Project description:Inflammation plays an essential role in eliminating microbial pathogens and repairing tissues, while sustained inflammation accelerates kidney damage and disease progression. Therefore, understanding the mechanisms of the inflammatory response is vital for developing therapies for inflammatory kidney diseases like acute kidney injury (AKI), which currently lacks effective treatment. Here, we identified N-acetyltransferase 10 (NAT10) as a significant mediator of inflammation. NAT10, the only known ‘writer’ protein for N4-acetylcytidine (ac4C) acetylation, is elevated in renal tubules across various AKI models, human biopsies and cultured tubular epithelial cells (TECs). Conditional knockout (cKO) of NAT10 in mouse kidneys attenuates renal dysfunction, inflammation, and infiltration of macrophages and neutrophils, whereas its conditional knock-in (cKI) exacerbates these effects. Mechanistically, our findings from ac4C-RIP-seq and RNA-seq analyses reveal that NAT10-mediated ac4C acetylation enhances the mRNA stability of a range of key chemokines, including C-C motif chemokine ligand 2 (CCL2) and C-X-C motif chemokine ligand 1(CXCL1), promoting macrophage and neutrophil recruitment and accelerating renal inflammation. Additionally, CCL2 and CXCL1 neutralizing antibodies or their receptor inhibitors, abrogated renal inflammation in NAT10-overexpression TECs or NAT10-cKI mice. Importantly, inhibiting NAT10, either through Adeno-associated virus 9 (AAV9)-mediated silencing or pharmacologically with our newly identified inhibitor Cpd-155, significantly reduces renal inflammation and injury. Thus, targeting the NAT10/CCL2/CXCL1 axis presents a promising therapeutic strategy for treating inflammatory kidney diseases.
Project description:C12orf43-NAT10 interaction was found to accelerate breast cancer progression and chemotherapy resistance by activating the BMI-P53 signaling axis in a N4-acetylcytidine-dependent manner.
Project description:Despite the advance in diagnosis and treatment, the prognosis of osteosarcoma patients remains unsatisfied. Therefore, it is imperative to identify novel therapeutic targets for osteosarcoma. Through RNA sequencing (RNA-seq) combined with functional screening, N4-acetylcytidine (ac4C) acetyltransferase 10 (NAT10) was identified as a candidate therapeutic target in osteosarcoma. Upregulated NAT10 correlated with poor prognosis in osteosarcoma patients and NAT10 knockout drastically inhibited cell proliferation and metastasis in vitro and in vivo. NAT10 enhanced mRNA stability and translation efficiency of activating transcription factor 4 (ATF4) through ac4C modification. ATF4 induced transcription of asparagine synthetase (ASNS), which catalyzes asparagine (Asn) biosynthesis. Asn promote protein and nucleotide synthesis, facilitating osteosarcoma progression. Overexpression of ATF4, ASNS or supplementation of asparagine rescue the tumor inhibitory effect of NAT10 knockout.
Project description:Despite the advance in diagnosis and treatment, the prognosis of osteosarcoma patients remains unsatisfied. Therefore, it is imperative to identify novel therapeutic targets for osteosarcoma. Through RNA sequencing (RNA-seq) combined with functional screening, N4-acetylcytidine (ac4C) acetyltransferase 10 (NAT10) was identified as a candidate therapeutic target in osteosarcoma. Upregulated NAT10 correlated with poor prognosis in osteosarcoma patients and NAT10 knockout drastically inhibited cell proliferation and metastasis in vitro and in vivo. NAT10 enhanced mRNA stability and translation efficiency of activating transcription factor 4 (ATF4) through ac4C modification. ATF4 induced transcription of asparagine synthetase (ASNS), which catalyzes asparagine (Asn) biosynthesis. Asn promote protein and nucleotide synthesis, facilitating osteosarcoma progression. Overexpression of ATF4, ASNS or supplementation of asparagine rescue the tumor inhibitory effect of NAT10 knockout.
Project description:N4-acetylcytidine (ac4C), a newly identified epigenetic modification within mRNAs, has been characterized as a crucial regulator of mRNA stability and translation efficiency. And NAT10 is the only known RNA acetyltransferase. In our study, we documented the down-regulated expression of both ac4C and NAT10 during meiotic maturation of mouse oocytes. NAT10 knockdown resulted in ac4C reduction and impaired mouse oocyte maturation in vitro. These results indicated that NAT10-mediated ac4C modification plays a critical regulatory role in oocyte meiotic maturation. We further performed high-throughput sequencing with NAT10-overexpressed HEK293T cells and NAT10-binding transcripts to investigate the genes modulated by NAT10-mediated ac4C modification.
Project description:N-acetyltransferase 10 (NAT10)-mediated N4-acetylcytidine (ac4C) modification is crucial for mRNA stability and translation efficiency, yet the underlying function in mammalian preimplantation embryos remains unclear.
