Silencing of the tRNA Modification Enzyme Cdkal1 Effects Functional Insulin Synthesis in NIT-1 Cells: tRNALys3 Lacking ms2- (ms2t6A37) is Unable to Establish Sufficient Anticodon:Codon Interactions to Decode the Wobble Codon AAG

Abstract

Human Genome Wide Association Studies found a significant risk of Type 2 Diabetes Mellitus (T2DM) in single nucleotide polymorphisms in the cdkal1 gene. The cdkal1 gene is remote from the insulin gene and with the surprising function of a specific tRNA modification. Population studies and case control studies acquired evidences of the connection between Cdkal1 protein and insulin production over the years. To obtain biochemical proofs directly linking potential SNPs to their roles in insulin production and availability is challenging, but the development of Cdkal1 knock out mice and knock out cell lines made it possible to extend our knowledge towards therapeutic field of diabetic research. Supporting the evidences, here we show that knock down of the cdkal1 gene using small interfering and short hairpin RNA in the NIT-1 cell line, a β-cell line inducible for insulin resulted in reduced levels of cdkal1 and mature insulin mRNAs, increased the level of precursor insulin mRNA, decreased Cdkal1 and insulin proteins, and diminished modification of tRNALys3 from t6A37 to ms2t6A37, the specified function of Cdkal1. tRNALys3 lacking ms2- is incapable of establishing sufficient hydrogen bonding energy and hydrophobic stabilization to decode the wobble codon AAG.

The Association between CDKAL1 Gene rs10946398 Polymorphism and Post-Transplant Diabetes in Kidney Allograft Recipients Treated with Tacrolimus

“…CDKAL1 plays a key role in post-translational modification and insulin secretion, as well as in the differentiation and development of pancreatic β-island cells. In mice lacking the CDKAL1 gene, pancreatic β-cells produced reduced amounts of insulin after glucose stimulation [ 28 ]. CDKAL1 affects the proper differentiation and function of pancreatic β-cells and thus insulin secretion.…”

Elucidating the influence of RNA modifications and Magnesium ions on tRNAPheconformational dynamics inS. cerevisiae: Insights from Replica Exchange Molecular Dynamics simulations

“…The role of modifications in tRNA has been previously investigated by targeting a single or a few modifications or by comparing the fully modified and the unmodified tRNA (15). Examples include RNA modifications & Magnesium in tRNA Dynamics ASL modifications that affect translation like Queuosine (Q) at position 37 of tRNA His in Escherichia coli (8), 2-thiocytidine (s 2 C), and Inosine (I) on tRNA Arg in Saccharomyces cerevisiae (16), lysidine (L) or granulation at position 34 and 2-methylthio-N6-threonylcarbamoyladenosine (ms 2 t 6 A), on tRNA Lys (17)(18)(19), mcm 5 U at position 34 of tRNA Asp in humans (20) and Wybutosine (yW), located at position 37 on tRNA Gly (9) However, structures and biochemical assays of unmodified tRNAs demonstrate that PTMs do not alter the overall L-shaped structure of the tRNA but they are essential to locally orient the nucleotides to maintain functionally active form for various biological processes (21,22). In addition to current experimental techniques, molecular dynamics simulations have emerged as a powerful tool to reveal the mechanistic details of how these PTMs collectively interact and influence RNA dynamics and structure (23).…”