The Reg1-interacting proteins, Bmh1, Bmh2, Ssb1, and Ssb2, have roles in maintaining glucose repression in Saccharomyces cerevisiae.
In Saccharomyces cerevisiae, a type 1 protein phosphatase complex composed of the Glc7 catalytic subunit and the Reg1 regulatory subunit represses expression of many glucose-regulated genes. Here we show that the Reg1-interacting proteins Bmh1, Bmh2, Ssb1, and Ssb2 have roles in glucose repression. Deleting both BMH genes causes partially constitutive ADH2 expression without significantly increasing the level of Adr1 protein, the major activator of ADH2 expression. Adr1 and Bcy1, the regulatory subunit of cAMP-dependent protein kinase, are both required for this effect indicating that constitutive expression in Deltabmh1Deltabmh2 cells uses the same activation pathway that operates in Deltareg1 cells. Deletion of both BMH genes and REG1 causes a synergistic relief from repression, suggesting that Bmh proteins also act independently of Reg1 during glucose repression. A two-hybrid interaction with the Bmh proteins was mapped to amino acids 187-232, a region of Reg1 that is conserved in different classes of fungi. Deleting this region partially releases SUC2 from glucose repression. This indicates a role for the Reg1-Bmh interaction in glucose repression and also suggests a broad role for Bmh proteins in this process. An in vivo Reg1-Bmh interaction was confirmed by copurification of Bmh proteins with HA(3)-TAP-tagged Reg1. The nonconventional heat shock proteins Ssb1 and Ssb2 are also copurified with HA(3)-TAP-tagged Reg1. Deletion of both SSB genes modestly decreases repression of ADH2 expression in the presence of glucose, suggesting that Ssb proteins, perhaps through their interaction with Reg1, play a minor role in glucose repression.
Dombek
Kenneth M
KM
Department of Biochemistry, University of Washington, Seattle, Washington 98195-7350, USA. kmd@u.washington.edu
Kacherovsky
Nataly
N
Young
Elton T
ET
D048948:14-3-3 Proteins; D000595:Amino Acid Sequence; D015153:Blotting, Western; D002455:Cell Division; D000242:Cyclic AMP / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000502:physiology*; D017353:Gene Deletion; D005838:Genotype; D005947:Glucose / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000502:physiology*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010749:Phosphoprotein Phosphatases / Q000502:physiology*; D010957:Plasmids / Q000378:metabolism; D054645:Protein Phosphatase 1; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000502:physiology*; D029702:Schizosaccharomyces pombe Proteins / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques; D001616:beta-Galactosidase / Q000378:metabolism
D048948:14-3-3 Proteins; C075852:BMH1 protein, S cerevisiae; C097164:BMH2 protein, S cerevisiae; D004268:DNA-Binding Proteins; D018840:HSP70 Heat-Shock Proteins; C095192:SSB1 protein, S cerevisiae; C101143:SSB2 protein, S pombe; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D000242:Cyclic AMP; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; C070829:REG1 protein, S cerevisiae; D001616:beta-Galactosidase; D005947:Glucose