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Background: It has been reported that the most common post-transcriptional modification of eukaryotic RNA is N6-methyladenosine (m6A). Previous studies show m6A is a key regulator for viral infection and immune response. However, whether there is a pathogen stimulus-dependent m6A regulation in invertebrate shrimp has not been studied. Results: In this study, we performed a transcriptome-wide profiling of mRNA m6A methylation in shrimp (Marsupenaeus japonicus) after white spot syndrome virus (WSSV) infection by methylated RNA immunoprecipitation sequencing (MeRIP-seq). A total of 15,436 m6A peaks were identified in the shrimp, distributed in 8,108 genes, mainly enriched in the CDS, 3′ UTR region and near the stop codon. After WSSV infection, we identified 2,260 m6A peaks with significantly changes, of which 1,973 peaks were significantly up-regulated and 287 peaks were significantly down-regulated. 1,795 genes were identified as differentially methylated genes. GO and KEGG analysis showed that hyper-methylated genes or hypo-methylated genes were highly associated with innate immune process and related to metabolic pathways including HIF-1 signaling pathway, lysine degradation and Wnt signaling pathway. Combined analysis showed a positive correlation between m6A methylation levels and mRNA expression levels. In addition, computational predictions of protein-protein interaction indicated that genes with altered levels of m6A methylation and mRNA expression clustered in metabolism, DNA replication, and protein ubiquitination. ZC3H12A and HIF-1 were two hub genes in protein-protein interaction (PPI) network that involved in immune and metabolism processes, respectively. Conclusion: Our study explored the m6A methylation pattern of mRNA in shrimp after WSSV infection, exhibited the first m6A map of shrimp at the stage of WSSV induced metabolic reprogramming. These findings may reveal the possible mechanisms of m6A-mediated innate immune response in invertebrates. [ABSTRACT FROM AUTHOR]

Abstract Background It has been reported that the most common post-transcriptional modification of eukaryotic RNA is N6-methyladenosine (m6A). Previous studies show m6A is a key regulator for viral infection and immune response. However, whether there is a pathogen stimulus-dependent m6A regulation in invertebrate shrimp has not been studied. Results In this study, we performed a transcriptome-wide profiling of mRNA m6A methylation in shrimp (Marsupenaeus japonicus) after white spot syndrome virus (WSSV) infection by methylated RNA immunoprecipitation sequencing (MeRIP-seq). A total of 15,436 m6A peaks were identified in the shrimp, distributed in 8,108 genes, mainly enriched in the CDS, 3′ UTR region and near the stop codon. After WSSV infection, we identified 2,260 m6A peaks with significantly changes, of which 1,973 peaks were significantly up-regulated and 287 peaks were significantly down-regulated. 1,795 genes were identified as differentially methylated genes. GO and KEGG analysis showed that hyper-methylated genes or hypo-methylated genes were highly associated with innate immune process and related to metabolic pathways including HIF-1 signaling pathway, lysine degradation and Wnt signaling pathway. Combined analysis showed a positive correlation between m6A methylation levels and mRNA expression levels. In addition, computational predictions of protein-protein interaction indicated that genes with altered levels of m6A methylation and mRNA expression clustered in metabolism, DNA replication, and protein ubiquitination. ZC3H12A and HIF-1 were two hub genes in protein-protein interaction (PPI) network that involved in immune and metabolism processes, respectively. Conclusion Our study explored the m6A methylation pattern of mRNA in shrimp after WSSV infection, exhibited the first m6A map of shrimp at the stage of WSSV induced metabolic reprogramming. These findings may reveal the possible mechanisms of m6A-mediated innate immune response in invertebrates.

