Publications (2)2.34 Total impact
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Article: Overproduction and functional analysis of DNA primase subunits from yeast and mouse
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ABSTRACT: Eukaryotic DNA primases are composed of two distinct subunits of 48–50 and 58–60 kDa. The amino acid sequences derived from the nucleotide sequences of the cloned genes are known only for the yeast and mouse polypeptides, and the extensive homology between the corresponding mouse and yeast subunits suggests conservation of functional domains. We were able to express in Saccharomyces cerevisiae the homologous and mouse primase-encoding genes under the control of both the constitutive ADH1 and the inducible GAL1 strong promoters, thus obtaining strains producing relevant amounts of the different polypeptides. In vivo complementation studies showed that neither one of the wild-type mouse primase-encoding encoding genes was able to rescue the lethal or temperature-sensitive phenotype caused by mutations in the yeast PRI1 or PRI2 genes, indicating that these proteins, even if structurally and functionally very similar, might be involved in critical species-specific interactions during DNA replication.Gene 113(2):199-205. · 2.34 Impact Factor -
Article: De novo synthesis of budding yeast DNA polymerase alpha and POL1 transcription at the G<sub>1</sub>/S boundary are not required for entrance into S phase
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ABSTRACT: The POL1 gene, encoding DNA polymerase α(pol α) in Saccharomyces cerevisiae , is transiently transcribed during the cell cycle at the G<sub>1</sub>/S phase boundary. Here we show that yeast pol α is present at every stage of the cell cycle, and its level only slightly increases following the peak of POL1 transcription. POL1 mRNA synthesis driven by a GAL1 promoter can be completely abolished without affecting the growth rate of logarithmically growing yeast cultures for several cell divisions, although the amount of the pol α polypeptide drops below the physiological level. Moreover, α-factor-arrested cells can enter S phase and divide synchronously even if POL1 transcription is abolished. These results indicate that the level of yeast pol α is not rate limiting and de novo synthesis of the enzyme is not required for entrance into S phase.
