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Gonadotropin-releasing hormone (GnRH) plays an important role in regulating the activities of other components downstream of the hypothalamic-pituitary-gonadal (HPG) axis and maintaining the normal reproductive cycle of animals. However, the molecular mechanisms by which GnRH synthesis and secretion are regulated in sheep remains unclear. In this study, a series of eight recombinant vectors with deletion fragments were constructed and cotransfected with pGL3-Basic and pRL-SV40 into sheep hypothalamic neuronal cells. After treatment with 1 nM kisspeptin, the core promoter of the sheep GnRH gene was identified to be in the region of -1912 bp to -1461 bp by dual-luciferase reporter assay. Bioinformatics analysis showed that there was a binding site for the transcription factor Otx-2 in the core promoter region (-1786 to -1770 bp) that was highly conserved among different species. The expression patterns of Kiss-1, Otx-2 and GnRH in the sheep hypothalamus were the same, and the expression of Kiss-1, Otx-2 and GnRH was significantly higher in the breeding season than in nonbreeding season (P < 0.01). In addition, when hypothalamic neurons were cultured in vitro with kisspeptin, kisspeptin induced the expression of GnRH and Otx-2. In conclusion, these results provide evidence that the core promoter region (-1786 to -1770 bp) of the GnRH gene is involved in the regulation of hypothalamic activity by kisspeptin and that binding of the transcription factor Otx-2 mediates this activation., (Copyright © 2019 Elsevier B.V. All rights reserved.)

In this study, we detect depressive effect on leptin receptor (LEPR) by LEPR-specific short hairpin RNA (shRNA) expression plasmids in chicken preadipocytes, and effect on messenger RNA (mRNA) expression levels of genes related to signal transduction, including JAK2, STAT3, SOCS3 as well as CPT1, which is associated with fatty acid metabolism. shRNA expression vectors targeting LEPR were constructed and transfected into chicken preadipocytes. The transfection efficiency was evaluated by fluorescence microscopy. Real-time PCR was used to detect its effect on mRNA expression levels of JAK2, STAT3, SOCS3 and CPT1. Results showed that LEPR mRNA was knocked down by 99% (P < 0.01) after transfection for 72 h. In the knockdown preadipocytes, the mRNA levels of JAK2 and CPT1 were down-regulated by 47.56% (P < 0.01) and 42.26% (P < 0.05), respectively; while expression of STAT3 and SOCS3 increased 7.72-fold (P < 0.01), 1.71-fold (P < 0.01), respectively. It is concluded that knockdown of LEPR influences mRNA expression of its down-stream genes, suggesting that chicken LEPR play a certain role in regulating genes in the complicated gene network of preadipocytes., (© 2016 Japanese Society of Animal Science.)

Pinellia pedatisecta agglutinin (PPA)is a very basic protein that accumulates in the tuber of P.pedatisecta .PPA is a hetero-tetramer protein of 40 kDa,composed of two polypeptide chains A (about 12 kDa)and two polypeptides chains B (about 12 kDa).The full-length cDNA of PPA was cloned from P.pedatisecta using SMART RACE-PCR technology; it was 1146 bp and contained a 771 bp open reading frame (ORF)encoding a lectin precursor of 256 amino acid residues with a 24 amino acid signal peptide.The PPA precursor contained 3 mannose-binding sites (QXDXNXVXY) and two conserved domains of 43% identity,PPA-DOM 1 (polypeptides A)and PPA-DOM 2 (polypeptides B).PPA shared varying identities,ranging from 40% to 85%,with mannose-binding lectins from other species of plant families such as Araceae, Alliaceae, Iridaceae, Liliaceae, Amaryllidaceae and Bromeliaceae. Southern blot analysis indicated that ppa belonged to a multi-copy gene family. Expression pattern analysis revealed that ppa expressed in most tested tissues, with high expression being found in spadix,spathe and tuber.Cloning of the ppa gene not only provides a basis for further investigation of its structure,expression and regulatory mechanism,but also enables us to test its potential role in controlling pests and fungal diseases by transferring the gene into plants in the future.

Using RNA extracted from Zingiber officinale rhizomes and primers designed according to the conservative regions of monocot mannose-binding lectins, the full-length cDNA of Z. officinale agglutinin (ZOA) was cloned by rapid amplification of cDNA ends (RACE). The full-length cDNA of zoa was 746 bp and contained a 510 bp open reading frame (ORF) encoding a lectin precursor of 169 amino acids with a signal peptide. ZOA was a mannose-binding lectin with three typical mannose-binding sites (QDNY). Semi-quantitative RT-PCR analysis revealed that zoa expressed in all the tested tissues of Z. officinale including leaf, root and rhizome, suggesting it to be a constitutively expressing form. ZOA protein was successfully expressed in Escherichia coli with the molecular weight expected. To our knowledge, this is the first mannose-binding lectin cDNA cloned from the family Zingiberaceae. Our results demonstrate that monocot mannose-binding lectins also occur within the family Zingiberaceae.

Myoblast proliferation and differentiation are essential for skeletal muscle development. In this study, we generated the expression profiles of mRNAs, long noncoding RNAs (lncRNAs), and microRNAs (miRNAs) in different developmental stages of chicken primary myoblasts (CPMs) using RNA sequencing (RNA-seq) technology. The dual luciferase reporter system was performed using chicken embryonic fibroblast cells (DF-1), and functional studies quantitative real-time polymerase chain reaction (qPCR), cell counting kit-8 (CCK-8), 5-Ethynyl-2′-deoxyuridine (EdU), flow cytometry cycle, RNA fluorescence in situ hybridization (RNA-FISH), immunofluorescence, and western blotting assay. Our research demonstrated that miR-301a-5p had a targeted binding ability to lncMDP1 and ChaC glutathione-specific gamma-glutamylcyclotransferase 1 (CHAC1). The results revealed that lncMDP1 regulated the proliferation and differentiation of myoblasts via regulating the miR-301a-5p/CHAC1 axis, and CHAC1 promotes muscle regeneration. This study fulfilled the molecular regulatory network of skeletal muscle development and providing an important theoretical reference for the future improvement of chicken meat performance and meat quality. all: "A transcriptome study of chicken primary myoblast proliferation and differentiation shows that the lncMDP1/miRNA-301a-5p/CHAC1 axis plays an important role in skeletal muscle development." [ABSTRACT FROM AUTHOR]

