Bloom DNA helicase facilitates homologous recombination between diverged homologous sequences.
Bloom syndrome caused by inactivation of the Bloom DNA helicase (Blm) is characterized by increases in the level of sister chromatid exchange, homologous recombination (HR) associated with cross-over. It is therefore believed that Blm works as an anti-recombinase. Meanwhile, in Drosophila, DmBlm is required specifically to promote the synthesis-dependent strand anneal (SDSA), a type of HR not associating with cross-over. However, conservation of Blm function in SDSA through higher eukaryotes has been a matter of debate. Here, we demonstrate the function of Blm in SDSA type HR in chicken DT40 B lymphocyte line, where Ig gene conversion diversifies the immunoglobulin V gene through intragenic HR between diverged homologous segments. This reaction is initiated by the activation-induced cytidine deaminase enzyme-mediated uracil formation at the V gene, which in turn converts into abasic site, presumably leading to a single strand gap. Ig gene conversion frequency was drastically reduced in BLM(-/-) cells. In addition, BLM(-/-) cells used limited donor segments harboring higher identity compared with other segments in Ig gene conversion event, suggesting that Blm can promote HR between diverged sequences. To further understand the role of Blm in HR between diverged homologous sequences, we measured the frequency of gene targeting induced by an I-SceI-endonuclease-mediated double-strand break. BLM(-/-) cells showed a severer defect in the gene targeting frequency as the number of heterologous sequences increased at the double-strand break site. Conversely, the overexpression of Blm, even an ATPase-defective mutant, strongly stimulated gene targeting. In summary, Blm promotes HR between diverged sequences through a novel ATPase-independent mechanism.
Kikuchi
Koji
K
Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshidakonoe, Kyoto 606-8501, Japan.
Abdel-Aziz
H Ismail
HI
Taniguchi
Yoshihito
Y
Yamazoe
Mitsuyoshi
M
Takeda
Shunichi
S
Hirota
Kouji
K
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D002453:Cell Cycle; D002460:Cell Line; D002645:Chickens; D004247:DNA / Q000235:genetics / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded / Q000528:radiation effects; D004260:DNA Repair; D045643:DNA Repair Enzymes / Q000235:genetics / Q000378:metabolism; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000378:metabolism; D005720:Gamma Rays; D006801:Humans; D007136:Immunoglobulins / Q000235:genetics; D009154:Mutation; D053484:RecQ Helicases / Q000235:genetics / Q000378:metabolism*; D011995:Recombination, Genetic / Q000235:genetics*; D012854:Sister Chromatid Exchange / Q000235:genetics
D007136:Immunoglobulins; D004247:DNA; C114331:EXO1 protein, human; D005090:Exodeoxyribonucleases; D000251:Adenosine Triphosphatases; C097151:Bloom syndrome protein; D053484:RecQ Helicases; D045643:DNA Repair Enzymes