Repetitious structure and transcription control of a polyubiquitin gene in Volvox carteri

Abstract

Southern analysis indicated the presence of at least four ubiquitin gene loci in the Volvox carteri genome. Three of these, a polyubiquitin gene described here and a non-segregating ubiquitin gene pair, were assigned to two different linkage groups by RFLP mapping; the non-polymorphic fourth gene locus remained unassigned. The polyubiquitin gene was cloned and its 2,116-bp sequence determined. It contains six exons each interrupted by an intron at Gly35, and it encodes a pentameric polyubiquitin polypeptide consisting of five runs of 76 identical amino-acid residues and a C-terminal extension of one leucine. The five tandem repeats of coding units plus introns exhibit an unusually high degree of overall sequence identity indicating an efficient process of gene homogenization in this region of the V. carteri genome. S1 mapping revealed two closely-spaced transcription starts, 24 and 28 nucleotides downstream from a putative TATA sequence. Preceding the TATA box are two 14-bp conserved heat-shock elements (HSEs) and two octameric sequences closely resembling an yesat HSE. Consistent with a 1.6-kb transcript seen on Northern blots are two polyadenylation signals (TGTAA) located 99 bp and 169 bp downstream from the TGA translational stop. The polyubiquitin gene was transcribed throughout the Volvox life cycle with peaks in the 1.6-kb mRNA levels during pre-cleavage, cleavage, and post-inversion. In contrast, an 0.6-kb monoubiquitin transcript was abundant only at the pre-cleavage stage suggesting a different type of gene control. Heat shock increased the level of polyubiquitin mRNA, whereas the level of monoubiquitin mRNA was down-regulated.