China leads the world in freshwater pearl production, an industry in which the triangle sail mussel (Sinohyriopsis cumingii) plays a pivotal role. In this paper, we report a high-quality chromosome-level genome assembly of S. cumingii with a size of 2.90 Gb—the largest yet reported among bivalves—and 89.92% anchorage onto 19 linkage groups. The assembled genome has 37,696 protein-coding genes and 50.86% repeat elements. A comparative genomic analysis revealed expansions of 752 gene families, mostly associated with biomineralization, and 237 genes under strong positive selection. Notably, the fibrillin gene family exhibited gene family expansion and positive selection simultaneously, and it also exhibited multiple high expressions after mantle implantation by transcriptome analysis. Furthermore, RNA silencing and an in vitro calcium carbonate crystallization assay highlighted the pivotal role played by one fibrillin gene in calcium carbonate deposition and aragonite transformation. This study provides a valuable genomic resource and offers new insights into the mechanism of pearl biomineralization. [ABSTRACT FROM AUTHOR]

Background: The clinical characteristics and efficacy of human epidermal growth factor receptor‐2 (HER‐2)‐directed agents against HER2 mutations and HER2 fusions in breast cancer are obscure due to their low frequency. Methods: We conducted a retrospective study in patients with advanced breast cancer harboring HER2 mutations and/or HER2 fusions between January 1, 2017 and January 1, 2021. Results: Among a total of 22 patients, 17 HER2 mutations were detected, including L755S, S310F, R100=, V777L, R897W, T862A, 440‐17C > G, H878Y, V842I, 73 + 9G > C, T278fs, E1069K, L755P, 226‐11C > T, 574 + 12C>T, L114V and P128L. The majority of patients had ductal carcinoma, which mostly coexisted with HER2 amplification/overexpression. The median progression‐free survival (PFS) of the 22 patients was 6.9 months (95% CI: 4.7, 9.1) in the first‐line setting. The median PFS of patients who received first‐line trastuzumab‐based regimens was significantly longer than that of patients who received a first‐line tyrosine kinase inhibitor (TKI) (10.8 months [95% CI: 2.9, 18.7] vs. 1.9 months [95% CI: 0.8, 3.0], p < 0.005). A total of 14 patients were treated with anti‐HER2 antibody‐drug conjugate (ADC), among whom the median treatment line of first‐time of administration of anti‐HER2 ADC was 4.5 (range, 1–10). Anti‐HER2 ADC reached an objective response rate (ORR) of 42.9%, a disease control rate (DCR) of 85.7% and a median PFS of 7.3 months (95% CI: 4.4–10.1) from the first‐time of administration. Conclusion: Our data demonstrated the clinical benefit of anti‐HER2 treatment in Chinese breast cancer patients harboring HER2 mutation and/or HER2 fusion. The value of immunotherapy and treatment selection among individual HER2 variants needs further study. [ABSTRACT FROM AUTHOR]

Sepsis is a life-threatening condition characterized by a harmful host response to infection with organ dysfunction. Annually about 20 million people are dead owing to sepsis and its mortality rates is as high as 20%. However, no studies have been carried out to investigate sepsis from the system biology point of view, as previous research predominantly focused on individual genes without considering their interactions and associations. Here, we conducted a comprehensive exploration of genome-wide expression alterations in both mRNAs and long non-coding RNAs (lncRNAs) in sepsis, using six microarray datasets. Co-expression networks were conducted to identify mRNA and lncRNA modules, respectively. Comparing these sepsis modules with normal modules, we observed a homogeneous expression pattern within the mRNA/lncRNA members, with the majority of them displaying consistent expression direction. Moreover, we identified consistent modules across diverse datasets, consisting of 20 common mRNA members and two lncRNAs, namely CHRM3-AS2 and PRKCQ-AS1, which are potential regulators of sepsis. Our results reveal that the up-regulated common mRNAs are mainly involved in the processes of neutrophil mediated immunity, while the down-regulated mRNAs and lncRNAs are significantly overrepresented in T-cell mediated immunity functions. This study sheds light on the co-expression patterns of mRNAs and lncRNAs in sepsis, providing a novel perspective and insight into the sepsis transcriptome, which may facilitate the exploration of candidate therapeutic targets and molecular biomarkers for sepsis. [ABSTRACT FROM AUTHOR]

Simple Summary: The study of skeletal muscle growth and development has historically been viewed as an entry point and as a breakthrough event in the improvement of animal breeding in the livestock industry. Muscle is the major constituent of any meat product, and its growth performance and the resulting product quality are the core foci of modern breeding. In this study, we found that creatine kinase M-type-like (CKM) was specifically highly expressed in chicken muscle tissue. The quantitative real-time PCR (qPCR) results were consistent with the sequencing results. By overexpressing CKM and interfering with CKM expression in chicken primary myoblasts, we found that CKM could inhibit the proliferation and promote the apoptosis and differentiation of chicken primary myoblasts. Moreover, the transcriptome sequencing results after interference with CKM expression indicated that CKM is involved in chicken skeletal muscle development. These results suggest that CKM may play a crucial role in chicken muscle growth and development. Skeletal muscle is an essential tissue in meat-producing animals, and meat-producing traits have been a hot topic in chicken genetic breeding research. Current research shows that creatine kinase M-type-like (CKM) is one of the most abundant proteins in skeletal muscle and plays an important role in the growth and development of skeletal muscle, but its role in the development of chicken skeletal muscle is still unclear. Via RNA sequencing (RNA-seq), we found that CKM was highly expressed in chicken breast muscle tissue. In this study, the expression profile of CKM was examined by quantitative real-time PCR (qPCR), and overexpression and RNA interference techniques were used to explore the functions of CKM in the proliferation, apoptosis and differentiation of chicken primary myoblasts (CPMs). It was shown that CKM was specifically highly expressed in breast muscle and leg muscle and was highly expressed in stage 16 embryonic muscle, while CKM inhibited proliferation, promoted the apoptosis and differentiation of CPMs and was involved in regulating chicken myogenesis. Transcriptome sequencing was used to identify genes that were differentially expressed in CPMs after CKM disruption, and bioinformatics analysis showed that CKM was involved in regulating chicken myogenesis. In summary, CKM plays an important role in skeletal muscle development during chicken growth and development. [ABSTRACT FROM AUTHOR]