Salt is one of the most important environmental factors in crop growth and development. N6-methyladenosine (m6A) is an epigenetic modification that regulates plant–environment interaction at transcriptional and translational levels. Sugar beet is a salt-tolerant sugar-yielding crop, but how m6A modification affects its response to salt stress remains unknown. In this study, m6A-seq was used to explore the role of m6A modification in response to salt stress in sugar beet (Beta vulgaris). Transcriptome-wide m6A methylation profiles and physiological responses to high salinity were investigated in beet roots. After treatment with 300 mM NaCl, the activities of peroxidase and catalase, the root activity, and the contents of Na+, K+, and Ca2+ in the roots were significantly affected by salt stress. Compared with the control plants, 6904 differentially expressed genes (DEGs) and 566 differentially methylated peaks (DMPs) were identified. Association analysis revealed that 243 DEGs contained DMP, and 80% of these DEGs had expression patterns that were negatively correlated with the extent of m6A modification. Further analysis verified that m6A methylation may regulate the expression of some genes by controlling their mRNA stability. Functional analysis revealed that m6A modifications primarily affect the expression of genes involved in energy metabolism, transport, signal transduction, transcription factors, and cell wall organization. This study provides evidence that a post-transcriptional regulatory mechanism mediates gene expression during salt stress by affecting the stability of mRNA in the root. [ABSTRACT FROM AUTHOR]

Abstract Background The number of myofiber is determined during the embryonic stage and does not increase during the postnatal period for birds, including goose. Thus, muscle production of adult goose is pre-determined during embryogenesis. Previous studies show N 6 -methyladenosine (m6A) is an important regulator for skeletal muscle development of birds and miRNAs play as a co-regulator for the skeletal muscle development in birds. Herein, we sequenced m6A and miRNA transcriptomes to investigate the profiles of m6A and their potential mechanism of regulating breast muscle development in Dingan Goose. Results We selected embryonic 21th day (E21) and embryonic 30th day (E30) to investigate the roles of transcriptome-wide m6A modification combining with mRNAs and miRNAs in goose breast muscle development. In this study, m6A peaks were mainly enriched in coding sequence (CDS) and start codon and397 genes were identified as differentially methylated genes (DMGs). GO and KEGG analysis showed that DMGs were highly related to cellular and metabolic process and that most DMGs were enriched in muscle-related pathways including Wnt signaling pathway, mTOR signaling and FoxO signaling pathway. Interestingly, a negative correlation between m6A methylation level and mRNA abundance was found through the analysis of m6A-RNA and RNA-seq data. Besides, we found 26 muscle-related genes in 397 DMGs. We also detected 228 differentially expressed miRNAs (DEMs), and further found 329 genes shared by the target genes of DEMs and DMGs (m6A-miRNA-genes), suggesting a tightly relationship between DEMs and DMGs. Among the m6A-miRNA-genes, we found 10 genes are related to breast muscle development. We further picked out an m6A-miRNA-gene, PDK3, from the 10 genes to visualize it and the result showed differentially methylated peaks on the mRNA transcript consistent with our m6A-seq results. Conclusion GO and KEGG of DMGs between E21 and E30 showed most DMGs were muscle-related. In total, 228 DEMs were found, and the majority of DMGs were overlapped with the targets of DEGs. The differentially methylated peaks along with an m6A-miRNA-gene, PDK3, showed the similar results with m6A-seq results. Taken together, the results presented here provide a reference for further investigation of embryonic skeletal muscle development mechanism in goose.

Hepatic inflammation is always accompanied with abnormal lipid metabolism. Whether N6-methyladenosine (m6A) mRNA methylation affects irregular inflammatory lipid level is unclear. Here, the m6A modification patterns in chicken liver at the acute stage of LPS-stimulated inflammation and at the normal state were explored via m6A and RNA sequencing and bioinformatics analysis. A total of 7,815 m6A peaks distributed in 5,066 genes were identified in the normal chicken liver and were mostly located in the CDS, 3′UTR region, and around the stop codon. At 2 h after the LPS intraperitoneal injection, the m6A modification pattern changed and showed 1,200 different m6A peaks. The hyper- and hypo-m6A peaks were differentially located, with the former mostly located in the CDS region and the latter in the 3′UTR and in the region near the stop codon. The hyper- or hypo-methylated genes were enriched in different GO ontology and pathways. Co-analysis revealed a significantly positive relationship between the fold change of m6A methylation level and the relative fold change of mRNA expression. Moreover, computational prediction of protein–protein interaction (PPI) showed that genes with altered m6A methylation and mRNA expression levels were clustered in processes involved in lipid metabolism, immune response, DNA replication, and protein ubiquitination. CD18 and SREBP-1 were the two hub genes clustered in the immune process and lipid metabolism, respectively. Hub gene AGPAT2 was suggested to link the immune response and lipid metabolism clusters in the PPI network. This study presented the first m6A map of broiler chicken liver at the acute stage of LPS induced inflammation. The findings may shed lights on the possible mechanisms of m6A-mediated lipid metabolism disorder in inflammation. [ABSTRACT FROM AUTHOR]

