The Prolylhydroxylase PHD2 Modulates Centromere Function through the Hydroxylation of CENP-N

Abstract

Successful segregation of chromosomes during mitosis requires that each sister chromatid is captured by microtubules emanating from opposite spindle poles. A multiprotein complex called the kinetochore provides an attachment site on chromosomes for microtubules. We have found that the prolylhydroxylase PHD2 is a critical regulator of the assembly of the kinetochore. PHD2 hydroxylates the kinetochore component CENP-N on P311 and is essential for CENP-N localization to kinetochores. Either depletion of PHD2, or expression of a hydroxylation-deficient mutant, results in loss of the histone H3 variant CENP-A from centromeres. Loss of CENP-N from chromatin bound protein complexes is not due to decreased protein stability but is a consequence of lowered affinity of CENP-N for binding CENP-L. Loss of hydroxylation also results in increased targeting of CENP-L to chromatin. Hydroxylation by PHD2 thus plays an important role in controlling the stoichiometry of mitotic kinetochore components.