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Chu, Chieh Min

University of British Columbia

Diabetes affects around 500 million people worldwide. Diabetes is marked by raised blood sugar levels owing to a relative or absolute lack of insulin, which is produced in pancreatic islet beta-cells. Pancreatic beta-cells are heterogeneous, with many studies reporting functional and morphological differences. However, it is unclear whether insulin gene activity is also heterogeneous in beta-cells and whether this heterogeneity is static or dynamic. We utilized a mouse model in which the endogenous Ins2 gene has been replaced with GFP (Ins2GFP), to study endogenous insulin gene activity. We found that beta-cells can be found in two states, Ins2(GFP)HIGH and Ins2(GFP)LOW. We also found that gene activity at the Ins2 loci was dynamic, and that cells could transition between the two beta-cell states over time. We profiled the two cell states at the transcriptome and proteome levels, and perturbed beta-cell functions using a small molecule screen to determine the mechanisms behind Ins2 dynamics and cell state transitions. We found that the Ins2(GFP)HIGH state was associated with increased beta-cell maturity factors and increased fragility, and cAMP pathways were the strongest effectors of Ins2 gene activity. We next investigated INS gene expression using an INS-EGFP stem cell-derived beta-cell line. Similar to the studies using beta-cells from Ins2GFP mice, we found that INS gene expression was bimodal and dynamic over time in stem cell-derived beta-like cells. We interrogated the two INS gene expression states with a combination of transcriptomics and proteomic analyses. We found that cells with high EGFP expression had increased mRNA expression and protein abundance in the endoplasmic reticulum (ER) and Golgi. To understand the relationship between INS gene dynamics and beta-cell survival, we also performed a high-throughput screen of prioritized ligands, curated from a combination of proteomic and transcriptomic data. We found that several factors, namely FGF5, FGF19 and FGF8F had protective effects on SC beta-cells. Overall, our studies defined the heterogeneity in insulin gene activity in both mice and SC beta-cell models and reveal pathways which modulate insulin production. These observations may inform new cell therapies for diabetes.

Text

2024-08-28

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/89073

Cell and Developmental Biology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/