Rpo26p, a subunit common to yeast RNA polymerases, is essential for the assembly of RNA polymerases I and II and for the stability of the largest subunits of these enzymes.
Eukaryotic nuclear RNA polymerases (RNAPs) are composed of two large subunits and a number of small polypeptides, some of which are common among these enzymes. To understand the function of Rpo26p, one of the five subunits common to yeast RNAPs, 34 different mutations have been isolated in RP026 that cause cell death in a strain carrying a temperature-sensitive (ts) mutation in the gene (RP021) encoding the largest subunit of RNAPII. These mutant alleles were grouped into three phenotypic classes (null, ts, and neutral) on the basis of the phenotype they imposed in combination with wild-type RP021. The function of Rpo26p was addressed by biochemical analysis of the ts rpo26-31 allele. The steady-state level of rpo26-31p was reduced at high temperature; this was accompanied by a decrease in the level of at least two other subunits, the largest subunits of RNAPI (A190p) and RNAPII (Rpo21p). Pulse-chase metabolic labeling and immunoprecipitation of RNAPII showed that at high temperature, rpo26-31 did not lead to dissociation of Rpo26p from the polymerase but prevented the assembly of RNAPII. Overexpression of rpo26-31 partially suppressed the ts phenotype and led to accumulation of the mutant subunit. However, overexpression only marginally suppressed the assembly defect of RNAPII. Furthermore, A190p and Rpo21p continued to accumulate at low levels under these conditions. We suggest that Rpo26p is essential for the assembly of RNAPI and RNAPII and for the stability of the largest subunits of these enzymes.
Nouraini
S
S
Department of Genetics, Hospital for Sick Children, Toronto, Canada.
Archambault
J
J
Friesen
J D
JD
D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D005656:Fungal Proteins / Q000096:biosynthesis* / Q000737:chemistry; D015870:Gene Expression; D005800:Genes, Fungal; D005838:Genotype; D006801:Humans; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010641:Phenotype; D010957:Plasmids; D012318:RNA Polymerase I / Q000096:biosynthesis* / Q000737:chemistry; D012319:RNA Polymerase II / Q000096:biosynthesis* / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D017386:Sequence Homology, Amino Acid; D013696:Temperature
D005656:Fungal Proteins; D046911:Macromolecular Substances; D012319:RNA Polymerase II; D012318:RNA Polymerase I