASTOCHRON1 (PLA1), which regulates the bract outgrowth and thus affects the pattern switch between vegetative and reproductive branching. Suppression of the bract results in global reprogramming of transcriptome and chromatin accessibility following the reproductive transition, while these processes are largely dysregulated in the mutants of these genes. These discoveries contribute to our understanding of the dynamic plant architecture and provide novel insights for improving crop yields., (Copyright © 2021 The Author. Published by Elsevier Inc. All rights reserved.)

Post-transcriptional regulation is involved in the regulation of many inflammatory genes. Zinc finger protein 36 (ZFP36) family proteins are RNA-binding proteins involved in messenger RNA (mRNA) metabolism pathways. The ZFP36 family is composed of ZFP36 (also known as tristetraprolin, TTP), ZFP36L1, ZFP36L2, and ZFP36L3 (only in rodents). The ZFP36 family proteins contain two tandemly repeated CCCH-type zinc-finger motifs, bind to adenine uridine-rich elements in the 3'-untranslated regions (3' UTR) of specific mRNA, and lead to target mRNA decay. Although the ZFP36 family members are structurally similar, they are known to play distinct functions and regulate different target mRNAs, probably due to their cell-type-specific expression patterns. For instance, ZFP36 has been well-known to function as an anti-inflammatory modulator in murine models of systemic inflammatory diseases by down-regulating the production of various pro-inflammatory cytokines, including TNF-α. Meanwhile, ZFP36L1 is required for the maintenance of the marginal-zone B cell compartment. Recently, we found that ZFP36L2 reduces the expression of  Ikzf2  (encoding HELIOS) and suppresses regulatory T cell function. This review summarizes the current understanding of the post-transcriptional regulation of immunological responses and inflammatory diseases by RNA-binding ZFP36 family proteins., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Makita, Takatori and Nakajima.)

Purpose: This study aimed to illustrate the biological role of hsa&#95;circ&#95;0005721 in the development of osteosarcoma and the molecular mechanism., Methods: hsa&#95;circ&#95;0005721 levels in 30 pairs of osteosarcoma and non-tumor tissues were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Functional experiments were conducted to assess the influence of hsa&#95;circ&#95;0005721 on proliferative, metastatic and apoptotic rates of osteosarcoma cells. The downstream target of hsa&#95;circ&#95;0005721 and their co-regulatory mechanism in malignant development of osteosarcoma were analyzed by dual-luciferase reporter assay and rescue experiments, respectively., Results: hsa&#95;circ&#95;0005721 was upregulated in osteosarcoma tissues and cell lines. Knockdown of hsa&#95;circ&#95;0005721 suppressed proliferative and metastatic rates of U-2OS and Saos-2 cells, and stimulated apoptosis. Serving as a ceRNA, hsa&#95;circ&#95;0005721 upregulated the linear transcript TEP1 by competitively binding miR-16-5p, thus exerting its biological functions in regulating osteosarcoma development., Conclusions: This study for the first time identified the upregulated hsa&#95;circ&#95;0005721 in osteosarcoma, which triggers the malignant development of osteosarcoma by upregulating the linear transcript TEP1.