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This study aimed to evaluate the effect of miR-10b on growth hormone (GH) in pituitary cells of Yanbian yellow cattle. According to analysis of GH and somatostatin receptor 2 (SSTR2) mRNA and protein expression levels, we found that miR-10b targeted 3'UTR of SSTR2. Compared with the negative control (NC) group, GH mRNA transcription and protein expression in pituitary cells of Yanbian yellow cattle were significantly increased by adding miR-10b mimics (p < .01), while these were significantly decreased by adding miR-10b inhibitor (p < .05); compared with the NC group, SSTR2 mRNA transcription and protein expression were significantly inhibited by the addition of miR-10b mimics (p < .01), while these were significantly increased by the addition of miR-10b inhibitor compared with the iNC group (p < .05). This study suggested that miR-10b could regulate GH level by regulating SSTR2 gene expression in pituitary cells of Yanbian yellow cattle., (© 2020 Japanese Society of Animal Science.)

Mutations in the hairless (Hr) gene in both mice and humans have been implicated in the development of congenital atrichia, but the role of Hr in skin and hair follicle (HF) biology remains unknown. Here, we established transgenic mice (TG) overexpressing mutant Hr to investigate its specific role in the development of HF. Three transgenic lines were successfully constructed, and two of them (TG3 and TG8) displayed a pattern of hair loss and regrowth with alternation in the expression of HR protein. The mutant Hr gene inhibited the expression of the endogenous gene in transgenic individuals, which led to the development of alopecia. Interestingly, the hair regrew with the increase in the endogenous expression levels resulting from decreased mutant Hr expression. The findings of our study indicate that the changes in the expression of Hr result in hair loss or regrowth.

The mechanism underlying autophagy in paroxysmal nocturnal hemoglobinuria (PNH) remains largely unknown. We previously sequenced the entire genome exon of the CD59– cells from 13 patients with PNH and found genes such as CUX1 encoding Cut-like homeobox 1. Peripheral blood samples from 9 patients with PNH and 7 healthy control subjects were obtained to measure CUX1 expression. The correlation between CUX1 messenger RNA expression and PNH clinical indicators was analyzed. To simulate CUX1 expression in patients with PNH, we generated a panel of PNH cell lines by knocking out PIGA in K562 cell lines and transfected lentivirus with CUX1. CCK-8 and EDU assay assessed cell proliferation. Western blotting was used to detect Beclin-1, LC3A, LC3B, ULK1, PI3K, AKT, p-AKT, mTOR, and p-mTOR protein levels. Autophagosomes were observed with transmission electron microscopy. Chloroquine was used to observe CUX1 expression in PNH after autophagy inhibition. Leukocytes from patients with PNH had lower levels of CUX1 messenger RNA expression and protein content than healthy control subjects. The lactose dehydrogenase level and the percentage of PNH clones were negatively correlated with CUX1 relative expression. We reduced CUX1 expression in a PIGA knockout K562 cell line, leading to increased cell proliferation. Levels of autophagy markers Beclin-1, LC3B, LC3A, and ULK1 increased, and autophagosomes increased. Furthermore, PI3K/AKT/mTOR protein phosphorylation levels were lower. CUX1 expression did not change and cell proliferation decreased in CUX1 knocked down PNH cells after inhibition of autophagy by chloroquine. In brief, CUX1 loss-of-function mutation resulted in stronger autophagy in PNH. [ABSTRACT FROM AUTHOR]

Introduction: Ovarian aging is characterized by a gradual decline in quantity and quality of oocytes and lower chance of fertility. Better understanding the genetic modulation during ovarian aging can further address available treatment options for aging-related ovarian diseases and fertility preservation. Methods: A novel technique spatial transcriptomics (ST) was used to investigate the spatial transcriptome features of rat ovaries. Transcriptomes from ST spots in the young and aged ovaries were clustered using differentially expressed genes. These data were analyzed to determine the spatial organization of age-induced heterogeneity and potential mechanisms underlying ovarian aging. Results: In this study, ST technology was applied to profile the comprehensive spatial imaging in young and aged rat ovary. Fifteen ovarian cell clusters with distinct gene-expression signatures were identified. The gene expression dynamics of granulosa cell clusters revealed three sub-types with sequential developmental stages. Aged ovary showed a significant decrease in the number of granulosa cells from the antral follicle. Besides, a remarkable rearrangement of interstitial gland cells was detected in aging ovary. Further analysis of aging-associated transcriptional changes revealed that the disturbance of oxidative pathway was a crucial factor in ovarian aging. Conclusions: This study firstly described an aging-related spatial transcriptome changes in ovary and identified the potential targets for prevention of ovarian aging. These data may provide the basis for further investigations of the diagnosis and treatment of aging-related ovarian disorders. [ABSTRACT FROM AUTHOR]

