Multiplicity of glutathione S-transferase genes in the rat and association with a type 2 Alu repetitive element

Abstract

Southern blot analysis of rat genomic DNA using glutathione S-transferase Ya and Yc cDNA probes was employed to estimate the size of the Ya/Yc multigene family. A minimum of five to seven Ya/Yc genes were detected; at least two of these are Yc genes. The presence of multiple genes was further supported by the isolation of three nonoverlapping genomic clones from a rat EcoRI library that hybridized to a Ya cDNA clone, pGTB38. However, not all EcoRI bands seen in genomic blots were represented in the clones, suggesting that not all Ya/Yc genes have been isolated. The organization of a Ya gene in one of these EcoRI genomic clones, .lambda.GTB38-3, and an overlapping clone, .lambda.GTB45-1, isolated from a HaeIII library, was investigated with 5'' and 3'' probes prepared from Ya and Yc cDNA clones. Restriction endonuclease mapping and hybridization studies revealed that the gene spans over 10 kilobases and contains at least three introns. Sequences upstream from the 5'' untranslated region of the gene, and within an intron in the 5'' coding region, were found to contain sequences homologous to a type 2 Alu repetitive element from the rat growth hormone gene [Page, G.S., Smith, S., and Goodman, H.M. (1981) Nucleic Acids Res. 9, 2087-2104]. The repetitive sequences in .lambda.GTB38-3 were identified by hybridization to a novel Ya cDNA clone, pGTB45. This cDNA clone was isolated from a cDNA library previously described [Telakowski-Hopkins, C.A., Rodkey, J.A., Bennett, C.D., Lu, A. Y. H., and Pickett, C.B. (1985) J. Biol. Chem. 260, 5820-5825] with nick-translated intron sequences as probes. pGTB45 is virtually identical with pGTR261 [Tu, C.-P.D., Lai, H.-C. J., Li, N.-Q., Weiss, M.J., and Reddy, C. C. (1984) J. Biol. Chem. 259, 9434-9439], except that the 3'' untranslated region extends 231 base pairs beyond the polyadenylation signal of pGTR261. This elongated 3'' untranslated sequence is unique in that it contains a full-length type 2 Alu repetitive element, which includes two additional, overlapping polyadenylation signals.