The structure of rat preproinsulin genes

Abstract

In rat there are two nonallelic insulins, I and II. We have cloned and sequenced double stranded cDNA copies of both preproinsulin mRNA I and II. Using the cloned sequence as probe, we established by the Southern blotting technique a restriction map of the two chromosomal genes. This map indicates that an intron exists within the insulin II gene. To examine this in more detail, we have isolated both genes from a library of rat DNA cloned in phage lambda. Restriction endonuclease analysis and direct DNA sequencing revealed that gene II contains two introns: a 490 base pair intron between the region encoding amino acids 38 and 39 of proinsulin, and a 119 base pair intron, which is 17 base pairs upstream from the initiation codon. Gene I is not interrupted within the protein coding region, but possesses an intron homologous to the 119 base pair intron of insulin II. We are studying the structure of insulin genes from other species to determine if the 490 base pair intron was lost or inserted in the duplicated gene. We have identified nuclear RNA molecules larger than preproinsulin mRNA which contain the transcribed intronic sequences. These molecules represent a new precursor in insulin biosynthesis.