Depsides and depsidones have attracted attention for biosynthetic studies due to their broad biological activities and structural diversity. Previous structure‒activity relationships indicated that triple halogenated depsidones display the best anti-pathogenic activity. However, the gene cluster and the tailoring steps responsible for halogenated depsidone nornidulin (3) remain enigmatic. In this study, we disclosed the complete biosynthetic pathway of the halogenated depsidone through in vivo gene disruption, heterologous expression and in vitro biochemical experiments. We demonstrated an unusual depside skeleton biosynthesis process mediated by both highly-reducing polyketide synthase and non-reducing polyketide synthase, which is distinct from the common depside skeleton biosynthesis. This skeleton was subsequently modified by two in-cluster enzymes DepG and DepF for the ether bond formation and decarboxylation, respectively. In addition, the decarboxylase DepF exhibited substrate promiscuity for different scaffold substrates. Finally, and interestingly, we discovered a halogenase encoded remotely from the biosynthetic gene cluster, which catalyzes triple-halogenation to produce the active end product nornidulin (3). These discoveries provide new insights for further understanding the biosynthesis of depsidones and their derivatives. A new and concise biosynthetic pathway of nornidulin (3) was elucidated. The depside skeleton 2 is synthesized by a collaborating hrPKS and nrPKS pair, followed by etherification, decarboxylation and halogenation. [Display omitted] [ABSTRACT FROM AUTHOR]

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare clonal disease of hematopoietic stem cells (HSCs). Long noncoding RNAs (lncRNAs) perform a wide range of biological functions, including the regulation of gene expression, cell differentiation, and proliferation, but their role in PNH remains unclear. CD59− and CD59+ granulocytes and monocytes from 35 PNH patients were sorted. High-throughput sequencing was analyzed in 5 PNH patients, and differentially expressed lncRNAs and mRNAs were identified. The mRNAs with fragments per kilobase of exon model per million mapped fragments (FPKM) > 10 in at least 3 patients were selected, and experiments were performed to identify their upstream regulatory lncRNAs. The expression of selected mRNAs and lncRNAs was verified by qRT‒PCR, and the correlation of these expression patterns with clinical data from other 30 PNH patients was analyzed. Then, the functions of the lncRNAs were studied in the PIGA-KO-THP-1 cell line. Transcription analysis revealed 742 upregulated and 1376 downregulated lncRNAs and 3276 upregulated and 213 downregulated mRNAs. After deep screening, 8 highly expressed mRNAs that were related to the NF-κB pathway were analyzed to determine coexpression patterns. LINC01002, FAM157C, CTD-2530H12.2, XLOC-064331 and XLOC-106677 were correlated with the 8 mRNAs. After measuring the expression of these molecules in 30 PNH patients by qRT‒PCR, lncRNA FAM157C was verified to be upregulated in the PNH clone, and its expression levels were positively correlated with the LDH levels and CD59− granulated and monocyte cell ratios. After knockdown of the FAM157C gene in the PIGA-KO-THP-1 cell line, we found that the cells were arrested in the G0/G1 phase and S phase, the apoptosis rate increased, and the cell proliferation decreased. LncRNA FAM157C was proven to promote PNH clone proliferation, and this is the first study to explore the role of lncRNAs in PNH. [ABSTRACT FROM AUTHOR]

Paroxysmal nocturnal hemoglobinuria (PNH) is a disease involving hematopoietic stem cell membrane defects caused by acquired phosphatidylinositol glycan anchor biosynthesis class A (PIGA) mutations. In this study, 97 target genes were selected as a target gene panel and screened in 23 PNH patients via the sequencing of specific DNA target regions. Through functional analysis, we identified that suppressor‐of‐Zeste 12 (SUZ12) may be involved in the proliferation of PNH clones. mRNA and protein expression levels of SUZ12 and the trimethylation level of histone H3 at lysine 27 (H3K27) in CD59– peripheral blood leukocytes from PNH patients were higher than those in CD59+ cells from PNH patients and peripheral blood leukocytes from healthy controls. In addition, the relative expression of SUZ12 in PNH patients was positively correlated with Ret% and the proportion of PNH clones. When we knocked down SUZ12 expression in a PIGA knockdown THP‐1 cell line (THP‐1 KD cells), the trimethylation of histone H3K27(H3K27me3) and cell proliferation decreased, apoptosis increased, and cell cycle arrest occurred in G0/G1 phase. In conclusion, SUZ12 participates in the proliferation of PNH clones by regulating histone H3K27me3 levels. Our results may provide new therapeutic targets and possibilities for PNH patients. [ABSTRACT FROM AUTHOR]