A growing number of studies have shown that members of the ankyrin repeat and suppressors of cytokine signaling (SOCS) box-containing protein (ASB) family are extensively involved in biological processes such as cell growth, tissue development, insulin signaling, ubiquitination, protein degradation, and skeletal muscle membrane protein formation, while the specific biological role of ankyrin-repeat and SOCS box protein 9 (ASB9) remains unclear. In this study, a 21 bp indel in the intron of ASB9 was identified for the first time in 2641 individuals from 11 different breeds and an F2 resource population, and differences were observed among individuals with different genotypes (II, ID, and DD). An association study of a cross-designed F2 resource population revealed that the 21-bp indel was significantly related to growth and carcass traits. The significantly associated growth traits were body weight (BW) at 4, 6, 8, 10, and 12 weeks of age; sternal length (SL) at 4, 8, and 12 weeks of age; body slope length (BSL) at 4, 8, and 12 weeks of age; shank girth (SG) at 4 and 12 weeks of age; tibia length (TL) at 12 weeks of age; and pelvic width (PW) at 4 weeks of age (p < 0.05). This indel was also significantly correlated with carcass traits including semievisceration weight (SEW), evisceration weight (EW), claw weight (CLW), breast muscle weight (BMW), leg weight (LeW), leg muscle weight (LMW), claw rate (CLR), and shedding weight (ShW) (p < 0.05). In commercial broilers, the II genotype was the dominant genotype and underwent extensive selection. Interestingly, the ASB9 gene was expressed at significantly higher levels in the leg muscles of Arbor Acres broilers than those of Lushi chickens, while the opposite was true for the breast muscles. In summary, the 21-bp indel in the ASB9 gene significantly influenced the expression of the ASB9 gene in muscle tissue and was associated with multiple growth and carcass traits in the F2 resource population. These findings suggested that the 21-bp indel within the ASB9 gene could be used in marker-assisted selection breeding for traits related to chicken growth. [ABSTRACT FROM AUTHOR]

Vestigial-like (Vgll) genes are widespread in vertebrates and play an important role in muscle development. In this study, we used bioinformatics methods to systematically identify the chicken VGLL family in the whole genome and investigated its evolutionary history and gene structure features. Tissue expression spectra combined with real-time PCR data were used to analyze the organizational expression pattern of the genes. Based on the maximum likelihood method, a phylogenetic tree of the VGLL family was constructed, and 94 VGLL genes were identified in 24 breeds, among which four VGLL family genes were identified in the chicken genome. Ten motifs were detected in the VGLL genes, and the analysis of introns combined with gene structure revealed that the family was conserved during evolution. Tissue expression analysis suggested that the expression profiles of the VGLL family genes in 16 tissues differed between LU Shi and AA broilers. In addition, a single gene (VGLL2) showed increased expression in chickens at embryonic days 10–16 and was involved in the growth and development of skeletal muscle in chickens in the embryonic stage. In summary, VGLL genes are involved in chicken muscle growth and development, which provides useful information for subsequent functional studies of VGLL genes. [ABSTRACT FROM AUTHOR]

Immune response and reproductive success are two vital energy-consuming processes in living organisms. However, it is still unclear which process is prioritized when both are required. Therefore, the present study was designed to examine this question arising for one of the world’s most destructive agricultural pests, the migratory locust, Locusta migratoria. Transcripts from the ovaries and fat bodies of newly emerged locusts were analyzed, using RNA-seq based transcriptome and qualitative real-time PCR, at 4 h and 6 d after being infected with the gram-positive bacteria Micrococcus luteus. Changes in the main biological pathways involved in reproduction and immunization were analyzed using bioinformatics. After 4 h of infection, 348 and 133 transcripts were up- and down-regulated, respectively, whereas 5699 and 44 transcripts were up- and down-regulated, respectively, at 6 d after infection. Moreover, KEGG analysis indicated that vital pathways related with immunity and reproduction, such as Insulin resistance, FoxO signaling, Lysosome, mTOR signaling, and Toll-like receptor signaling pathways were up-regulated. Among the differentially expressed genes, 22 and 17 were related to immunity and reproduction, respectively. The expression levels of PPO1 and antimicrobial peptide defensin 3 were increased (log2FC = 5.93 and 6.75, respectively), whereas those of VgA and VgB were reduced (log2FC = -17.82 and -18.13, respectively). These results indicated that locust allocate energy and resources to maintain their own survival by increasing immune response when dealing with both immune and reproductive processes. The present study provides the first report of expression levels for genes related with reproduction and immunity in locusts, thereby providing a reference for future studies, as well as theoretical guidance for investigations of locust control. [ABSTRACT FROM AUTHOR]

