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MicroRNAs (miRNAs) are post-transcriptional regulators that target specific mRNAs for repression and thus play key roles in many biological processes, including insect wing morphogenesis. miR-2 is an invertebrate-specific miRNA family that has been predicted in the fruit fly, Drosophila melanogaster, to be involved in regulating the Notch signaling pathway. We show here that miR-2 plays a critical role in wing morphogenesis in the silkworm, Bombyx mori, a lepidopteran model insect. Transgenic over-expression of a miR-2 cluster using a Gal4/UAS system results in deformed adult wings, supporting the conclusion that miR-2 regulates functions essential for normal wing morphogenesis. Two genes, abnormal wing disc (awd) and fringe (fng), which are positive regulators in Notch signaling, are identified as miR-2 targets and validated by a dual-luciferase reporter assay. The relative abundance of both awd and fng expression products was reduced significantly in transgenic animals, implicating them in the abnormal wing phenotype. Furthermore, somatic mutagenesis analysis of awd and fng using the CRISPR/Cas9 system and knock-out mutants also resulted in deformed wings similar to those observed in the miR-2 overexpression transgenic animals. The critical role of miR-2 in Bombyx wing morphogenesis may provide a potential target in future lepidopteran pest control.

Simple Summary: The problem of low fertility in Tibetan pigs has a long history. In order to solve the problem of low fertility in Tibetan pigs, in this study, we successfully constructed cDNA libraries of the transcriptome from ovarian tissues of Tibetan and Yorkshire pigs using RNA-Seq technology, analyzed and screened the genes significantly enriched in the pathways closely related to the regulation of reproduction, and finally identified AR, CYP11A1, CYP17A1, INHBA, ARRB2, EGFR, ETS1, HSD17B1, IGF1R, MIF, SCARB1, and SMAD4 as important candidate genes. The 12 differentially expressed genes screened in relation to reproduction were validated using RT-qPCR. The results showed that six genes were highly significantly (p < 0.01) higher in expression in Tibetan pigs than in Yorkshire pigs, i.e., AR, CYP17A1, EGFR, ETS1, IGF1R, and SMAD4; on the other hand, six genes were highly significantly (p < 0.01) lower in expression in Tibetan pigs than in Yorkshire pigs, i.e., CYP11A1, INHBA, ARRB2, HSD17B, MIF, SCARB1. This study provides a theoretical basis for improving the regulatory genes in Tibetan pigs. The purpose of this study was to explore and verify genes that regulate the reproductive traits of Tibetan pigs at the mRNA level. The ovarian tissues of Tibetan pigs (TPs) and Yorkshire pigs (YPs) were selected as research objects, and cDNA libraries of the ovarian tissue transcripts of Tibetan pigs and Yorkshire pigs were successfully constructed by the RNA-Seq technique. A total of 651 differentially expressed genes (DEGs) were screened, including 414 up-regulated genes and 237 down-regulated genes. Through GO and KEGG enrichment analysis, it was found that these differentially expressed genes were significantly enriched in cell process, reproductive process, reproduction, cell proliferation, binding, and catalytic activity, as well as oxidative phosphorylation, endocrine resistance, thyroid hormone, Notch, and other signal transduction pathways. Genes significantly enriched in pathways closely related to reproductive regulation were analyzed and selected, and the AR, CYP11A1, CYP17A1, INHBA, ARRB2, EGFR, ETS1, HSD17B1, IGF1R, MIF, SCARB1, and SMAD4 genes were identified as important candidate genes. Twelve differentially expressed genes related to reproduction were verified by RT-qPCR. The results showed that the expression of the AR, CYP17A1, EGFR, ETS1, IGF1R, and SMAD4 genes was significantly higher in Tibetan pigs than in Yorkshire pigs, while the expression of the CYP11A1, INHBA, ARRB2, HSD17B, MIF, and SCARB1 genes in Tibetan pigs was significantly lower than in Yorkshire pigs. The purpose of this study is to provide a theoretical basis for exploring the molecular mechanism of reproductive trait effect genes and the application of molecular breeding in Tibetan pigs. [ABSTRACT FROM AUTHOR]

Simple Summary: Muscle growth is essential for the economic sustainability of the swine industry. Gaining a deeper understanding of pig muscle development is crucial for improving both the quantity and quality of pork production. This study employed RNA-seq technology to conduct a transcriptomic analysis of the longissimus dorsi muscle (LD) in three different pig breeds with varying body sizes and growth rates: Tibetan pigs (TP), Wujin pigs (WJ), and large white pigs (LW). Significant differences in gene expression were observed among the different breeds. We identified that myosin light chain 4 (MYL4) plays a role in influencing muscle growth. Experimental validation confirmed that MYL4 promotes skeletal muscle cell proliferation and inhibits degradation. These findings reveal the molecular mechanisms that control muscle growth and provide valuable insights for improving pig breeding practices, ultimately enhancing the efficiency and quality of meat production. Muscle growth stands as a pivotal economic trait within pig production, governed by a complex interplay of multiple genes, each playing a role in its quantitative manifestation. Understanding the intricate regulatory mechanisms of porcine muscle development is crucial for enhancing both pork yield and quality. This study used the GSE99749 dataset downloaded from the GEO database, conducting a detailed analysis of the RNA-seq results from the longissimus dorsi muscle (LD) of Tibetan pigs (TP), Wujin pigs (WJ) and large white pigs (LW) at 60 days of gestation, representing diverse body sizes and growth rates. Comparative analyses between TPvsWJ and TPvsLW, along with differential gene expression (DEG) analysis, functional enrichment analysis, and protein–protein interaction (PPI) network analysis, revealed 1048 and 1157 significantly differentially expressed genes (p < 0.001) in TPvsWJ and TPvsLW, respectively. With stricter screening criteria, 37 DEGs were found to overlap between the 2 groups. PPI analysis identified MYL5, MYL4, and ACTC1 as the three core genes. This article focuses on exploring the MYL4 gene. Molecular-level experimental validation, through overexpression and interference of the MYL4 gene combined with EDU staining experiments, demonstrated that overexpression of MYL4 significantly promoted the proliferation of porcine skeletal muscle satellite cells (PSMSC), while interference with MYL4 inhibited their proliferation. Furthermore, by examining the effects of overexpressing and interfering with the MYL4 gene on the muscle hypertrophy marker Fst gene and the muscle degradation marker FOXO3 gene, the pivotal role of the MYL4 gene in promoting muscle growth and preventing muscle degradation was further confirmed. These findings offer a new perspective on the molecular mechanisms behind porcine muscle growth and development, furnishing valuable data and insights for muscle biology research. [ABSTRACT FROM AUTHOR]