Background: Eucalyptus is a highly diverse genus of the Myrtaceae family and widely planted in the world for timber and pulp production. Tissue culture induced callus has become a common tool for Eucalyptus breeding, however, our knowledge about the genes related to the callus maturation and shoot regeneration is still poor. Results: We set up an experiment to monitor the callus induction and callus development of two Eucalyptus species - E. camaldulensis (high embryogenic potential) and E. grandis x urophylla (low embryogenic potential). Then, we performed transcriptome sequencing for primary callus, mature callus, shoot regeneration stage callus and senescence callus. We identified 707 upregulated and 694 downregulated genes during the maturation process of the two Eucalyptus species and most of them were involved in the signaling pathways like plant hormone and MAPK. Next, we identified 135 and 142 genes that might play important roles during the callus development of E. camaldulensis and E. grandis x urophylla, respectively. Further, we found 15 DEGs shared by these two Eucalyptus species during the callus development, including Eucgr.D00640 (stem-specific protein TSJT1), Eucgr.B00171 (BTB/POZ and TAZ domain-containing protein 1), Eucgr.C00948 (zinc finger CCCH domain-containing protein 20), Eucgr.K01667 (stomatal closure-related actinbinding protein 3), Eucgr.C00663 (glutaredoxin-C10) and Eucgr.C00419 (UPF0481 protein At3g47200). Interestingly, the expression patterns of these genes displayed "N" shape in the samples. Further, we found 51 genes that were dysregulated during the callus development of E. camaldulensis but without changes in E. grandis x urophylla, such as Eucgr.B02127 (GRF1-interacting factor 1), Eucgr.C00947 (transcription factor MYB36), Eucgr.B02752 (laccase-7), Eucgr.B03985 (transcription factor MYB108), Eucgr.D00536 (GDSL esterase/lipase At5g45920) and Eucgr.B02347 (scarecrow-like protein 34). These 51 genes might be associated with the high propagation ability of Eucalyptus and 22 might be induced after the dedifferentiation. Last, we performed WGCNA to identify the co-expressed genes during the callus development of Eucalyptus and qRT-PCR experiment to validate the gene expression patterns. Conclusions: This is the first time to globally study the gene profiles during the callus development of Eucalyptus. The results will improve our understanding of gene regulation and molecular mechanisms in the callus maturation and shoot regeneration. [ABSTRACT FROM AUTHOR]

Background: Puerarin, an active compound of radix puerariae, is a major compound used in Chinese herbal medicines and it has been well known for its pharmacological effects, including antioxidant, anti‑inflammatory, neuroprotective and cardioprotective properties. The aim of the present study was to determine the role of puerarin (Pue) in complete Freund's adjuvant (CFA)-induced trigeminal neuralgia (TN) and the effects of this compound on Sirt1 activity and on the progression of CFA-induced TN. Methods: Mice were injected with CFA on the unilateral face to induce TN. A cell model of inflammation-associated TN was established by interleukin-1β (IL-1β; 10 ng/mL) and tumor necrosis factor-α (TNF-α; 50 ng/mL) stimulation of neurons. Reverse transcription-quantitative PCR and Western blot analyses were performed to analyze mRNA and protein expression levels in trigeminal ganglion and nerve cells. Terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) staining was used to determine nerve cell apoptosis following IL-1β/TNF-α or Pue treatment. Results: Pue is a conceivable Sirtuin1 (Sirt1) activator used for the prevention of trigeminal nerve injury that attenuates CFA-induced TN and inflammatory cytokine-evoked overactivation of neuronal inflammation and apoptosis. Treatment of mice with inflammatory cytokines induced upregulation of cleaved caspase-3 protein expression, which was neutralized by Pue supplementation. Both in vivo and in vitro experiments led to the conclusion that Pue modulated Sirt1 activation and repressed transforming growth factor-β 1 (TGF-β 1) protein expression and drosophila mothers against decapentaplegic homolog3 (Smad3) phosphorylation in order to exert neuroprotection. Conclusion: The findings suggested that Pue functioned as a potential Sirt1 activator to improve neuroinflammation-induced TN and neuronal apoptosis via the suppression of TGF-β 1/Smad3 activity. The pharmacological activity of Pue provides a new perspective for the effective prevention and treatment of TN. [ABSTRACT FROM AUTHOR]