The formation mechanism underlying the blue eggshell characteristic has been discovered in birds, and SLCO1B3 is the key gene that regulates the blue eggshell color. Insertion of an endogenous retrovirus, EAV-HP, in the SLCO1B3 5′ flanking region promotes SLCO1B3 expression in the chicken shell gland, and this expression causes bile salts to enter the shell gland, where biliverdin is secreted into the eggshell, forming a blue shell. However, at different laying stages of the same group of chickens, the color of the eggshell can vary widely, and the molecular mechanism underlying the eggshell color change remains unknown. Therefore, to reveal the molecular mechanism of the blue eggshell color variations, we analyzed the change in the eggshell color during the laying period. The results indicated that the eggshell color in Lushi chickens can be divided into three stages: 20–25 weeks for dark blue, 26–45 weeks for medium blue, and 46–60 weeks for light blue. We further investigated the expression and methylation levels of the SLCO1B3 gene at eight different weeks, finding that the relative expression of SLCO1B3 was significantly higher at 25 and 30 weeks than at other laying weeks. Furthermore, the overall methylation rate of the SLCO1B3 gene in Lushi chickens increased gradually with increasing weeks of egg production, as shown by bisulfite sequencing PCR. Pearson correlation analysis showed that methylation of the promoter region of SLCO1B3 was significantly negatively correlated with both SLCO1B3 expression in the shell gland tissue and eggshell color. In addition, we predicted that CpG5 and CpG8 may be key sites for regulating SLCO1B3 gene transcription. Our findings show that as the level of methylation increases, methylation of the CpG5 and CpG8 sites hinders the binding of transcription factors to the promoter, reducing SLCO1B3 expression during the late period and resulting in a lighter eggshell color. [ABSTRACT FROM AUTHOR]

The cathepsin E-A-like, also known as ‘similar to nothepsin’, is a new member of the aspartic protease family, which may take part in processing of egg yolk macromolecules, due to it was identified in the chicken egg-yolk. Previously, studies have suggested that the expression of cathepsin E-A-like increased gradually during sexual maturation of pullets, but the exact regulation mechanism is poorly understood. In this study, to gain insight into the function and regulation mechanism of the gene in egg-laying hen, we cloned the cathepsin E-A-like gene and evaluated its evolutionary origin by using both phylogenetic and syntenic methods. The mode of the gene expression regulation was analysed through stimulating juvenile hens with 17β-estradiol and chicken embryo hepatocytes with 17β-estradiol combined with oestrogen receptor antagonists including MPP, ICI 182,780 and tamoxifen. Our results showed that cathepsin E-A-like was an orthologoues gene with nothepsin, which is present in birds but not in mammals. The expression of cathepsin E-A-like significantly increased in a dose-dependent manner after the juvenile hens were treated with 17β-estradiol (P<0.05). Compared with the 17β-estradiol treatment group, the expression of cathepsin E-A-like was not significantly changed when the hepatocytes were treated with 17β-estradiol combined with MPP (P<0.05). In contrast, compared with the 17β-estradiol combined with MPP treatment group, the expression of cathepsin E-A-like was significantly downregulated when the hepatocytes were treated with 17β-estradiol combined with tamoxifen or ICI 182,780 (P<0.05). These results demonstrated that cathepsin E-A-like shared the same evolutionary origin with nothepsin. The expression of cathepsin E-A-like was regulated by oestrogen, and the regulative effect was predominantly mediated through ER-β in liver of chicken. [ABSTRACT FROM AUTHOR]

Background: The molecular mechanisms underlying meat quality and muscle growth are not clear. The meat quality and growth rates of local chickens and commercial broilers are very different. The Ribo-Zero RNA-Seq technology is an effective means of analyzing transcript groups to clarify molecular mechanisms. The aim of this study was to provide a reference for studies of the differences in the meat quality and growth of different breeds of chickens. Results: Ribo-Zero RNA-Seq technology was used to analyze the pectoral muscle transcriptomes of Gushi chickens and AA broilers. Compared with AA broilers, 1649 genes with annotated information were significantly differentially expressed (736 upregulated and 913 downregulated) in Gushi chickens with Q≤0.05 (Q is the P-value corrected by multiple assumptions test) at a fold change ≥2 or ≤0.5. In addition, 2540 novel significantly differentially expressed (SDE) genes (1405 upregulated and 1135 downregulated) were discovered. The results showed that the main signal transduction pathways that differed between Gushi chickens and AA broilers were related to amino acid metabolism. Amino acids are important for protein synthesis, and they regulate key metabolic pathways to improve the growth, development and reproduction of organisms. Conclusion: This study showed that differentially expressed genes in the pectoral tissues of Gushi chickens and AA broilers were related to fat metabolism, which affects meat. Additionally, a large number of novel genes were found that may be involved in fat metabolism and thus may affect the formation of meat, which requires further study. The results of this study provide a reference for further studies of the molecular mechanisms of meat formation. [ABSTRACT FROM AUTHOR]

Abstract: MicroRNAs (miRNAs) are noncoding RNAs that impact almost every aspect of biology and disease. Until now, the cell-specific effects of miRNAs in cardiovascular system have not been established. In the current study, the cellular functions of miR-221 and miR-222 (miR-221/222) in vascular smooth muscle cells (VSMCs) and vascular endothelial cells (ECs) were compared. In cultured cells, we identified that the effects of miR-221/222 on proliferation, migration, and apoptosis are opposite between VSMCs and ECs. In VSMCs, miR-221/222 had effects of pro-proliferation, pro-migration, and anti-apoptosis. In contrast, miR-221/222 had effects of anti-proliferation, anti-migration, and pro-apoptosis in ECs. The different expression profiles of their target genes, p27(Kip1), p57(kip2), and c-kit between the two cell types might be related to the opposite effects. Finally, the opposite cellular effects of miR-221/222 were verified in vivo in balloon-injured rat carotid artery as demonstrated by different consequences in neointimal growth and re-endothelialization. The results suggest that the biological functions of miR-221/222 in vascular walls are cell-specific. The opposite cellular effects of miR-221/222 on VSMCs and ECs may have important therapeutic applications in many vascular diseases such as atherosclerosis and restenosis after angioplasty. [Copyright &y& Elsevier]

