MTOR1C-dependent suppression of autophagic activity in somatic cell nuclear transfer mouse embryos
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ABSTRACT:
PROVIDER: PRJNA1311358 | ENA |
REPOSITORIES: ENA
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Project description:Activation of autophagy after fertilization is essential for mammalian embryonic development, as it supplies embryos with nutrients and energy. Somatic cell nuclear transfer (SCNT) embryos frequently exhibit developmental arrest, largely because of incomplete genomic reprogramming; however, the role of non-genomic factors remains unclear. Here, we investigated autophagy dynamics in mouse SCNT embryos using immunostaining and live-cell fluorescence imaging. In fertilized embryos, autophagy increased markedly from the late 2-cell stage and peaked at the morula stage. SCNT embryos followed a similar timeline but consistently showed reduced autophagic activity. Notably, the autophagic activity levels varied among SCNT embryos and positively correlated with their developmental potential. Attempts to enhance genomic reprogramming, including the removal of somatic histone methylation, did not restore autophagy. Instead, transcriptome analysis revealed ectopic activation of mTORC1 signaling as a likely cause of impaired autophagy. Consistently, treatment with an mTORC1 inhibitor successfully rescued the autophagic activity in SCNT embryos. These findings identify a persistent autophagy defect during preimplantation development in SCNT embryos and suggest that modulation of non-genomic pathways, such as mTORC1 signaling, could improve SCNT efficiency.
2025-09-01 | GSE306587 | GEO
Project description:Suppression of endogenous retroviral enhancers in somatic cell nuclear transfer embryos
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Project description:We report a novel long noncoding RNA, LINC00622, which modulates the mTORC1-regulated autophagy in cutaneous melanoma. Functionally, LINC00622 acts as an oncogene to promote the proliferation, colony formation, migration and invasion in melanoma while suppressing cell death. Mechanically, LINC00622 associates and recruits BTF3 to transcriptionally enhance RRAGD expression for activating mTORC1 and inhibiting autophagic cell death, which contributes to the development of cutaneous melanoma. Our findings not only demonstrated the oncogenic role of LINC00622 via RRAGD/mTORC1 axis to repress autophagic cell death in cutaneous melanoma, but also offer novel treatment targets for melanoma therapy.
2025-02-19 | GSE275315 | GEO
Project description:Endogenous retroviruses (ERVs) in the mammalian genome play diverse roles in embryonic development. These development-related ERVs are generally repressed in somatic cells and are therefore likely repressed in embryos derived from somatic cell nuclear transfer (SCNT). In this study, we sought to identify ERVs that are repressed in SCNT-derived morula embryos, which may consequently cause previously unexplained embryonic deaths shortly after implantation. Our transcriptome analysis revealed that, amongst ERV families, ERVK was specifically and strongly downregulated in SCNT embryos while other transposable elements including LINE and ERVL were unchanged. Among subfamilies of ERVK, RLTR45-int was most repressed in SCNT embryos despite its highest expression in control fertilized embryos. Interestingly, the nearby genes (within 5–50 kb, n = 19) of the repressed RLTR45-int loci were also repressed in SCNT embryos with a significant correlation between them. Furthermore, lysine H3K27 acetylation was enriched around the RLTR45-int loci. These findings indicate that RLTR45-int elements function as enhancers of nearby genes. Indeed, deletion of two sequential RLTR45-int loci on chromosome 4 or 18 resulted in downregulations of nearby genes at the morula stage. We also found that RLTR45-int loci, especially SCNT-low, enhancer-like loci, were strongly enriched with H3K9me3, a repressive histone mark. Importantly, these H3K9me3-enriched regions were resistant to histone demethylase Kdm4d in SCNT embryos. Thus, we identified ERVK subfamily RLTR45-int, putative enhancer elements, as a strong reprogramming barrier for SCNT. A large-scale downregulations of their regulating genes in SCNT morulae might cumulatively affect postimplantation development of SCNT embryos.