Project description:N4-acetylcytidine (ac4C) is an evolutionarily conserved RNA modification that is deposited on diverse RNAs by N-acetyltransferase 10 (NAT10), a protein with high biological significance for aging and cancer. Here, we performed a comprehensive survey of ac4C using metabolic labeling, sodium cyanoborohydride chemical treatment coupled to next generation sequencing (NGS) and ac4C antibody-based cell and molecular biology techniques. Our NGS analysis confirms robust ac4C modification of rRNA and specific tRNA species in a NAT10-dependent manner, but suggests low or spurious ac4C acetylation in mRNA. Analysis of RNA-seq data also revealed an induction of inflammatory responses as well as mutagenesis at transcriptionally active sites in NAT10-KO cells. This finding led us to further explore the role of NAT10 in R-loops, which have recently been shown to induce APOBEC3B-mediated mutagenesis. Our analysis revealed that R-loops are modified with ac4C in a NAT10-dependent manner. Furthermore, NAT10 restrains the levels of R-loops at a subset of differentially expressed genes in a manner dependent on its catalytic activity. Together with cellular biology data showing ac4C containing RNA in endosomal structures, we propose that increased levels of ac4C-unmodified RNAs, likely derived from R-loops, in endosomal structures induce inflammatory responses.
Project description:N4-acetylcytidine (ac4C) is an evolutionarily conserved RNA modification that is deposited on diverse RNAs by N-acetyltransferase 10 (NAT10), a protein with high biological significance for aging and cancer. Here, we performed a comprehensive survey of ac4C using metabolic labeling, sodium cyanoborohydride chemical treatment coupled to next generation sequencing (NGS) and ac4C antibody-based cell and molecular biology techniques. Our NGS analysis confirms robust ac4C modification of rRNA and specific tRNA species in a NAT10-dependent manner, but suggests low or spurious ac4C acetylation in mRNA. Analysis of RNA-seq data also revealed an induction of inflammatory responses as well as mutagenesis at transcriptionally active sites in NAT10-KO cells. This finding led us to further explore the role of NAT10 in R-loops, which have recently been shown to induce APOBEC3B-mediated mutagenesis. Our analysis revealed that R-loops are modified with ac4C in a NAT10-dependent manner. Furthermore, NAT10 restrains the levels of R-loops at a subset of differentially expressed genes in a manner dependent on its catalytic activity. Together with cellular biology data showing ac4C containing RNA in endosomal structures, we propose that increased levels of ac4C-unmodified RNAs, likely derived from R-loops, in endosomal structures induce inflammatory responses.
Project description:N4-acetylcytidine (ac4C) is an evolutionarily conserved RNA modification that is deposited on diverse RNAs by N-acetyltransferase 10 (NAT10), a protein with high biological significance for aging and cancer. Here, we performed a comprehensive survey of ac4C using metabolic labeling, sodium cyanoborohydride chemical treatment coupled to next generation sequencing (NGS) and ac4C antibody-based cell and molecular biology techniques. Our NGS analysis confirms robust ac4C modification of rRNA and specific tRNA species in a NAT10-dependent manner, but suggests low or spurious ac4C acetylation in mRNA. Analysis of RNA-seq data also revealed an induction of inflammatory responses as well as mutagenesis at transcriptionally active sites in NAT10-KO cells. This finding led us to further explore the role of NAT10 in R-loops, which have recently been shown to induce APOBEC3B-mediated mutagenesis. Our analysis revealed that R-loops are modified with ac4C in a NAT10-dependent manner. Furthermore, NAT10 restrains the levels of R-loops at a subset of differentially expressed genes in a manner dependent on its catalytic activity. Together with cellular biology data showing ac4C containing RNA in endosomal structures, we propose that increased levels of ac4C-unmodified RNAs, likely derived from R-loops, in endosomal structures induce inflammatory responses.
Project description:Increased differentiation or activity of osteoclasts is the key pathogenic factor of postmenopausal osteoporosis (PMOP). N4‐acetylcytidine (ac4C) modification, catalyzed by Nat10, is a novel post-transcriptional mRNA modification related to many diseases. However, its impact on regulating osteoclast activation in PMOP remains uncertain. Here, we initially observed that Nat10-mediated ac4C positively correlates with osteoclast differentiation of monocytes and low bone mass in PMOP. The specific knockout of Nat10 in monocytes and remodelin, a Nat10 inhibitor, alleviates ovariectomized (OVX)-induced bone loss by downregulating osteoclast differentiation. Mechanistically, epitranscriptomic analyses reveal that the nuclear factor of activated T cells cytoplasmic 1 (Nfatc1) is the key downstream target of ac4C modification during osteoclast differentiation. Subsequently, translatomic results demonstrate that Nat10-mediated ac4C enhances the translation efficiency of Nfatc1, thereby inducing Nfatc1 expression and consequent osteoclast maturation. Cumulatively, these findings reveal the promotive role of Nat10 in osteoclast differentiation and PMOP from a novel field of RNA modifications and suggest that Nat10 can be a target of epigenetic therapy for preventing bone loss in PMOP.