Sox21 is thought to function as a counteracting partner of SoxB1 ( Sox1, 2, 3) genes and is involved in cell fate determination. In this study, we comparatively analyzed the expression patterns and conserved cis-regulatory elements of the duplicated sox21 genes in zebrafish. In embryogenesis, sox21b is predominantly expressed in the telencephalon, hypothalamus, mesencephalon and lens, and sox21a is solely expressed in the midbrain-hindbrain boundary, olfactory placode and lateral line, while both genes are expressed in the hindbrain, spinal cord and ear. In adult, sox21a is expressed in the brain, skin, ovary and intestine, while sox21b is expressed in the brain and testis. Interestingly, all 16 pan-vertebrate conserved non-coding elements (CNEs) are asymmetrically preserved in the sox21b locus, whereas two fish-specific elements are kept in the sox21a locus, and this is correlated with increased evolutionary rate of the sox21a protein sequence. Transient transgenic reporter analysis revealed that six sox21b CNEs and two sox21a CNEs drove green fluorescent protein (GFP) expression in tissues correlated with the partitioning of expression in two orthologues. These results indicate that sox21a and sox21b have reciprocally lost expression domains of the ancestral gene reflected by degeneration of certain CNEs in their genomic loci and provide clear evidence for evolution of the duplicated sox21 genes by subfunctionalization. In addition, our data suggest that some CNEs-based regulatory pathways have been predominantly preserved in the sox21b locus. [ABSTRACT FROM AUTHOR]

Abstract: In mammals, alternative splicing of the leptin receptor (LEPR) produces several C-terminal truncated isoforms that are believed to play a role in the transport, cellular internalisation and degradation of the hormone leptin. The chicken leptin receptor (chLEPR) is similar to its mammalian counterparts in terms of its intron/exon structure and conserved motifs. However, it is unknown whether the chLEPR also undergoes alternative splicing. To test this, structural analysis of intron 19 of the chLEPR, equivalent to the intron in which alternative splicing occurs in mammals, was combined with 3′-rapid amplification of cDNA ends (3′-RACE) to search for chLEPR splice variants. A 44-amino acid alternative exon 20 was identified that is spliced to generate a short isoform of the chLEPR (chLEPR-SF). Comparative sequence analysis of intron 19 identified two regions that are highly conserved between the chicken and mammals, indicating their possible importance as intronic elements in the regulation of alternative splicing of the LEPR in vertebrates. Tissue expression of the chLEPR-SF was lower and more restricted than that of the chLEPR long isoform. Collectively these data demonstrate that the chLEPR is alternatively spliced to produce at least one short isoform, as is the case in mammals. [Copyright &y& Elsevier]

The genetic basis of variation in complex traits remains poorly understood, and few genes underlying variation have been identified. Previous work identified a quantitative trait locus (QTL) responsible for much of the response to selection on growth in mice, effecting a change in body mass of approximately 20%. By fine-mapping, we have resolved the location of this QTL to a 660-kb region containing only two genes of known function, Gpc3 and Gpc4, and two other putative genes of unknown function. There are no non-synonymous polymorphisms in any of these genes, indicating that the QTL affects gene regulation. Mice carrying the high-growth QTL allele have approximately 15% lower Gpc3 mRNA expression in kidney and liver, whereas expression differences at Gpc4 are non-significant. Expression profiles of the two other genes within the region are inconsistent with a factor responsible for a general effect on growth. Polymorphisms in the 3′ untranslated region of Gpc3 are strong candidates for the causal sequence variation. Gpc3 loss-of-function mutations in humans and mice cause overgrowth and developmental abnormalities. However, no deleterious side-effects were detected in our mice, indicating that genes involved in Mendelian diseases also contribute to complex trait variation. Furthermore, these findings show that small changes in gene expression can have substantial phenotypic effects. The genetic basis of variation in complex traits remains poorly understood. Here, a genetic variant is discovered that leads to increased body size in mice. [ABSTRACT FROM AUTHOR]

Acute myeloid leukemia (AML) presents ongoing therapeutic challenges due to its intricate molecular pathogenesis. This study aimed to elucidate the role of RNA binding motif protein 39 (RBM39) in AML cell proliferation, apoptosis, and chemosensitivity, and its potential modulation of the PI3K/AKT pathway. In vitro and in vivo experiments were conducted using AML cell lines (K562 and U937) and bone marrow mononuclear cells (BM-MNCs) from AML patients and healthy donors. RBM39 mRNA and protein levels were measured using qRT-PCR and Western blotting. Cells were transfected with sh-RBM39 or sh-control, and then treated with daunorubicin (DNR) or homoharringtonine (HHT) at varied concentrations. Cell proliferation, chemosensitivity, and apoptosis were assessed through CCK-8 assay and Annexin V–APC/PI staining. RNA sequencing identified differentially expressed genes (DEGs) post RBM39 knockdown. An in vivo xenograft AML model using E7070, a selective RBM39 inhibitor, was employed to evaluate RBM39 modulation effects. Elevated RBM39 levels were found in AML patients and cell lines compared to controls. RBM39 knockdown promoted apoptosis, curtailed cell proliferation, and enhanced chemosensitivity to DNR and HHT in vitro. Drug-resistant or relapsed AML patients displayed higher RBM39 levels. RNA sequencing after RBM39 knockdown revealed downregulated PI3K/AKT signaling. The xenograft model validated in vitro results, as E7070 treatment suppressed AML xenograft growth via RBM39-mediated PI3K/AKT pathway suppression. RBM39 plays a pivotal role in AML progression through the PI3K/AKT signaling pathway. Targeting RBM39, potentially with E7070, could inhibit proliferation and induce apoptosis in AML cells, offering a promising avenue for future AML research and treatment. A schematic model depicting the role of the RBM39–PI3K/AKT axis in AML. [Display omitted] • Elevated RBM39 levels found in AML patients and cell lines compared to healthy controls • RBM39 knockdown enhances apoptosis, curtails proliferation, and boosts chemosensitivity in AML. • Higher RBM39 levels linked to drug-resistant or relapsed AML cases • RBM39 modulation downregulates the PI3K/AKT signaling pathway. • E7070, a selective RBM39 inhibitor, suppressed AML growth in vivo via the PI3K/AKT pathway [ABSTRACT FROM AUTHOR]