Introduction: Severe aplastic anemia (SAA) is a disease characterized by severe pancytopenia and hematopoietic failure of bone marrow. Natural killer (NK) cells are a class of large granular lymphocytes that perform killing and immunomodulatory functions. Our previous study demonstrated that NK cells played the "protective" role in SAA, which is weakened. However, the mechanism remains unclear. Methods: Peripheral blood NK cells from SAA patients and normal controls were sorted and total proteins were extracted. Then, mass spectrometry was performed to screen differentially expressed proteins (DEPs). Results: Significant differences in the expression levels of 93 proteins were observed in NK cells of SAA patients compared with normal controls. Among them, 48 were upregulated proteins, including histone H1.2, histone H1.3, heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1), and interferon regulatory factor 1 (IRF‐1), and 45 were downregulated proteins, including actin‐related complex (ARP2/3), histone H3, histone H4, phosphoglycerate kinase 1 (PGK1), talin‐1. Gene Ontology (GO) function indicated that the DEPs most involved were vesicle‐mediated transport, innate immune response, and DNA binding. KEGG analysis showed 3 upregulated and 12 downregulated pathways, in which cell endocytosis and FC‐γ receptor‐mediated phagocytosis were most closely related to NK cell functions. Conclusion: Our study is the first analysis of proteomic profile in NK cells in SAA and found many DEPs involving in dysfunction of NK cells, which provides potential targets for deeper research of inadequate immunomodulation. [ABSTRACT FROM AUTHOR]

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease and a severe form of pulmonary fibrosis. Epithelial-mesenchymal transition (EMT) of alveolar epithelial cells is induced in response to epithelial injury, which leads to the accumulation of extracellular matrix in the lung parenchyma and contributes to pulmonary fibrosis. NPAS2 (neuronal PAS domain protein 2) is significantly increased in the lung tissues of IPF patients according to microarray dataset GSE10667 and NPAS2 is downregulated in differentiated human pulmonary type 2 epithelial cells in vitro based on microarray dataset GSE3306 from Gene Expression Omnibus (GEO). In this study, we demonstrated that NPAS2 was increased in bleomycin (BLM)- induced fibrotic lungs in mice. Knockdown of NPAS2 inhibited EMT in primary mouse lung alveolar type 2 epithelial (pmATII) cells and human lung alveolar type 2 epithelial cell line A549 cells under BLM challenge in vitro. Moreover, the silence of NPAS2 alleviated the BLM-induced pulmonary fibrosis in a murine model. Mechanistically, NPAS2 promotes EMT through positively regulating hairy and enhancer of split 1 (HES1) expression. In this study, we present novel findings that have not been previously reported, emphasizing that p53 transcriptionally activates NPAS2 in ATII cells and overexpression of NPAS2 weakens the effects of TP53 knockdown on EMT of pmATII and A549 cells. Our results suggest NPAS2 is a novel target gene of p53 in regulating BLM-mediated EMT in ATII cells and pulmonary fibrosis. [Display omitted] • NPAS2 is upregulated in the lung tissues of IPF patients. • NPAS2 is increased in the bleomycin (BLM)- induced fibrotic lungs in mice. • NPAS2 is a novel target gene of p53 in regulating BLM-induced EMT in ATII cells. [ABSTRACT FROM AUTHOR]

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired clonal proliferative disease of hematopoietic stem cells. Various gene mutations, including the phosphatidylinositol glycan anchor biosynthesis class A (PIG-A) gene, may contribute to the proliferation of PNH clones. In order to explore the mechanism of PNH clone proliferation, a study was performed on 13 patients with PNH who underwent whole exome sequencing. The frequency of mutations in these patients was explored, and an additional 30 patients with PNH were selected for analysis of cluster of differentiation 59-negative (CD59−) cells. The mRNA expression of 13 genes, which were selected based on their high frequency in patients with PNH and the fact that they met four screening conditions, was determined in these CD59− cells. Cell proliferation, apoptosis and cell cycle were evaluated upon knocking down the recombinant signal binding protein of immunoglobulin κJ region (RBPJ) gene in 5 patients in vitro. The detection rate of PIG-A gene mutation was 61.54% (8/13), and additional mutations in somatic genes were detected, including RBPJ, zinc finger protein 717, polycomb repressive complex 2 subunit and tet methylcytosine dioxygenase. Upon screening according to the mutation frequency and expression level, the present study focused on the RBPJ gene. The expression level of RBPJ in CD59− cells was apparently higher than that in CD59+ cells and normal controls which was significantly correlated with clinical data. Furthermore, the expression of RBPJ in PNH primary cells could be effectively inhibited by small interfering RNA-RBPJ. Once the expression of RBPJ decreased remarkably, the apoptotic rate increased gradually, while cell proliferation activity decreased with transfection time and cells were blocked in G0/G1 phase. In conclusion, mutations and abnormal expression of the RBPJ gene may participate in the abnormal proliferation of PNH clones. [ABSTRACT FROM AUTHOR]

