Calpain-1 Cleaves Rad21 To Promote Sister Chromatid Separation

Abstract

Defining the mechanisms of chromosomal cohesion and dissolution of the cohesin complex from chromatids is important for understanding the chromosomal missegregation seen in many tumor cells. Here we report the identification of a novel cohesin-resolving protease and describe its role in chromosomal segregation. Sister chromatids are held together by cohesin, a multiprotein ring-like complex comprised of Rad21, Smc1, Smc3, and SA2 (or SA1). Cohesin is known to be removed from vertebrate chromosomes by two distinct mechanisms, namely, the prophase and anaphase pathways. First, PLK1-mediated phosphorylation of SA2 in prophase leads to release of cohesin from chromosome arms, leaving behind centromeric cohesins that continue to hold the sisters together. Then, at the onset of anaphase, activated separase cleaves the centromeric cohesin Rad21, thereby opening the cohesin ring and allowing the sister chromatids to separate. We report here that the calcium-dependent cysteine endopeptidase calpain-1 is a Rad21 peptidase and normally localizes to the interphase nuclei and chromatin. Calpain-1 cleaves Rad21 at L192, in a calcium-dependent manner. We further show that Rad21 cleavage by calpain-1 promotes separation of chromosome arms, which coincides with a calcium-induced partial loss of cohesin at several chromosomal loci. Engineered cleavage of Rad21 at the calpain-cleavable site without activation of calpain-1 can lead to a loss of sister chromatid cohesion. Collectively, our work reveals a novel function of calpain-1 and describes an additional pathway for sister chromatid separation in humans.DNA replication in S phase generates two identical molecules of chromosomal DNA, known as sister chromatids, which must equally segregate to the two daughter cells in the subsequent mitosis. Therefore, it is important that the sister chromatids are held together from the S phase until proper kinetochore assembly is complete. Such cohesion of the sisters is mediated by a multiprotein ring-like complex called cohesin, which is conserved from Saccharomyces cerevisiae to humans. Cohesin is comprised of four core proteins, namely, Rad21 (Scc1/Mcd1), Smc1, Smc3, and either SA2/Stag2 or SA1/Stag1 (13,22,25,39; reviewed in reference 28). Cohesin undergoes a series of physical and chemical changes during the course of the cell cycle in order to orchestrate the dynamic cohesion and separation between the sister chromatids that are essential for high-fidelity transmission of the genetic material (32). The cohesin cycle includes loading cohesins onto the chromatin of the unpaired chromosome, forming cohesion between the sister chromatids in S phase, regulated freeing of chromosome arms in early mitosis, separation of sister chromatids to allow segregation to opposite poles, and recycling of cohesin molecules for subsequent usage.Cohesin is loaded onto the chromatin in G 1 phase in yeast (25) and in late telophase in vertebrates (21, 39). Loading of cohesin onto chromatin depends on two proteins, Scc2 and Scc4 (8,46). Once it is loaded onto...

Molecular Basis of Calpain Cleavage and Inactivation of the Sodium-Calcium Exchanger 1 in Heart Failure

“…The empty vectors pcDNA3.1 and EGFP-N1 were obtained from Invitrogen and Clontech, respectively. Tobacco etch virus (TEV) protease in pCS2MT was kindly provided by Prof. Pati (17).…”

Augmented generation of protein fragments during wakefulness as the molecular cause of sleep: a hypothesis

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Calcium mobilization is both required and sufficient for initiating chromatin decondensation during activation of peripheral T-cells

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