To assess the role of eukaryotic DNA primase in vivo, we have produced conditional and lethal point mutations by random in vitro mutagenesis of the PRI1 and PRI2 genes, which encode the small and large subunits of yeast DNA primase. We replaced the wild-type copies of PRI1 and PRI2 with two pri1 and two pri2 conditional alleles. When shifted to the restrictive temperature, these strains showed altered DNA synthesis and reduced ability to synthesize high molecular weight DNA products, thus providing in vivo evidence for the essential role of DNA primase in eukaryotic DNA replication. Furthermore, mapping of the mutations at the nucleotide level has shown that the two pri1 and two pri2 conditional alleles and one pri2 lethal allele have suffered single base-pair substitutions causing a change in amino acid residues conserved in the corresponding mouse polypeptide.
|Titolo:||Mutations in conserved yeast DNA primase domains impair DNA replication in vivo|
PLEVANI, PAOLO (Ultimo)
|Parole Chiave:||DNA Primase; Centrifugation, Density Gradient; DNA, Fungal; Base Sequence; Alleles; Genes, Fungal; DNA Mutational Analysis; Molecular Sequence Data; Amino Acid Sequence; RNA Nucleotidyltransferases; DNA Replication; Saccharomyces cerevisiae|
|Settore Scientifico Disciplinare:||Settore BIO/11 - Biologia Molecolare|
|Data di pubblicazione:||1-mag-1991|
|Digital Object Identifier (DOI):||10.1073/pnas.88.9.3877|
|Appare nelle tipologie:||01 - Articolo su periodico|