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, MECI, RAD9, and CHK1. Phosphorylated Ho is recruited for ubiquitylation via the SCF ubiquitin ligase complex by the F-box protein, MI. Here we identify a further DNA damage-inducible protein, the UbL-UbA protein Ddil, specifically required for Ho degradation. Ho interacts only with Ddil; it does not interact with the other UbL-UbA proteins, Rad23 or Dsk2. Ho must be ubiquitylated to interact with Ddil, and there is no interaction when Ho is produced in mec1 or Delta ufo1 mutants that do not support its degradation. Ddil binds the proteasome via its N-terminal ubiquitinlike domain (UbL) and interacts with ubiquitylated Ho via its ubiquitin-associated domain (UbA); both domains of Ddil 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 Ddil, ubiquitylated Ho is stabilized and accumulates in the cytoplasm. These results establish a role for Ddil in the degradation of a natural ubiquitylated substrate. The specific interaction between Ho and Ddil identifies an additional function associated with DNA damage involved in its degradation.