Hepatic inflammation is always accompanied with abnormal lipid metabolism. Whether N6-methyladenosine (m6A) mRNA methylation affects irregular inflammatory lipid level is unclear. Here, the m6A modification patterns in chicken liver at the acute stage of LPS-stimulated inflammation and at the normal state were explored via m6A and RNA sequencing and bioinformatics analysis. A total of 7,815 m6A peaks distributed in 5,066 genes were identified in the normal chicken liver and were mostly located in the CDS, 3′UTR region, and around the stop codon. At 2 h after the LPS intraperitoneal injection, the m6A modification pattern changed and showed 1,200 different m6A peaks. The hyper- and hypo-m6A peaks were differentially located, with the former mostly located in the CDS region and the latter in the 3′UTR and in the region near the stop codon. The hyper- or hypo-methylated genes were enriched in different GO ontology and pathways. Co-analysis revealed a significantly positive relationship between the fold change of m6A methylation level and the relative fold change of mRNA expression. Moreover, computational prediction of protein–protein interaction (PPI) showed that genes with altered m6A methylation and mRNA expression levels were clustered in processes involved in lipid metabolism, immune response, DNA replication, and protein ubiquitination. CD18 and SREBP-1 were the two hub genes clustered in the immune process and lipid metabolism, respectively. Hub gene AGPAT2 was suggested to link the immune response and lipid metabolism clusters in the PPI network. This study presented the first m6A map of broiler chicken liver at the acute stage of LPS induced inflammation. The findings may shed lights on the possible mechanisms of m6A-mediated lipid metabolism disorder in inflammation.

Background: The number of myofiber is determined during the embryonic stage and does not increase during the postnatal period for birds, including goose. Thus, muscle production of adult goose is pre-determined during embryogenesis. Previous studies show N6-methyladenosine (m6A) is an important regulator for skeletal muscle development of birds and miRNAs play as a co-regulator for the skeletal muscle development in birds. Herein, we sequenced m6A and miRNA transcriptomes to investigate the profiles of m6A and their potential mechanism of regulating breast muscle development in Dingan Goose. Results: We selected embryonic 21th day (E21) and embryonic 30th day (E30) to investigate the roles of transcriptome-wide m6A modification combining with mRNAs and miRNAs in goose breast muscle development. In this study, m6A peaks were mainly enriched in coding sequence (CDS) and start codon and397 genes were identified as differentially methylated genes (DMGs). GO and KEGG analysis showed that DMGs were highly related to cellular and metabolic process and that most DMGs were enriched in muscle-related pathways including Wnt signaling pathway, mTOR signaling and FoxO signaling pathway. Interestingly, a negative correlation between m6A methylation level and mRNA abundance was found through the analysis of m6A-RNA and RNA-seq data. Besides, we found 26 muscle-related genes in 397 DMGs. We also detected 228 differentially expressed miRNAs (DEMs), and further found 329 genes shared by the target genes of DEMs and DMGs (m6A-miRNA-genes), suggesting a tightly relationship between DEMs and DMGs. Among the m6A-miRNA-genes, we found 10 genes are related to breast muscle development. We further picked out an m6A-miRNA-gene, PDK3, from the 10 genes to visualize it and the result showed differentially methylated peaks on the mRNA transcript consistent with our m6A-seq results. Conclusion: GO and KEGG of DMGs between E21 and E30 showed most DMGs were muscle-related. In total, 228 DEMs were found, and the majority of DMGs were overlapped with the targets of DEGs. The differentially methylated peaks along with an m6A-miRNA-gene, PDK3, showed the similar results with m6A-seq results. Taken together, the results presented here provide a reference for further investigation of embryonic skeletal muscle development mechanism in goose. [ABSTRACT FROM AUTHOR]

Myogenic progenitor/stem cells retain their skeletal muscle differentiation potential by maintaining myogenic transcription factors such as MyoD. However, the mechanism of how MyoD expression is maintained in proliferative progenitor cells has not been elucidated. Here, we found that MyoD expression was reduced at the mRNA level by cell cycle arrest in S and G2 phases, which in turn led to the absence of skeletal muscle differentiation. The reduction of MyoD mRNA was correlated with the reduced expression of factors regulating RNA metabolism, including methyltransferase like 3 (Mettl3), which induces N6-methyladenosine (m6A) modifications of RNA. Knockdown of Mettl3 revealed that MyoD RNA was specifically downregulated and that this was caused by a decrease in processed, but not unprocessed, mRNA. Potential m6A modification sites were profiled by m6A sequencing and identified within the 5′ untranslated region (UTR) of MyoD mRNA. Deletion of the 5′ UTR revealed that it has a role in MyoD mRNA processing. These data showed that Mettl3 is required for MyoD mRNA expression in proliferative myoblasts.