Breast cancer is the second leading cause of cancer-associated mortality in women worldwide. Strong evidence has suggested that Ran, which is a small GTP binding protein involved in the transport of RNA and protein across the nucleus, may be a key cellular protein involved in the metastatic progression of cancer. The present study investigated Ran gene expression in breast cancer tissue samples obtained from 140 patients who had undergone surgical resection for breast cancer. Western blot analysis of Ran in breast cancer tissues and paired adjacent normal tissues showed that expression of Ran was significantly increased in breast cancer tissues. Immunohistochemistry analyses conducted on formalin-fixed paraffin-embedded breast cancer tissue sections revealed that Ran expression was associated with tumor histological grade, nerve invasion and metastasis, vascular metastasis and Ki-67 expression (a marker of cell proliferation). Kaplan-Meier survival analysis showed that increased Ran expression in patients with breast cancer was positively associated with a poor survival prognosis. Furthermore, in vitro experiments demonstrated that highly migratory MDA-MB-231 cancer cells treated with Ran-si-RNA (si-Ran), which knocked down expression of Ran, exhibited decreased motility in trans-well migration and wound healing assays. Cell cycle analysis of Ran knocked down MDA-MB-231 cells implicated Ran in cell cycle arrest and the inhibition of proliferation. Furthermore, a starvation and re-feeding (CCK-8) assay was performed, which indicated that Ran regulated breast cancer cell proliferation. Taken together, the results provide strong in vitro evidence of the involvement of Ran in the progression of breast cancer and suggest that it could have high potential as a therapeutic target and/or marker of disease. [ABSTRACT FROM AUTHOR]

The flowering time of soybean is a highly important agronomic characteristic, which affects the adaptability and yield. AtMRF1 , a MORN-repeat motif gene, acts as a floral promoter in Arabidopsis , its functions in soybean are not yet understood. Here, we employed qRT-PCR to analyze the tissue expression patten of MRF1 homologs in soybean and determined that the GmMRF2 gene, containing a MORN-motif, highly expressed in the shoot and responded to photoperiod. GmMRF2 overexpression soybean lines exhibited earlier flowering time under long-day (LD) conditions, and increased plant height under both LD and short-day (SD) conditions compared to wild-type (WT) plants. The expression levels of gibberellic acid (GA) pathway genes that positively regulate plant height genes and flowering-promoting genes were up-regulated in the GmMRF2 overexpression lines, were up-regulated in the GmMRF2 overexpression lines. Further study revealed that Gm MRF2 interacted with GmTCP15 to co-induce the expression of GmSOC1b. Together, our results preliminarily reveal the functions and mechanisms of GmMRF2 in regulating flowering time and plant height, provide a new promising gene for soybean crop improvement. • GmMRF2 overexpression promoted flowering and increased plant height in soybean. • The expression level of GA pathway and flowering related genes was alerted in the GmMRF2 overexpression lines. • GmMRF2 interacted with GmTCP15 to co-induce the expression of GmSOC1b to accelerate soybean flowering. [ABSTRACT FROM AUTHOR]

SOX17, a member of SRY-related high-mobility-group box (SOX) family, involves in endoderm formation, angiogenesis and carcinogenesis, and its expression characteristics are different in spermatogenesis among several vertebrates. In this study, we cloned a full-length cDNA sequence of sox17 from Zhikong scallop Chlamys farreri , and determined its expression characteristics in gonad at mRNA and protein levels. The cDNA sequence was 2802 bp in length, predicted to encode a protein of 511 amino acids and contained a conservative HMG-box of SOX family, while lacked the C-terminal region of SOX17 comparing to vertebrates. Semi-quantitative RT-PCR showed that C. farreri sox17 ( Cf-sox17 ) mRNA exhibited a different tissue distribution, and the transcript abundance was the highest in the gonads. In situ hybridization determined that the Cf-sox17 mRNA was located in various germ cells in testis and ovary. Similar result of Cf- SOX17 protein was also observed by immunohistochemical detection. The location in gonad is different from that of mammals and fish in which SOX17 is only located in some specific germ cells. Our finding revealed a different characteristic of sox17 expression in gametogenesis between scallop and vertebrates, which implied that Cf-sox17 may involve in gametogenesis of bivalves and the function may differ from that in vertebrates. [ABSTRACT FROM AUTHOR]