Spt16 and Pob3 of Saccharomyces cerevisiae form an essential, abundant heterodimer that is nuclear, chromatin-associated, and copurifies with DNA polymerase alpha.
Previously we showed that the yeast proteins Spt16 (Cdc68) and Pob3 are physically associated, and interact physically and genetically with the catalytic subunit of DNA polymerase alpha, Pol1 [Wittmeyer and Formosa (1997) Mol. Cell. Biol. 17, 4178-4190]. Here we show that purified Spt16 and Pob3 form a stable, abundant, elongated heterodimer and provide evidence that this is the functional form of these proteins. Genetic interactions between mutations in SPT16 and POB3 support the importance of the Spt16-Pob3 interaction in vivo. Spt16, Pob3, and Pol1 proteins were all found to localize to the nucleus in S. cerevisiae. A portion of the total cellular Spt16-Pob3 was found to be chromatin-associated, consistent with the proposed roles in modulating chromatin function. Some of the Spt16-Pob3 complex was found to copurify with the yeast DNA polymerase alpha/primase complex, further supporting a connection between Spt16-Pob3 and DNA replication.
Department of Biochemistry, University of Utah, Salt Lake City 84132, USA.
J
J
Wittmeyer
L
L
Joss
T
T
Formosa
D000123:Acetyltransferases / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002843:Chromatin / Q000378:metabolism*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004256:DNA Polymerase I / Q000302:isolation & purification* / Q000378:metabolism; D019281:Dimerization; D017886:Durapatite; D004789:Enzyme Activation; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D051548:Histone Acetyltransferases; D015151:Immunoblotting; D046911:Macromolecular Substances; D009687:Nuclear Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010441:Peptide Chain Elongation, Translational; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002843:Chromatin; D005656:Fungal Proteins; D046911:Macromolecular Substances; D009687:Nuclear Proteins; C106768:POB3 protein, S cerevisiae; C071238:SPT16 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; D017886:Durapatite; D000123:Acetyltransferases; D051548:Histone Acetyltransferases; D004256:DNA Polymerase I; D000251:Adenosine Triphosphatases