Hepatic inflammation is always accompanied with abnormal lipid metabolism. Whether N 6 -methyladenosine (m 6 A) mRNA methylation affects irregular inflammatory lipid level is unclear. Here, the m 6 A modification patterns in chicken liver at the acute stage of LPS-stimulated inflammation and at the normal state were explored via m 6 A and RNA sequencing and bioinformatics analysis. A total of 7,815 m 6 A peaks distributed in 5,066 genes were identified in the normal chicken liver and were mostly located in the CDS, 3'UTR region, and around the stop codon. At 2 h after the LPS intraperitoneal injection, the m 6 A modification pattern changed and showed 1,200 different m 6 A peaks. The hyper- and hypo-m 6 A peaks were differentially located, with the former mostly located in the CDS region and the latter in the 3'UTR and in the region near the stop codon. The hyper- or hypo-methylated genes were enriched in different GO ontology and pathways. Co-analysis revealed a significantly positive relationship between the fold change of m 6 A methylation level and the relative fold change of mRNA expression. Moreover, computational prediction of protein-protein interaction (PPI) showed that genes with altered m 6 A methylation and mRNA expression levels were clustered in processes involved in lipid metabolism, immune response, DNA replication, and protein ubiquitination. CD18 and SREBP-1 were the two hub genes clustered in the immune process and lipid metabolism, respectively. Hub gene AGPAT2 was suggested to link the immune response and lipid metabolism clusters in the PPI network. This study presented the first m 6 A map of broiler chicken liver at the acute stage of LPS induced inflammation. The findings may shed lights on the possible mechanisms of m 6 A-mediated lipid metabolism disorder in inflammation., 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 © 2022 Guo, Zhang, Ma, Yu, Wang, Gao, Wang, Xu, Wei and Jing.)

Aim: To systematically profile the global m 6 A modification pattern in clear cell renal cell carcinoma (ccRCC). Methods: m 6 A modification patterns in ccRCC and normal tissues were described via m 6 A sequencing and RNA sequencing, followed by bioinformatics analysis. m 6 A-related RNAs were immunoprecipitated and validated by quantitative real-time PCR (qPCR). Results: In total, 6919 new m 6 A peaks appeared with the disappearance of 5020 peaks in ccRCC samples. The unique m 6 A-related genes in ccRCC were associated with cancer-related pathways. We identified differentially expressed mRNA transcripts with hyper-methylated or hypo-methylated m 6 A peaks in ccRCC. Conclusion: This study presented the first m 6 A transcriptome-wide map of human ccRCC, which may shed lights on possible mechanisms of m 6 A-mediated gene expression regulation.

Myogenic progenitor/stem cells retain their skeletal muscle differentiation potential by maintaining myogenic transcription factors such as MyoD. However, the mechanism of how MyoD expression is maintained in proliferative progenitor cells has not been elucidated. Here, we found that MyoD expression was reduced at the mRNA level by cell cycle arrest in S and G2 phases, which in turn led to the absence of skeletal muscle differentiation. The reduction of MyoD mRNA was correlated with the reduced expression of factors regulating RNA metabolism, including methyltransferase like 3 (Mettl3), which induces N 6 -methyladenosine (m 6 A) modifications of RNA. Knockdown of Mettl3 revealed that MyoD RNA was specifically downregulated and that this was caused by a decrease in processed, but not unprocessed, mRNA. Potential m 6 A modification sites were profiled by m 6 A sequencing and identified within the 5' untranslated region (UTR) of MyoD mRNA. Deletion of the 5' UTR revealed that it has a role in MyoD mRNA processing. These data showed that Mettl3 is required for MyoD mRNA expression in proliferative myoblasts., (© 2017 The Authors.)