2022-10-08 | GSE214878 | GEO
Project description:Endogenous retroviruses (ERVs) in the mammalian genome play diverse roles in embryonic development. These developmentally related ERVs are generally repressed in somatic cells and therefore are likely repressed in embryos derived from somatic cell nuclear transfer (SCNT). In this study, we sought to identify ERVs that are repressed in SCNT-derived morulae, which might cause previously unexplained embryonic deaths shortly after implantation. Our transcriptome analysis revealed that, amongst ERV families, ERVK was specifically, and strongly downregulated in SCNT-derived embryos while other transposable elements including LINE and ERVL were unchanged. Among the subfamilies of ERVK, RLTR45-int was most repressed in SCNT-derived embryos despite its highest expression in control fertilized embryos. Interestingly, the nearby genes (within 5-50 kb, n = 18; 50-200 kb, n = 63) of the repressed RLTR45-int loci were also repressed in SCNT-derived embryos, with a significant correlation between them. Furthermore, lysine H3K27 acetylation was enriched around the RLTR45-int loci. These findings indicate that RLTR45-int elements function as enhancers of nearby genes. Indeed, deletion of two sequential RLTR45-int loci on chromosome 4 or 18 resulted in downregulations of nearby genes at the morula stage. We also found that RLTR45-int loci, especially SCNT-low, enhancer-like loci, were strongly enriched with H3K9me3, a repressive histone mark. Importantly, these H3K9me3-enriched regions were not activated by overexpression of H3K9me3 demethylase Kdm4d in SCNT-derived embryos, suggesting the presence of another epigenetic barrier repressing their expressions and enhancer activities in SCNT embryos. Thus, we identified ERVK subfamily RLTR45-int, putative enhancer elements, as a strong reprogramming barrier for SCNT (253 words).
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Project description:We recently reported that the in vitro and in vivo survival of Rickettsia australis are Atg5-dependent, in association with an inhibited level of anti-rickettsial cytokine, IL-1β. In the present study, we sought to investigate how R. australis interacts with host innate immunity via an Atg5-dependent autophagic response. We found that the serum levels of IFN-γ and G-CSF in R. australis-infected Atg5flox/floxLyz-Cre mice were significantly less compared to Atg5flox/flox mice, accompanied by significantly lower rickettsial loads in tissues with inflammatory cellular infiltrations including neutrophils. R. australis infection differentially regulated a significant number of genes in bone marrow-derived macrophages (BMMs) in an Atg5-depdent fashion as determined by RNA sequencing and Ingenuity Pathway Analysis, including genes in the molecular networks of IL-1 family cytokines and PI3K-Akt-mTOR. The secretion levels of inflammatory cytokines, such as IL-1α, IL-18, TNF-α, and IL-6, by R. australis-infected Atg5flox/floxLyz-Cre BMMs were significantly greater compared to infected Atg5flox/flox BMMs. Interestingly, R. australis significantly increased the levels of phosphorylated mTOR and P70S6K at a time when the autophagic response is induced. Rapamycin treatment nearly abolished the phosphorylated mTOR and P70S6K but did not promote significant autophagic flux during R. australis infection. These results highlight that R. australis modulates an Atg5-dependent autophagic response, which is not sensitive to regulation by mTORC1 signaling in macrophages. Overall, we demonstrate that R. australis counteracts host innate immunity including IL-1β-dependent inflammatory response to support the bacterial survival via an mTORC1-resistant autophagic response in macrophages.
2021-04-06 | GSE171160 | GEO
Project description:Trp53 Mutation-Mediated mTORC1 Inhibition and Inflammation Suppression Confer Resistance to Checkpoint Immunotherapy
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Project description:Phosphatidylinositol-5-phosphate 4-kinases (PIP4ks) are a family of lipid kinases that specifically use PI-5-P as a substrate to synthesize PI-4,5-P2. Suppression of PIP4k function in Drosophila results in smaller cells and reduced target of rapamycin complex 1 (TORC1) signaling. We showed that the γ isoform of PIP4k stimulated signaling through mammalian TORC1 (mTORC1). Knockdown of PIP4kγ reduced cell mass in cells in which mTORC1 is constitutively activated by Tsc2 deficiency. In Tsc2 null cells mTORC1 activation was partially independent of amino acids or glucose and glutamine. PIP4kγ knockdown inhibited the nutrient-independent activation of mTORC1 in Tsc2 knockdown cells and reduced basal mTORC1 signaling in wild-type cells. PIP4kγ was phosphorylated by mTORC1 and associated with the complex. Phosphorylated PIP4kγ was enriched in light microsomal vesicles, whereas the unphosphorylated form was enriched in heavy microsomal vesicles associated with the Golgi. Furthermore, basal mTORC1 signaling was enhanced by overexpression of an unphosphorylated wild-type PIP4kγ and phosphorylation-defective mutant and decreased by overexpression of a phosphorylation mimetic mutant. Together these results demonstrate that PIP4kγ and mTORC1 interact in a self-regulated feedback loop to maintain low and tightly regulated mTORC1 activation during starvation.
2016-05-04 | PXD001340 | Pride