The DNA damage-inducible UbL-UbA protein Ddi1 participates in Mec1-mediated degradation of Ho endonuclease.
Ho endonuclease initiates a mating type switch by making a double-strand break at the mating type locus, MAT. Ho is marked by phosphorylation for rapid destruction by functions of the DNA damage response, MEC1, RAD9, and CHK1. Phosphorylated Ho is recruited for ubiquitylation via the SCF ubiquitin ligase complex by the F-box protein, Ufo1. Here we identify a further DNA damage-inducible protein, the UbL-UbA protein Ddi1, specifically required for Ho degradation. Ho interacts only with Ddi1; it does not interact with the other UbL-UbA proteins, Rad23 or Dsk2. Ho must be ubiquitylated to interact with Ddi1, and there is no interaction when Ho is produced in mec1 or Deltaufo1 mutants that do not support its degradation. Ddi1 binds the proteasome via its N-terminal ubiquitin-like domain (UbL) and interacts with ubiquitylated Ho via its ubiquitin-associated domain (UbA); both domains of Ddi1 are required for association of ubiquitylated Ho with the proteasome. Despite being a nuclear protein, Ho is exported to the cytoplasm for degradation. In the absence of Ddi1, ubiquitylated Ho is stabilized and accumulates in the cytoplasm. These results establish a role for Ddi1 in the degradation of a natural ubiquitylated substrate. The specific interaction between Ho and Ddi1 identifies an additional function associated with DNA damage involved in its degradation.
Kaplun
Ludmila
L
Department of Life Sciences, Ben Gurion University of Negev, P.O. Box 653, Beersheba 84105, Israel.
Tzirkin
Regina
R
Bakhrat
Anya
A
Shabek
Nitzan
N
Ivantsiv
Yelena
Y
Raveh
Dina
D
D003593:Cytoplasm / Q000378:metabolism; D004249:DNA Damage; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins; D009154:Mutation; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D025801:Ubiquitin / Q000378:metabolism
C108125:DDI1 protein, S cerevisiae; D005656:Fungal Proteins; D047908:Intracellular Signaling Peptides and Proteins; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; C088937:MEC1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; C530845:HO protein, S cerevisiae; C037725:SCEI protein, S cerevisiae; D015252:Deoxyribonucleases, Type II Site-Specific; D046988:Proteasome Endopeptidase Complex