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PMID: 8355715 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

SPK1 is an essential S-phase-specific gene of Saccharomyces cerevisiae that encodes a nuclear serine/threonine/tyrosine kinase.

Molecular and cellular biology ·Vol. 13 ·No. 9 ·1993-09-00 ·Pages 5829-42

Zheng P, Fay DS, Burton J, Xiao H, Pinkham JL, Stern DF

Abstract

SPK1 was originally discovered in an immunoscreen for tyrosine-protein kinases in Saccharomyces cerevisiae. We have used biochemical and genetic techniques to investigate the function of this gene and its encoded protein. Hybridization of an SPK1 probe to an ordered genomic library showed that SPK1 is adjacent to PEP4 (chromosome XVI L). Sporulation of spk1/+ heterozygotes gave rise to spk1 spores that grew into microcolonies but could not be further propagated. These colonies were greatly enriched for budded cells, especially those with large buds. Similarly, eviction of CEN plasmids bearing SPK1 from cells with a chromosomal SPK1 disruption yielded viable cells with only low frequency. Spk1 protein was identified by immunoprecipitation and immunoblotting. It was associated with protein-Ser, Thr, and Tyr kinase activity in immune complex kinase assays. Spk1 was localized to the nucleus by immunofluorescence. The nucleotide sequence of the SPK1 5' noncoding region revealed that SPK1 contains two MluI cell cycle box elements. These elements confer S-phase-specific transcription to many genes involved in DNA synthesis. Northern (RNA) blotting of synchronized cells verified that the SPK1 transcript is coregulated with other MluI box-regulated genes. The SPK1 upstream region also includes a domain highly homologous to sequences involved in induction of RAD2 and other excision repair genes by agents that induce DNA damage. spk1 strains were hypersensitive to UV irradiation. Taken together, these findings indicate that SPK1 is a dual-specificity (Ser/Thr and Tyr) protein kinase that is essential for viability. The cell cycle-dependent transcription, presence of DNA damage-related sequences, requirement for UV resistance, and nuclear localization of Spk1 all link this gene to a crucial S-phase-specific role, probably as a positive regulator of DNA synthesis.

Related Genes
PHR1 RAD2 RAD23 RAD7 SPK1
MeSH Terms
Base Sequence Cell Cycle Proteins Checkpoint Kinase 2 Chromosome Mapping DNA Repair Fungal Proteins/metabolism Gene Expression Gene Expression Regulation, Fungal/radiation effects Genes, Fungal Molecular Sequence Data Mutagenesis, Insertional Nuclear Proteins/metabolism Oligodeoxyribonucleotides/chemistry Promoter Regions, Genetic Protein Kinases/metabolism Protein Serine-Threonine Kinases RNA, Messenger/genetics Restriction Mapping S Phase Saccharomyces cerevisiae/enzymology Saccharomyces cerevisiae Proteins Sequence Alignment Substrate Specificity
Chemicals
Cell Cycle Proteins Fungal Proteins Nuclear Proteins Oligodeoxyribonucleotides RNA, Messenger Saccharomyces cerevisiae Proteins Protein Kinases Checkpoint Kinase 2 Protein Serine-Threonine Kinases RAD53 protein, S cerevisiae
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Zheng P
Department of Pathology, Yale University School of Medicine, New Haven, Connecticut 06510.
Fay D S
Burton J
Xiao H
Pinkham J L
Stern D F
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.13.9.5829-5842.1993
Published
1993-09-00
Pages
5829-42
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC360328
Subset
IM
Grants
NCRR NIH HHS · 2-SO7-RR058 · United States
PHS HHS · 9MO7223-16 · United States
NCI NIH HHS · CA45708 · United States
External Links
PubMed source DOI: 10.1128/mcb.13.9.5829-5842.1993 PMC Full Text View journal details
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