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

Human Genome Wide Association Studies found a significant risk of Type 2 Diabetes Mellitus (T2DM) in single nucleotide polymorphisms in the gene. The 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 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 and mature insulin mRNAs, increased the level of precursor insulin mRNA, decreased Cdkal1 and insulin proteins, and diminished modification of tRNA from tA to mstA, the specified function of Cdkal1. tRNA lacking ms- is incapable of establishing sufficient hydrogen bonding energy and hydrophobic stabilization to decode the wobble codon AAG.