Showing papers by "Marian Carlson published in 1989"

Mutational analysis of the Saccharomyces cerevisiae SNF1 protein kinase and evidence for functional interaction with the SNF4 protein.

Abstract: The SNF1 gene of Saccharomyces cerevisiae encodes a protein-serine/threonine kinase that is required for derepression of gene expression in response to glucose limitation. We present evidence that the protein kinase activity is essential for SNF1 function: substitution of Arg for Lys in the putative ATP-binding site results in a mutant phenotype. A polyhistidine tract near the N terminus was found to be dispensable. Deletion of the large region C terminal to the kinase domain only partially impaired SNF1 function, causing expression of invertase to be somewhat reduced but still glucose repressible. The function of the SNF4 gene, another component of the regulatory system, was required for maximal in vitro activity of the SNF1 protein kinase. Increased SNF1 gene dosage partially alleviated the requirement for SNF4. C-terminal deletions of SNF1 also reduced dependence on SNF4. Our findings suggest that SNF4 acts as a positive effector of the kinase but does not serve a regulatory function in signaling glucose availability.

Molecular analysis of the SNF4 gene of Saccharomyces cerevisiae: evidence for physical association of the SNF4 protein with the SNF1 protein kinase.

Abstract: The SNF4 gene is required for expression of glucose-repressible genes in response to glucose deprivation in Saccharomyces cerevisiae. Previous evidence suggested that SNF4 is functionally related to SNF1, another essential gene in this global regulatory system that encodes a protein kinase. Increased SNF1 gene dosage partially compensates for a mutation in SNF4, and the SNF4 function is required for maximal SNF1 protein kinase activity in vitro. We have cloned SNF4 and identified its 1.2-kilobase RNA, which is not regulated by glucose repression. A 36-kilodalton SNF4 protein is predicted from the nucleotide sequence. Disruption of the chromosomal SNF4 locus revealed that the requirement for SNF4 function is less stringent at low temperature (23 degrees C). A bifunctional SNF4-lacZ gene fusion that includes almost the entire SNF4 coding sequence was constructed. The fusion protein was shown by immunofluorescence microscopy to be distributed throughout the cell, with partial localization to the nucleus. The SNF4-beta-galactosidase protein coimmunoprecipitated with the SNF1 protein kinase, thus providing evidence for the physical association of the two proteins.