Cholestasis is a commonly occurring disorder induced by impaired bile flow, for which there is no effective treatment so far. Qingre Lidan decoction (QRLD) is a clinically used herbal compound for the long-term treatment of bile circulation disorders arising from inflammation and obstruction in the gallbladder and bile ducts. The objective of this study was to investigate the protective effect of QRLD on cholestatic liver injury and its possible mechanism. α-Naphthyl isothiocyanate (ANIT) was used to induce cholestatic liver injury in rats. Liver histopathology and serum biochemical markers were used to assess QRLD's protective impact. The possible biomarkers and mechanism of the therapeutic benefits of QRLD were investigated using a UHPLC-based Q-Exactive Orbitrap MS / MS untargeted serum metabolomics technique together with 16 S rRNA microbiota profiling. Afterwards, using RT-qPCR as well as Western Blot techniques, the expression of pertinent indicators was determined. The intervention effect of QRLD was stronger at medium and high dosages than at low doses, and it dramatically decreased the levels of serum biochemical markers in cholestatic rats reflecting alterations in liver function and relieving ANIT-induced abnormalities in the liver's histopathology. Serum metabolomics showed that QRLD could affect the metabolic profile of cholestatic rats, mainly related to glycerophospholipid metabolism, taurine and hypotaurine metabolism, alanine, aspartate and glutamate metabolism, and histidine metabolic pathway. Additionally, analysis of 16 S rRNA gene sequencing indicated that QRLD could moderate ANIT-induced microbiota disorders, particularly Romboutsia, Bifidobacterium, Fusicatenibacter, Prevotella_9, Prevotellaceae_NK3B31_group and Prevotella_1. Other experimental results showed that QRLD significantly upregulated the mRNA and protein expression of PPARα, CYP7A1 and NTCP in the liver, inhibited the expression of p-IκBα, p-p65 and TNFα while increasing the anti-inflammatory factor IL-10, and downregulated the expression of MDA (a peroxidation product) and D -lactic acid (an intestinal barrier indicator) while increasing the expression of SOD and GSH. QRLD can effectively regulate endogenous metabolites and microbiota disorders in cholestatic rats that are correlated with the attenuation of inflammation and oxidative stress. [Display omitted] • Using 16S rRNA gene sequencing and metabolomics techniques to assess the protective effect of QRLD against cholestasis. • ANIT-induced liver injury leads to disturbances in serum metabolites and fecal microbiota. • Bacterial genera and metabolites were significantly altered and restored to normal levels after QRLD treatment. • QRLD significantly activated PPARα and inhibited NF-κB signaling pathway. [ABSTRACT FROM AUTHOR]

Background:Reactive oxygen species arise in the mitochondria as byproducts of respiration and oxidase activity and have important roles in many physiological and pathophysiological conditions. The level of reactive oxygen species is regulated by a number of enzymes and physiological antioxidants, including HO-1, Sod2, catalase and COX-2, etc. And HO-1 against oxidative stress requires an increase in stress-responsive genes, such as Sod2 and catalase. Especially for the activity of HO-1, cobalt protoporphyrin is known to be a potent and effective inducer in many tissues. The transcription factor, FOXO1 is resistant to oxidative stress through downregulating reactive oxygen species production. Previous study showed that FOXO1 induces HO-1 expression by binding to HO-1 promoter. The question whether cobalt protoporphyrin induces HO-1 expression mediated by FOXO1 and subsequently lessens reactive oxygen species production remains to be elucidated. Results:Cobalt protoporphyrin enhances the expression of FOXO1 and facilitates FOXO1 binding to HO-1 promoter and increasing its transcriptional activity without influencing the FOXO1 protein stability. CoPP induces HO-1 and other oxidative stress-responsive genes expression, such as catalase, cytochrome c, Sod2, and COX-2, and decreases mitochondria-derived reactive oxygen species production, which are mediated partially by FOXO1. Conclusions:Cobalt protoporphyrin induces HO-1 and other oxidative stress-responsive genes expression mediated partially by FOXO1, and has an important role in reducing cellular reactive oxygen species level. Cobalt protoporphyrin may be a more promising therapeutic agent to upregulate some antioxidantive genes. [ABSTRACT FROM AUTHOR]

The "KNDy neurons" located in the hypothalamic arcuate nucleus (ARC) of mammals are known to co-express kisspeptin, neurokinin B (NKB), and dynorphin (DYN), and have been identified as key mediators of the feedback regulation of steroid hormones on gonadotropin-releasing hormone (GnRH). However, in birds, the genes encoding kisspeptin and its receptor GPR54 are genomic lost, leaving unclear mechanisms for feedback regulation of GnRH by steroid hormones. Here, the genes tachykinin 3 ( TAC3 ) and prodynorphin ( PDYN ) encoding chicken NKB and DYN neuropeptides were successfully cloned. Temporal expression profiling indicated that TAC3, PDYN and their receptor genes (TACR3, OPRK1) were mainly expressed in the hypothalamus, with significantly higher expression at 30W than at 15W. Furthermore, overexpression or interference of TAC3 and PDYN can regulate the GnRH mRNA expression. In addition, in vivo and in vitro assays showed that estrogen (E2) could promote the mRNA expression of TAC3, PDYN , and GnRH , as well as the secretion of GnRH/LH. Mechanistically, E2 could dimerize the nuclear estrogen receptor 1 (ESR1) to regulate the expression of TAC3 and PDYN , which promoted the mRNA and protein expression of GnRH gene as well as the secretion of GnRH. In conclusion, these results revealed that E2 could regulate the GnRH expression through TAC3 and PDYN systems, providing novel insights for reproductive regulation in chickens. [ABSTRACT FROM AUTHOR]

