Project description:β-cell specific Mettl14 knock-out mice display reduced N6-methyladenosine (m6A) levels and recapitulate human Type II diabetes (T2D) islet phenotype with early diabetes onset and mortality secondary to decreased β-cell proliferation and insulin degranulation. To gain insights into the role of m6A in regulating the IGF1/insulin -> AKT - > PDX1 pathway and to dissect the signaling networks modulating AKT phosphorylation, we subjected freshly isolated islets from control and Mettl14 knock-out mice to phospho-antibody microarrays.
Project description:m6A regulates virtually every step in RNA metabolism. However, its toles in limb development remains largely unknown. To understand the roles, we created a limb bud-specific conditional knockout (cKO) mice and control heterozygous (cHet) mice of the Mettl14 gene, which encodes an essential subunit in the m6A methyltransferase complex METTL3/METTL14. We harvested limb buds from the mice on E12.5 and applied the proteins to quantitative mass spectrometry to understand how the depletion of Mettl14 affected the proteomes.
Project description:N6-methyladenosine (m6A) is a widespread RNA modification that plays a crucial role in regulating gene expression, affecting processes such as RNA stability, splicing, and translation. I Previous studies have shown that m6A is involved in fine-tuning β-cell function, but its impact on α-cells, which secrete glucagon in response to low glucose and amino acids, has not been fully investigated. Here, we show that METTL14, a key m6A writer, is essential for maintaining α-cell function and identity. Using a combination of in vitro and in vivo models, we demonstrate that metabolic stimuli such as low glucose and amino acids upregulate METTL14, enhancing glucagon secretion, while insulin downregulates it. Importantly, α-cell-specific knockout of Mettl14 in mice results in impaired glucagon secretion, increased insulin levels, and a surprising conversion of α-cells into β-cells. These findings reveal that m6A is a critical regulator of α-cell identity and function, expanding our understanding of RNA modifications in islet biology. By uncovering a role for METTL14 in maintaining α-cell integrity, our study opens new avenues for exploring RNA-based therapies in diabetes and other metabolic disorders.
Project description:Mechanisms driving sex differences across islet cells is unknown. Thus, studying sex differences in islet regulation and function represent a unique avenue to understand the sex-specific heterogeneity in β cell failure in diabetes. We examined sex and race differences in human pancreatic islets from 15 donors using an orthogonal series of experiments including single cell RNA-seq (scRNA-seq), single nucleus assay for transposase-accessible chromatin sequencing (snATAC-seq), dynamic hormone secretion, and bioenergetics.
Project description:Post-transcriptional m6A methylation on mRNA plays a key role in neural development. Here, we specifically depleted Mettl14 (a key component of m6A complex) in retina progenitor cells by crossing Mettl14 fl/fl mice with the Chx10-Egfp/Cre mouse line and investigated cell type and transcriptome changes in the developing retinas using scRNA-seq.
Project description:We profiled the transcritpome and ATAC profiles of human pancreatic islets generated from pluripotent stem cells. Multiomic profiling was also performed on primary human islets and in vivo matured SC-islets for comparision. We catalogued the ATAC associated signatures for each cell types in SC-islets and compared them to their human primiary islet counterparts. In vivo maturation of SC-islets were also compared with in vitro SC-islets. In this study, we identified key regulators associated with islet identity during differentiation and maturation. Gene manipulation of CTCF affects differentiating SC-islet cell fate to enteroendocrine-like lineage. ARID1B knockdown caueses islet cells to present mature signatures. These gene altered SC-islets were also sequenced.
Project description:C57BL/6J mice were subjected to 30 Gy liver irradiation (5 Gy × 6 fractions). METTL14 overexpression was achieved via AAV-METTL14 tail-vein injection. Liver tissues were collected for RNA-seq analysis to identify differentially expressed genes (DEGs) related to cell death pathways.
Project description:Zfp92, a repressive KRAB domain-containing zinc-finger protein, was identified by Gene Co-expression Network analysis to be an interesting candidate gene involved in endocrine specification and maturation. We examined the role of Zfp92, a KRAB-ZFP that is highly expressed in pancreatic islets of adult mice, by analyzing global Zfp92 knockout (KO) mice. Adult Zfp92 KO animals exhibited only mild changes in glucose homeostasis and no change in islet structure, although, male KO mice exhibited decreased growth, and female KO mice exhibited increased body fat accumulation on a high fat diet. We found that Zfp92 regulates a subset of transposable elements as well as Mafb, a transcription factor involved in islet development.
