The DNA helicase ChlR1 is required for sister chromatid cohesion in mammalian cells

Abstract

It has recently been suggested that the Saccharomyces cerevisiae protein Chl1p plays a role in cohesion establishment. Here, we show that the human ATPdependent DNA helicase ChlR1 is required for sister chromatid cohesion in mammalian cells. Localization studies show that ChlR1 diffusely coats mitotic chromatin in prophase and then translocates from the chromatids to concentrate at the spindle poles during the transition to metaphase. Depletion of ChlR1 protein by RNA interference results in mitotic failure with replicated chromosomes failing to segregate after a pro-metaphase arrest. We show that depletion also results in abnormal sister chromatid cohesion, determined by increased separation of chromatid pairs at the centromere. Furthermore, biochemical studies show that ChlR1 is in complex with cohesin factors Scc1, Smc1 and Smc3. We conclude that human ChlR1 is required for sister chromatid cohesion and, hence, normal mitotic progression. These functions are important to maintain genetic fidelity.

Loss of ChlR1 Helicase in Mouse Causes Lethality Due to the Accumulation of Aneuploid Cells Generated by Cohesion Defects and Placental Malformation

“…A substantial fraction of cells (28% and 34% of the cells treated with RNAi constructs #5 and #7, respectively) also showed chromosome misalignment or missegregation (Supplementary Figure 3E). Since these phenotypes are ChlR1 is Required for Development and Chromosome Cohesion similar to the mouse phenotype and those published previously, 10 we concluded that the siRNA system was suitable for further studies.…”

“…In addition, depletion of either the cohesin molecule RAD21 or the ChlR proteins resulted in a loosened primary constriction or separated chromatids in HeLa cells. These studies verify the results of other ChlR1 RNAi studies 10 and are consistent with data obtained from our mutant mouse studies; as such, we have clearly eliminated the possibility that the effects of ChlR proteins on chromosome cohesion were artifacts encountered in siRNA experiments. Thus, we conclude that ChlR is required for centromeric cohesion in both mouse and human cells.…”

“…Since siRNA-mediated depletion of ChlR causes decreased centromeric cohesion in human cells, 10 we examined the morphology of the mouse embryo chromosomes. As shown in Figure 3E, sister chromatids from Ddx11 +/+ cells contained a shared region of pericentric heterochromatin representing areas of centromeric cohesion.…”

“…Since loosened primary constrictions indicate a loss of cohesion, [17][18][19] these data indicated the ChlR-depleted cells have a defect in sister-chromatid cohesion at centromeres as reported previously. 10 It is difficult to observe chromosome arm cohesion in mammalian cells because interphase cells do not shown a clear separation of sister signals by FISH, and arm cohesin is removed by the activity of Plk1, aurora B and possibly other mitotic kinases early in mitosis. However, since that dissociation of cohesin from mitotic chromosome arms requires the phosphorylation of cohesin component SA2 by polo-like kinase 1 (Plk1), [20][21][22] depletion of Plk1 prevents the removal of cohesin complexes from sister-chromatid arms.…”

“…A role for human ChlR proteins in the cell cycle was suggested by our previous data indicating that ChlR proteins are expressed ubiquitously in proliferative human tissues and cell lines, but not in quiescent cells; 9 and by recent studies using siRNA technology. 10 The siRNA studies revealed that human ChlR1 interacts with cohesion components and that this protein was required for sister-chromatid cohesion. DDX11 was also identified in a genetic study as a strong candidate gene for telomere length determination in humans.…”