The regulation of insulin secretory granule protein production by hnRNP A2/B1

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Post-transcriptional dynamic assembly of mRNAs with RNA-binding proteins (RBPs) into ribonucleoprotein complexes (RNPs) fine-tunes gene expression. Specifically, RBPs regulate the splicing, export, storage, translation and degradation of each individual mRNA, including its targeting to polysomes, the endoplasmic reticulum or cytoplasmic RNA granules. Moreover, binding of RBPs to motifs shared among functionally related mRNAs allows for the coordinated expression of the latter. Pancreatic islet beta cells exploit post-transcriptional mechanisms to quickly adjust their insulin production relative to glycaemia. Hyperglycaemia rapidly enhances the biosynthesis of insulin and other insulin secretory granule cargoes, e.g. PC1/3, PC2 and ICA512, without initially affecting the corresponding mRNAs levels. We discovered that in resting MIN6 cells a common set of RBPs binds the 5'-UTRs of Insulin1, Insulin2, spliced Insulin2, PC2 and ICA512 mRNAs, while another set of shared RBPs coordinate the translation of these mRNAs upon glucose stimulation. A novel RBP shared among all tested mRNAs for insulin secretory granule cargoes is hnRNP A2/B1. We mapped hnRNP A2/B1-binding sites in the 5'-UTR of Insulin1 mRNA and detected reduced hnRNP A2/B1 binding upon their mutation. Furthermore, Hnrnpa2b1 /- MIN6 cells displayed reduced Insulin1 mRNA and total insulin protein levels compared to WT cells. Finally, we found that in resting MIN6 cells in culture and in human beta cells in situ hnRNP A2/B1 is enriched in cytoplasmic RNA granules. Here we propose to develop further these studies to better understand the physiology and pathophysiology of beta cells. The first aim is to elucidate how hnRNP A2/B1 controls the dynamic expression of mRNAs for insulin and other secretory granule proteins. Next, we shall study RNP dynamics with FLIM-FRET imaging using RNAs tagged with RNA aptamers and fluorescent RBPs in order to decipher the RNP code of mRNAs for insulin granule proteins in a space- and time-resolved manner. Finally, we shall investigate hnRNP A2/B1’s role in human beta cells and if the presence of hnRNP A2/B1 positive RNA granules correlates with beta cell dysfunction in diabetes.