DNA-dependent protein kinase inhibits AID-induced antibody gene conversion.
Affinity maturation and class switching of antibodies requires activation-induced cytidine deaminase (AID)-dependent hypermutation of Ig V(D)J rearrangements and Ig S regions, respectively, in activated B cells. AID deaminates deoxycytidine bases in Ig genes, converting them into deoxyuridines. In V(D)J regions, subsequent excision of the deaminated bases by uracil-DNA glycosylase, or by mismatch repair, leads to further point mutation or gene conversion, depending on the species. In Ig S regions, nicking at the abasic sites produced by AID and uracil-DNA glycosylases results in staggered double-strand breaks, whose repair by nonhomologous end joining mediates Ig class switching. We have tested whether nonhomologous end joining also plays a role in V(D)J hypermutation using chicken DT40 cells deficient for Ku70 or the DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Inactivation of the Ku70 or DNA-PKcs genes in DT40 cells elevated the rate of AID-induced gene conversion as much as 5-fold. Furthermore, DNA-PKcs-deficiency appeared to reduce point mutation. The data provide strong evidence that double-strand DNA ends capable of recruiting the DNA-dependent protein kinase complex are important intermediates in Ig V gene conversion.
Cook
Adam J L
AJ
Centenary Institute and University of Sydney Faculty of Medicine, Sydney, New South Wales, Australia.
Raftery
Joanna M
JM
Lau
K K Edwin
KK
Jessup
Andrew
A
Harris
Reuben S
RS
Takeda
Shunichi
S
Jolly
Christopher J
CJ
D000818:Animals; D000906:Antibodies / Q000235:genetics*; D002460:Cell Line; D002645:Chickens; D002999:Clone Cells; D003564:Cytidine Deaminase / Q000378:metabolism*; D051747:DNA-Activated Protein Kinase / Q000378:metabolism*; D005434:Flow Cytometry; D005785:Gene Conversion; D007134:Immunoglobulin Switch Region; D009154:Mutation; D051981:Uracil-DNA Glycosidase / Q000378:metabolism; D000098830:AICDA (Activation-Induced Cytidine Deaminase)
D000906:Antibodies; D051747:DNA-Activated Protein Kinase; D051981:Uracil-DNA Glycosidase; D000098830:AICDA (Activation-Induced Cytidine Deaminase); D003564:Cytidine Deaminase