Long non-coding RNAs (lncRNAs) play an essential role in vertebrate myogenesis and muscle diseases. However, the dynamic expression patterns, biological functions, and mechanisms of lncRNAs in skeletal muscle development and regeneration remain largely unknown. In this study, a novel lncRNA (named lncMGR) was differentially expressed during breast muscle development in fast- and slow-growing chickens. Functionally, lncMGR promoted myoblast differentiation, inhibited myoblast proliferation in vitro , and promoted myofiber hypertrophy and injury repair in vivo. Mechanistically, lncMGR increased the mRNA and protein expression of skeletal muscle myosin heavy chain 1 A (MYH1A) via both transcriptional and post-transcriptional regulation. Nuclear lncMGR recruited cyclin-dependent kinase 9 (CDK9) to the core transcriptional activation region of the MYH1A gene to activate MYH1A transcription. Cytoplasmic lncMGR served as a competitive endogenous RNA (ceRNA) to competitively absorb miR-2131-5p away from MYH1A and subsequently protected the MYH1A from miR-2131-5p-mediated degradation. Besides miR-2131-5p, cytoplasmic lncMGR could also sponge miR-143-3p to reconcile the antagonist between the miR-2131-5p/ MYH1A -mediated inhibition effects and miR-143-3p-mediated promotion effects on myoblast proliferation, thereby inhibiting myoblast proliferation. Collectively, lncMGR could recruit CDK9 and sponge multiple miRNAs to regulate skeletal muscle development and regeneration, and could be a therapeutic target for muscle diseases. [ABSTRACT FROM AUTHOR]

The economic losses incurred due to reduced muscle pigmentation highlight the crucial role of melanin-based coloration in the meat of black-bone chickens. Melanogenesis in the breast muscle of black-bone chickens is currently poorly understood in terms of molecular mechanisms. This study employed whole-transcriptome sequencing to analyze black and white breast muscle samples from black-bone chickens, leading to the identification of 367 differentially expressed (DE) mRNAs, 48 DElncRNAs, 104 DEcircRNAs, and 112 DEmiRNAs involved in melanin deposition. Based on these findings, a competitive endogenous RNA (ceRNA) network was developed to better understand the complex mechanisms of melanin deposition. Furthermore, our analysis revealed key DEmRNAs (TYR, DCT, EDNRB, MLPH and OCA2) regulated by DEmiRNAs (g ga-miR-140-5p, gga-miR-1682, gga-miR-3529, gga-miR-499-3p, novel-m0012-3p, gga-miR-200b-5p, gga-miR-203a, gga-miR-6651-5p, gga-miR-7455-3p, gga-miR-31-5p, miR-140-x, miR-455-x, novel-m0065-3p, gga-miR-29b-1-5p, miR-455-y, novel-m0085-3p, and gga-miR-196-1-3p). These DEmiRNAs competitively interacted with DElncRNAs including MSTRG.2609.2, MSTRG.4185.1, LOC112530666, LOC112533366, LOC771030, LOC107054724, LOC121107411, LOC100859072, LOC101750037, LOC121108550, LOC121109224, LOC121110876, and LOC101749016 , as well as DEcircRNAs, such as novel_circ_000158, novel_circ_000623, novel_001518, and novel_circ_003596. The findings from this study provide insight into the mechanisms that regulate lncRNA, circRNA, miRNA, and mRNA expression in chicken melanin deposition. [ABSTRACT FROM AUTHOR]

In order to survive in extreme environments, plants have evolved multiple mechanisms, such as DNA methylation, to adapt to environmental stress. A large number of investigations indicate that abiotic stress triggers changes in DNA methylation, resulting in fluctuations of expression levels of stress-related genes, thereby enhancing stress tolerance in plants. This review summarized that DNA methylation in plants can enhance tolerance to abiotic stress, including nutrient, sulfur dioxide (SO 2), metal, temperature and osmotic stress. The dynamics of DNA methylation in plant under abiotic stress was discussed from the aspects of genome stability, gene expression, gene silencing and protein interaction. Moreover, several important pathways of methylation also are involved in response to stress. Firstly, methylation activated or inhibited gene expression and mediatted interaction of genes under abiotic stress. Secondly, the RNA-directed DNA methylation (RdDM) pathway can participate in gene expression. Third, up-regulation of DNA methylase and demethylase genes can also affect DNA methylation levels. Fourth, methylation mediated the response of plant antioxidant system. Fifth, DNA methylation had a potential regulatory role in plant cross-adaptation. Additionally, we summarized the methylation of abscisic acid and ethylene synthesis related-genes under stress. We also speculate that the interaction of methylation with other phytohormones can enhance plant stress resistance. Therefore, this review provides a better understanding of plant response to abiotic stress at the molecular level. • DNA methylation in plants can enhance tolerance to nutrient, sulfur dioxide, Aluminum, temperature and osmotic stress. [ABSTRACT FROM AUTHOR]

miR-19b-3p is reported to undertake various biological role, while its function and action mechanism in chicken hepatic lipid metabolism is unclear. Conservation analysis and tissue expression pattern of miR-19b-3p and its target gene were evaluated, respectively. Dual luciferase reporter system and Western blot technologies were adopted to validate miR-19b-3p target gene. Overexpression and knockdown assays were done to explore the biological functions of miR-19b-3p and target gene in Leghorn Male Hepatoma cell line (LMH). Regulatory approaches of estrogen on miR-19b-3p and target gene expressions are analyzed through site-directed mutation combined with estrogen receptors antagonist treatment assays. The results showed that chicken miR-19b-3p mature sequences are highly conserved among Capra hircus, Columba livia, Rattus norvegicus, Mus musculus, Cricetulus griseus, Danio rerio, Danio novaehollandiae, Orycodylus porosus, Crocodylus porosus, Gadus morhua , and widely expressed in lung, ovary, spleen, duodenum, kidney, heart, liver, leg muscle, and pectoral muscle tissues. miR-19b-3p could significantly increase intracellular triglyceride (TG) content and decrease intracellular cholesterol (TC) content via targeting methylsterol monooxygenase 1 ( MSMO1 ) and elongase of very long chain fatty acids 5 ( ELOVL5 ), which are highly conserved among species, in both mRNA and protein levels. Estrogen could inhibit miR-19b-3p expression, but directly promoted MSMO1 transcription via estrogen receptor α (ERα) and indirectly regulated ELOVL5 expression at the transcription level. Meanwhile, estrogen could also upregulate MSMO1 and ELOVL5 expression through inhibiting miR-19b-3p expression at the post-transcription level. Taken together, these results highlight the role and regulatory mechanism of miR-19b-3p in hepatic lipid metabolism in chicken, and might produce useful comparative information for human obesity studies and biomedical research. [ABSTRACT FROM AUTHOR]

