Dbf4-dependent kinase (DDK)-mediated proteolysis of CENP-A prevents mislocalization of CENP-A in Saccharomyces cerevisiae

• Autores: Eisenstatt, Jessica R.; Boeckmann, Lars; Au, Wei Chun; Garcia, Valerie; Bursch, Levi; Ocampo, Josefina; Costanzo, Michael; Weinreich, Michael; Sclafani, Robert A.; Baryshnikova, Anastasia; Myers, Chad L.; Boone, Charles; Clark, David J.; Baker, Richard; Basrai, Munira A.
• Año de publicación: 2020
• Idioma: inglés
• Tipo de recurso: artículo
• Materia: CENTROMERE; Cse4; CENP-A; DDK; Psh1; Cdc7
• Institución: Consejo Nacional de Investigaciones Científicas y Técnicas

The evolutionarily conserved centromeric histone H3 variant (Cse4 in budding yeast, CENP-A in humans) is essential for faithful chromosome segregation. Mislocalization of CENP-A to non-centromeric chromatin contributes to chromosomal instability (CIN) in yeast, fly, and human cells and CENP-A is highly expressed and mislocalized in cancers. Defining mechanisms that prevent mislocalization of CENP-A is an area of active investigation. Ubiquitin-mediated proteolysis of overexpressed Cse4 (GALCSE4)byE3 ubiquitin ligases such as Psh1 prevents mislocalization of Cse4, and psh1D strains display synthetic dosage lethality (SDL) with GALCSE4. We previously performed a genome-wide screen and identified five alleles of CDC7 and DBF4 that encode the Dbf4-dependent kinase (DDK) complex, which regulates DNA replication initiation, among the top twelve hits that displayed SDL with GALCSE4. We determined that cdc7-7 strains exhibit defects in ubiquitin-mediated proteolysis of Cse4 and show mislocalization of Cse4. Mutation of MCM5 (mcm5-bob1) bypasses the requirement of Cdc7 for replication initiation and rescues replication defects in a cdc7-7 strain. We determined that mcm5-bob1 does not rescue the SDL and defects in proteolysis of GALCSE4 in a cdc7-7 strain, suggesting a DNA replication-independent role for Cdc7 in Cse4 proteolysis. The SDL phenotype, defects in ubiquitin-mediated proteolysis, and the mislocalization pattern of Cse4 in a cdc7-7 psh1D strain were similar to that of cdc7-7 and psh1D strains, suggesting that Cdc7 regulates Cse4 in a pathway that overlaps with Psh1. Our results define a DNA replication initiation-independent role of DDK as a regulator of Psh1-mediated proteolysis of Cse4 to prevent mislocalization of Cse4.
Fil: Eisenstatt, Jessica R.. National Institutes of Health; Estados Unidos
Fil: Boeckmann, Lars. National Institutes of Health; Estados Unidos
Fil: Au, Wei Chun. National Institutes of Health; Estados Unidos
Fil: Garcia, Valerie. National Institutes of Health; Estados Unidos
Fil: Bursch, Levi. National Institutes of Health; Estados Unidos
Fil: Ocampo, Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentina. National Instituto of Child Health & Human Development; Estados Unidos
Fil: Costanzo, Michael. National Institutes of Health; Estados Unidos. University of Toronto; Canadá
Fil: Weinreich, Michael. Van Andel Research Institute; Estados Unidos
Fil: Sclafani, Robert A.. University of Colorado; Estados Unidos
Fil: Baryshnikova, Anastasia. University of Princeton; Estados Unidos
Fil: Myers, Chad L.. University of Minnesota; Estados Unidos
Fil: Boone, Charles. University of Toronto; Canadá. National Institutes of Health; Estados Unidos
Fil: Clark, David J.. National Institutes of Health; Estados Unidos
Fil: Baker, Richard. University of Massachusetts; Estados Unidos
Fil: Basrai, Munira A.. National Institutes of Health; Estados Unidos