Abundance of the Fanconi anaemia core complex is regulated by the RuvBL1 and RuvBL2 AAA+ ATPases

Abstract

Fanconi anaemia (FA) is a genome instability disease caused by defects in the FA DNA repair pathway that senses and repairs damage caused by DNA interstrand crosslinks. At least 8 of the 16 genes found mutated in FA encode proteins that assemble into the FA core complex, a multisubunit monoubiquitin E3 ligase. Here, we show that the RuvBL1 and RuvBL2 AAA+ ATPases co-purify with FA core complex isolated under stringent but native conditions from a vertebrate cell line. Depletion of the RuvBL1-RuvBL2 complex in human cells causes hallmark features of FA including DNA crosslinker sensitivity, chromosomal instability and defective FA pathway activation. Genetic knockout of RuvBL1 in a murine model is embryonic lethal while conditional inactivation in the haematopoietic stem cell pool confers profound aplastic anaemia. Together these findings reveal a function for RuvBL1-RuvBL2 in DNA repair through a physical and functional association with the FA core complex. Surprisingly, depletion of RuvBL1-RuvBL2 leads to co-depletion of the FA core complex in human cells. This suggests that a potential mechanism for the role of RuvBL1-RuvBL2 in maintaining genome integrity is through controlling the cellular abundance of FA core complex.

A Novel Interaction of Ecdysoneless (ECD) Protein with R2TP Complex Component RUVBL1 Is Required for the Functional Role of ECD in Cell Cycle Progression

“…The delineation of sequences in ECD and RUVBL1 that mediate their interaction should help directly test whether selective abrogation of this interaction is functionally critical in cell cycle progression, as well as to assess the potential role of ECD in other roles of RUVBL1 within the R2TP complex. Interestingly, mouse ECD and RUVBL1 knockouts are phenotypically similar, since both are embryonic lethal at the blastocyst stage (6,19). RUVBL1 is essential for cellular proliferation as seen in knockout cells or upon knockdown of RUVBL1 expression (18).…”

“…Recent studies have shown that, aside from the R2TP complex, RUVBL1/2 are also parts of other functionally relevant complexes, such as chromatin remodeling complexes TIP60, SWR/SRCAP, and INO80 and the Fanconi anemia core complex that controls DNA interstrand cross-link repair and function, and regulate telomerase biogenesis and mitosis (19,(43)(44)(45). Given the PIH1D1-independent interaction of ECD with RUVBL1, the potential roles of ECD via these alternative complexes will be of great future interest.…”

“…Recent studies have shown that RUVBL1 (Pontin) plays an important role in cell cycle regulation (17,18). Germ line deletion of Ruvbl1 was shown to be early embryonic lethal (18,19). Depletion of RUVBL1 in AML1-ETO fusion oncogene-expressing leukemic cells was shown to cause cell cycle arrest (17) and Cre-mediated deletion of Ruvbl1 in Ruvbl1 fl/fl cells also led to G 1 /S cell cycle arrest (18).…”

Reticulocytes Are an Enriched Source of the RUVBL1 Protein

“…RUVBL1 is essential for adult hematopoiesis in the mouse [5]. In U2OS cells (osteosarcoma cell line), depletion of RUVBL1 together with RUVBL2 leads to depletion of the Fanconi anemia core complex, which in turn leads to chromosomal instability [13]. Salzer et al [1] postulated that RUVBL1 may be part of a chaperone complex which could have a vital role in protecting mammalian RBCs under various stress conditions by associating with cytoskeletal or membrane components.…”

Epigenetic Regulation by ATP-Dependent Chromatin-Remodeling Enzymes

“…The helicases RUVBL1 and RUVBL2 (RUVBLs) exist in INO80, TIP60, and SRCAP yet their remodeling-related functions are largely unknown, due in part to the fact that the RUVBLs promiscuously participate in non-remodeling activities through the Fanconi Anemia complex [44] and in snoRNP assembly [45]. Assembly of INO80 and TIP60 has been shown to be dependent on the RUVBLs [46], [47].…”