Bloom's syndrome protein response to ultraviolet-C radiation and hydroxyurea-mediated DNA synthesis inhibition

Abstract

Bloom's syndrome (BS) arises through mutations in both copies of the BLM gene that encodes a RecQ 3'-5' DNA helicase. BS patients are predisposed to developing all the cancers that aect the general population, and BS cells exhibit marked genetic instability. We showed recently that BLM protein contributes to the cellular response to ionizing radiation by acting as downstream ATM kinase eector. We now show that following UVC treatment, BLM-de®cient cells exhibit a reduction in the number of replicative cells, a partial escape from the G2/ M cell cycle checkpoint, and have an altered p21 response. Surprisingly, we found that hydroxyureatreated BLM-de®cient cells exhibit an intact S phase arrest, proper recovery from the S phase arrest, and intact p53 and p21 responses. We also show that the level of BLM falls sharply in response to UVC radiation. This UVC-induced reduction in BLM does not require a functional ATM gene and does not result from a subcellular compartment change. Finally, we demonstrate that exposure to UVC and hydroxyurea treatment both induce BLM phosphorylation via an ATM-independent pathway. These results are discussed in the light of their potential physiological signi®cance with regard to the role of BLM in the cellular pathways activated by UVC radiation or HU-mediated inhibition of DNA synthesis.

Multiple Genetic Pathways Involving the Caenorhabditis elegans Bloom's Syndrome Genes him-6, rad-51, and top-3 Are Needed To Maintain Genome Stability in the Germ Line

“…We observed only partial checkpoint defects in him-6 C. elegans worms. Similarly, irradiation-induced apoptosis and G 2 /M cell cycle arrest is only partially compromised in human BS Ϫ/Ϫ cells (1,59). It was recently shown that HIM-6 physically interacts with the C. elegans ATR homolog, whose inactivation by RNAi leads to defective checkpoint processes in response to ionizing radiation (8), as well as to HU (A. Gartner, unpublished observation).…”

“…sgs1-deficient budding yeast cells are partially defective in arresting S-phase progression in response to HU (15,66). Human BS Ϫ/Ϫ cells, on the other hand, respond normally to HU (1). S. pombe cells lacking the RecQ-like helicase Rqh1p also react normally to HU but are defective in the recovery from S-phase arrest when exposed to HU (53).…”

Cleavage of BLM and Sensitivity of Bloom’s Syndrome Cells to Hydroxurea

“…We performed the same experiments using wild type Priess and IPM1 BS lymphoblastoid cells, and we did not detect BLM proteolytic products (data not shown). Moreover, the strong decrease in expression of the full-length BLM protein in response to UVC radiation, which we previously described for D1 cells, 14 was also observed in K562 and Ramos cells (this study), and in Priess and IPM1 cells (data not shown). Thus, the UVC-induced decrease in BLM expression apparently does not correlate with the appearance of BLM proteolytic products.…”

“…1B), as expected. 14 Furthermore, 6 and 16 h post-UVC, the full-length BLM protein was totally degraded to a single band of about 110 kDa (∆BLM) in Ramos cells (Fig. 1B, upper middle panel), whereas no BLM cleavage was observed in UVCtreated D1 or K562 cells (Fig.…”

“…13 These data, together with our results showing that BLM protein is modified by phosphorylation through an ATM-dependent pathway in response to ionizing radiation and through an ATM-independent pathway in response to UVC irradiation and HU treatment, prompted us to analyze BLM expression and BS cell sensitivity in response to apoptosis induction by UVC or HU. 7,14 …”