HUMAN RAB11A - TRANSCRIPTION, CHROMOSOME MAPPING AND EFFECT ON THE EXPRESSION LEVELS OF HOST GTP-BINDING PROTEINS

Rab11a is a member of the rab-branch of the ras-like small GTP-binding protein superfamily that is associated with both constitutive and reg ulated secretory pathways. Using a direct procedure for cDNA cloning o f small ras-related GTPases, that is based on the screening of eukaryo tic cDNA expression libraries using [alpha-P-32]GTP as a probe, we hav e isolated two cDNA clones encoding rab11a, Both clones share identica l coding sequences, but differ in the length and sequence of their 3' untranslated regions (3'-UTR). Northern blot hybridisation analysis of carious human tissues revealed indeed two mRNA species with lengths o f 1.0 and 2.3 kb, respectively. Sequence analysis of the cDNAs identif ied two different putative polyadenylation signals (AATAAA) at positio ns 927 and 2302 of the large's transcript. In addition, the 3'-UTR of the larger transcript exhibited several AU-rich elements (ARE) that ar e believed to control gene expression by regulating the rate of mRNA d egradation. Southern blots of human DNA digested with several rare res triction enzymes, and separated by pulse-field gel electrophoresis, yi elded the same macro-restriction fragment pattern when hybridised with probes that discriminate between the two transcripts. Taken together, these findings imply that the two mRNA species originate from a singl e gene, which we have mapped to 15q21.3-q22.31, by the use of diferent polyadenylation sites. As expected, both rab11a-cDNAs yielded the sam e protein product when transiently expressed in COS-1 cells, and surpr isingly, upregulated the proteome expression profile (de novo synthesi s or posttranslational modification of preexisting proteins) of a few other, yet unknown GTP-binding proteins. (C) 1998 Federation of Europe an Biochemical Societies.