Structure of rat calmodulin processed genes with implications for a mRNA-mediated process of insertion

Abstract

Two distinct processed calmodulin genes of rat (λSC8 and λSC9) were identified, cloned and their DNA sequences determined. The existence of direct repeats of 19 base-pairs for λSC8 or 9 base-pairs for λSC9 at both ends of the coding plus non-coding regions suggested a possible involvement of a mRNA-mediated process of insertion. Total genomic Southern hybridization suggested the existence of at least three different calmodulin-related genes in the rat genome. The other gene was the bona fide calmodulin gene (λSC4) which was split into at least five exons. λSC9 contained insertions of one nucleotide and two 17 base-pair direct repeats in the coding region. These insertions cause frameshift mutations probably preventing it from encoding a functional calmodulin. It also carried an insertion of a rat middle repetitive sequence, identifier sequence (IDS: Sutcliffe et al., 1982) in the 3′-non-coding region. Otherwise, it consisted of an almost identical DNA sequence to that of the bona fide calmodulin gene (λSC4), including the 3′-non-coding region down to the poly(A) recognition signal, A-A-T-A-A-A. On the other hand, λSC8 did not possess frameshift mutations in the coding region, and hence was capable of encoding a functional protein. In fact, a probe specific to the λSC8 sequence identified a band in Northern blotting whose size was 300 nucleotides smaller than that of authentic calmodulin mRNA. Comparison of the nucleotide sequences showed that only the coding regions of these two processed genes were homologous, indicating that the divergence of these two processed genes from the common ancestor calmodulin was an ancient event.