BLAP75, an essential component of Bloom's syndrome protein complexes that maintain genome integrity

Abstract

Bloom's syndrome (BS) is a rare human genetic disorder characterized by dwarfism, immunodeficiency, genomic instability and cancer predisposition. We have previously purified three complexes containing BLM, the helicase mutated in this disease. Here we demonstrate that BLAP75, a novel protein containing a putative OB-fold nucleic acid binding domain, is an integral component of BLM complexes, and is essential for their stability in vivo. Consistent with a role in BLM-mediated processes, BLAP75 colocalizes with BLM in subnuclear foci in response to DNA damage, and its depletion impairs the recruitment of BLM to these foci. Depletion of BLAP75 by siRNA also results in deficient phosphorylation of BLM during mitosis, as well as defective cell proliferation. Moreover, cells depleted of BLAP75 display an increased level of sisterchromatid exchange, similar to cells depleted of BLM by siRNA. Thus, BLAP75 is an essential component of the BLM-associated cellular machinery that maintains genome integrity.

Defining the molecular interface that connects the Fanconi anemia protein FANCM to the Bloom syndrome dissolvasome

“…A theme that emerges from this work and previous studies is that depletion of proteins in either the BLM dissolvasome or the FA core complex, or disruption of protein interfaces that stabilize each complex or link the two, lead to similar increases in SCE levels ( Fig. 4 and (7,11,12,19,20). Taken together, these data suggest that the RMI/FANCM interface is critical in bridging the BLM dissolvasome and the FA core complexes and that proper association of these two complexes is vital in SCE suppression.…”

“…BS cells display elevated levels of SCEs, which is consistent with the proposed role for the BLM helicase in the nonrecombinogenic repair of doubleHolliday junction (dHJ) DNA structures that can arise during repair. To catalyze this activity, BLM functions within a complex called the "dissolvasome" that also includes topoisomerase IIIα (Top3α) and the RecQ-mediated genome instability, or RMI, subcomplex (RMI1/RMI2 heterodimer) (7)(8)(9)(10)(11)(12)(13). The dissolvasome is thought to drive HJ branch migration (catalyzed by BLM) and DNA cleavage of the resulting hemi-catenated DNA (catalyzed by Top3α) in a reaction called "dissolution" (14).…”

Shu Proteins Promote the Formation of Homologous Recombination Intermediates That Are Processed by Sgs1-Rmi1-Top3

“…The yeast and human Rmi1 proteins have been proposed to be integral components of multienzyme complexes containing Sgs1 and Top3, and BLM and hTOPOIII␣, respectively (Johnson et al, 2000;Wu et al, 2000;Chang et al, 2005;Mullen et al, 2005;Yin et al, 2005). We examined, therefore, if mutation of SHU1 also suppresses rmi1 phenotypes.…”

“…This is exemplified by the cancer-predisposition disorder, Bloom's syndrome, which is caused by mutation in the human BLM gene (reviewed in German, 1993). Because the BLM protein, in conjunction with its associated proteins, hTOPOIII␣ and hRMI1 (Johnson et al, 2000;Wu et al, 2000;Yin et al, 2005), can catalyze dissolution of HRR intermediates in vitro Plank et al, 2006;Raynard et al, 2006;Wu et al, 2006), it is likely that unprocessed and/or aberrantly processed HRR intermediates at least partly contribute to the cellular defects in Bloom's syndrome. Indeed, Bloom's syndrome cells classically demonstrate elevated levels of sister chromatid exchanges, mitotic recombination, and genome instability.…”

“…RMI1 encodes a protein that associates with Sgs1 and Top3 in vivo, and rmi1 mutants phenotypically resemble top3 mutants (Chang et al, 2005;Mullen et al, 2005). This physical interaction is also evolutionarily conserved, because the human Rmi1 homologue, hRMI1, is an integral component of the BLM-hTOPOIII␣ complex in human cells (Yin et al, 2005). In this study, we have demonstrated that mutation of SHU1 suppresses poor growth caused by overexpression of TOP3 Y356F or deletion of RMI1, indicating that the Shu complex actively contributes to the cellular defects seen in cells lacking Sgs1, Top3, or Rmi1.…”