Molecular cloning and characteristics of rpc19+ and rpc40+ Schizosaccharomyces pombe genes, coding for common subunits of nuclear RNA polymerase I and III

Full-length copies of cDNAs of the rpc19+ and rpc40+ genes encoding the common subunits of nuclear RNA polymerases I and III and the corresponding fragments of chromosomes were isolated from genomic and cDNA libraries of Schizosaccharomyces pombe and characterized. It was established that the cloned genes are located on chromosomes III and II of the fission yeast, respectively. The rpc40+ gene lacks introns, and the rpc19+ gene contains two intervening sequences. The comparison of subunits Rpc19 (125 aa; M 13 722 Da; pI 4.51) and Rpc40 (348 aa; M 39 141 Da; pI 5.40) of Sz. pombe, whose characteristics were deduced from the sequences of their cDNAs, with the orthologous components of other eukaryotes allowed the most conserved structure-functional domains of these proteins to be identified.     

Distinct regions of RPB11 are required for heterodimerization with RPB3 in human and yeast RNA polymerase II

In Saccharomyces cerevisiae, RNA polymerase II assembly is probably initiated by the formation of the RPB3–RPB11 heterodimer. RPB3 is encoded by a single copy gene in the yeast, mouse and human genomes. The RPB11 gene is also unique in yeast and mouse, but in humans a gene family has been identified that potentially encodes several RPB11 proteins differing mainly in their C-terminal regions. We compared the abilities of both yeast and human proteins to heterodimerize. We show that the yeast RPB3/RPB11 heterodimer critically depends on the presence of the C-terminal region of RPB11. In contrast, the human heterodimer tolerates significant changes in RPB11 C-terminus, allowing two human RPB11 variants to heterodimerize with the same efficiency with RPB3. In keeping with this observation, the interactions between the conserved N-terminal ‘α-motifs’ is much more important for heterodimerization of the human subunits than for those in yeast. These data indicate that the heterodimerization interfaces have been modified during the course of evolution to allow a recent diversification of the human RPB11 subunits that remains compatible with heterodimerization with RPB3.     

New Genes on Human Chromosome 7: Bioinformation Analysis of a Cluster of Genes from the POLR2J Family

Four independent genes encoding various variants of the hRPB11 subunit of Homo sapiens RNA polymerase II were revealed in human chromosome 7. Three genes (POLR2 J1, POLR2 J2, and POLR2 J3) form a cluster of total length of 214 530 bp in the genetic locus 7q22.1 on the long arm of chromosome 7 (contig NT_007933). The fourth gene (POLR2 J4, 31 040 bp) was localized in the cytogenetic locus 7p13 of the short arm of chromosome 7 (contig NT_007819). An analysis enabled us to refine dissimilar experimental data on the mapping of the hRPB11 subunit gene on chromosome 7. In particular, the presence of three sites of its localization according to data on hybridization with fluorescent-labeled probes (the FISH method) was explained. It was established that, upon the expression of the four human POLR2 J genes, at least 14 types of mature mRNAs encoding somewhat differing hRPB11 isoforms can be synthesized. Eleven of these mRNAs were revealed (as full-length copies or clearly identifiable fragments) in the available databases of expressed sequence tags and cDNAs. The most probable scheme of origination of the multiple genes of the POLR2 J family as a result of three consecutive segmented duplications increasing in size was proposed and substantiated. On the basis of the scheme, some assumptions on the pathways of evolution of separate human genes and the mechanisms of generation of protein diversity in higher eukaryotes were made.