The elongation of very long chain fatty acids protein 6 (ELOVL6) encodes a fatty acid elongase that is responsible for the final step in endogenous saturated fatty acid synthesis and involves in de novo lipogenesis. Though the regulatory mechanism of ELOVL6 expression has been studied extensively, little is known about the role of miRNA in regulating ELOVL6 gene expression in chicken until now. To investigate the regulatory mechanism of miRNA on the expression of ELOVL6 gene, bioinformatics analysis was employed to predict the potential miRNAs that binding with the 3′untranslated region (3′UTR) of ELOVL6. The putative miRNA was further screened by comparative analysis with previous miRNA-seq results. Gga-miR-22-3p, which could bind with the 3′UTR of ELOVL6 and showed negative expression correlation with ELOVL6 gene in chicken liver, was obtained. Tissue expression profiles showed that gga-miR-22-3p and ELOVL6 are extensively expressed in many tissues, and ELOVL6 with high expression level in kidney and liver tissues, and gga-miR-22-3p with high expression in lung and heart. Dual-luciferase reporter assays results indicated that the expression of luciferase reporter gene linked with part sequence of the 3′UTR of chicken ELOVL6 gene was down-regulated by the overexpression of gga-miR-22-3p in the DF1 cells, and the down-regulation behavior was abolished when the gga-miR-22-3p binding site in 3′UTR of ELOVL6 was mutated ( P > 0.05). Furthermore, the ELOVL6 expression in chicken hepatocytes was down-regulated when miR-22-3p was over-expressed. Therefore, we concluded that miR-22-3p might involve in controlling the hepatic lipid composition through affecting the expression of ELOVL6 gene, and could serve as a regulator of lipid metabolism in the liver of egg-laying hen. [ABSTRACT FROM AUTHOR]

Glycoprotein nonmetastatic melanoma protein B (GPNMB) is a vital secreted factor that promotes the occurrence of obesity in mammals. However, the effects of GPNMB on abdominal fat deposition is still unknown in chickens. In this study, we looked into the genetic and expression association of GPNMB gene with abdominal fat traits in chicken, and found that a genetic variation rs31126482 in GPNMB promoter was significantly associated with abdominal fat weight (AFW , P < 0.05) and abdominal fat percentage (AFP , P < 0.01). Express profile analysis of the GPNMB indicated that the gene was mainly expressed in abdominal fat tissue, and its expression level was strongly positively correlated with AFW (R2 = 0.6356, P = 4.10E−05) and AFP (R2 = 0.6450, P = 2.90E−05). We then investigated biological function of GPNMB on adipogenesis in chicken, and found that GPNMB could inhibit abdominal preadipocyte proliferation, but promote abdominal preadipocyte differentiation and lipid deposition. Furthermore, we explored regulatory mechanism of GPNMB gene in chicken, and detected one nonclassical estrogen regulatory element (AP1) and one peroxisome proliferator-activated receptor α (PPARα) responsive element in the 2 kb promoter region of GPNMB gene, and demonstrated that estrogen could up-regulate GPNMB mRNA expression in adipose tissue and primary abdominal preadipocytes, while PPARα could down-regulate GPNMB expression in primary preadipocytes. Taken together, this study brings new insights into understanding the function and transcriptional control of GPNMB gene, and provides genetic markers for breeding selection to improve abdominal fat traits in chicken. [ABSTRACT FROM AUTHOR]

• A 31-bp indel located in LDB2 gene was detected in chicken for the first time. • The indel cause significant differences in growth and carcass traits of F2. • The expression of LDB2 gene in breast muscle was higher than that in other tissues. • In the breast muscle, it reached the highest at the age of 6 weeks. • The 31-bp indel can be used as a genetic marker in chicken molecular breeding. Many studies have shown that the LDB2 gene plays a regulatory role in retinal development and the cell cycle, but its biological role remains unclear. In this study, a 31-bp indel in the LDB2 gene was found for the first time on the basis of 2797 individuals from 10 different breeds, which led to different genotypes among individuals (II, ID and DD). Among these genotypes, DD was the most dominant. Association analysis of an F 2 resource population crossed with the Gushi (GS) chicken and Anka chicken showed that the DD genotype conferred a significantly greater semi-evisceration weight (SEW, 1108.665 g ± 6.263), evisceration weight (EW, 927.455 g ± 5.424), carcass weight (CW, 1197.306 g ± 6.443), breast muscle weight (BMW, 71.05 g ± 0.574), and leg muscle weight (LMW, 100.303 g ± 0.677) than the ID genotype (SEW, 1059.079 g ± 16.86; EW, 879.459 g ± 14.446; CW, 1141.821 g ± 17.176; BMW, 67.164 g ± 1.523; and LMW, 96.163 g ± 1.823). In addition, LDB2 gene expression in different breeds was significantly higher in the breast muscles and leg muscles than in other tissues. The expression level in the breast muscle differed significantly among stages of GS chicken development, with the highest expression observed at 6 weeks. The expression levels in the pectoral muscles differed significantly among Ross 308 genotypes. In summary, we studied the relationships between a 31-bp indel in the LDB2 gene and economic traits in chickens. The indel was significantly correlated with multiple growth and carcass traits in the F 2 resource population and affected the expression of the LDB2 gene in muscle tissue. In short, our study revealed that the LDB2 gene 31-bp indel can be used as a potential genetic marker for molecular breeding. [ABSTRACT FROM AUTHOR]