Aurora A-dependent CENP-A phosphorylation at inner centromeres protects bioriented chromosomes against cohesion fatigue

Abstract

Sustained spindle tension applied to sister centromeres during mitosis eventually leads to uncoordinated loss of sister chromatid cohesion, a phenomenon known as “cohesion fatigue.” We report that Aurora A-dependent phosphorylation of serine 7 of the centromere histone variant CENP-A (p-CENP-AS7) protects bioriented chromosomes against cohesion fatigue. Expression of a non-phosphorylatable version of CENP-A (CENP-AS7A) weakens sister chromatid cohesion only when sister centromeres are under tension, providing the first evidence of a regulated mechanism involved in protection against passive cohesion loss. Consistent with this observation, p-CENP-AS7 is detected at the inner centromere where it forms a discrete domain. The depletion or inhibition of Aurora A phenocopies the expression of CENP-AS7A and we show that Aurora A is recruited to centromeres in a Bub1-dependent manner. We propose that Aurora A-dependent phosphorylation of CENP-A at the inner centromere protects chromosomes against tension-induced cohesion fatigue until the last kinetochore is attached to spindle microtubules.

Phosphorylation of CENP-A on serine 7 does not control centromere function

“…What role, if any, CENP-A S7ph may provide remains an open question; however, our data strongly indicate that this post-translational modification is not essential nor required for centromere function, in contrast to previous proposals 20–22,24 . While our data do not exclude a function for S7 CENP-A phosphorylation in some cell events such as maintenance of proper cohesion via Shugoshin 24 , it argues that such function cannot be essential for long-term cell cycling and chromosome segregation. Moreover, CENP-A S7ph has never been found in mass spectrometry data designed to identify CENP-A PTMs across the cell cycle, including during mitosis 19 , suggesting that this PTM might be at low abundancy, as recently suggested 24 .…”

“…1) also highlight how the level of CENP-A is critical for centromere function (as in the case of the CENP-A S7E variant): too little or too much can have deleterious effects on cell viability, in agreement with previous results 2,29,30,39 . This might explain previously contradictory reports on the importance of CENP-A S7ph that were obtained with technologies that produced only partial CENP-A downregulation (achieved by RNAi) and/or transient rescue (known to lead to a range of expression levels including overexpression 40 ) with CENP-A mutants 20–22,24 , as observed for other PTMs of CENP-A 13–15 .…”

“…Recognizing that cells that have a more unstable karyotype might be more sensitive to centromere manipulation, we next tested the influence of CENP-A S7ph in a non-diploid cell line such as HeLa, a line used in previous reports to assess the importance of such PTM 22,24 . We tagged all endogenous HeLa CENP-A alleles with an AID tag to permit its rapid, inducible degradation (Supplementary Fig.…”

“…It is enriched in arginines that do not appear to be frequently modified and lacks most of the well-characterized lysines of histone H3, known to be hotspots of conserved PTMs that regulate histone function 17 . PTMs on CENP-A’s amino-terminal tail such as the α-N of glycine 1 and phosphorylation of Serine 7, 17, and 19 (hereafter named S7 18,19 , S16 and S18 due to first methionine digestion 18,19 ) have also been detected, all of them to some extent proposed to be important for CENP-A’s functionality 19–24 .…”

“…Indeed, quite surprisingly, they found that ectopic expression of a CENP-A S7A variant led to CENP-C loss and, consequently, to mitotic errors and cell lethality. Recently, it was found that preventing CENP-A S7ph led to sister chromatid cohesion defects but no evidence of defective kinetochore assembly was reported 24 .…”