Alternative end-joining catalyzes robust IgH locus deletions and translocations in the combined absence of ligase 4 and Ku70.
Class switch recombination (CSR) in B lymphocytes is initiated by introduction of multiple DNA double-strand breaks (DSBs) into switch (S) regions that flank immunoglobulin heavy chain (IgH) constant region exons. CSR is completed by joining a DSB in the donor S mu to a DSB in a downstream acceptor S region (e.g., S gamma1) by end-joining. In normal cells, many CSR junctions are mediated by classical nonhomologous end-joining (C-NHEJ), which employs the Ku70/80 complex for DSB recognition and XRCC4/DNA ligase 4 for ligation. Alternative end-joining (A-EJ) mediates CSR, at reduced levels, in the absence of C-NHEJ, even in combined absence of Ku70 and ligase 4, demonstrating an A-EJ pathway totally distinct from C-NHEJ. Multiple DSBs are introduced into S mu during CSR, with some being rejoined or joined to each other to generate internal switch deletions (ISDs). In addition, S-region DSBs can be joined to other chromosomes to generate translocations, the level of which is increased by absence of a single C-NHEJ component (e.g., XRCC4). We asked whether ISD and S-region translocations occur in the complete absence of C-NHEJ (e.g., in Ku70/ligase 4 double-deficient B cells). We found, unexpectedly, that B-cell activation for CSR generates substantial ISD in both S mu and S gamma1 and that ISD in both is greatly increased by the absence of C-NHEJ. IgH chromosomal translocations to the c-myc oncogene also are augmented in the combined absence of Ku70 and ligase 4. We discuss the implications of these findings for A-EJ in normal and abnormal DSB repair.
Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
Cristian
C
Boboila
Mila
M
Jankovic
Catherine T
CT
Yan
Jing H
JH
Wang
Duane R
DR
Wesemann
Tingting
T
Zhang
Alex
A
Fazeli
Lauren
L
Feldman
Andre
A
Nussenzweig
Michel
M
Nussenzweig
Frederick W
FW
Alt
D000818:Animals; D034961:Antigens, Nuclear / Q000235:genetics; D001402:B-Lymphocytes / Q000276:immunology; D001483:Base Sequence; D015139:Blotting, Southern; D053903:DNA Breaks, Double-Stranded; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004260:DNA Repair / Q000276:immunology*; D004268:DNA-Binding Proteins / Q000235:genetics; D016259:Genes, myc / Q000235:genetics; D017578:Immunoglobulin Class Switching / Q000235:genetics* / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D007134:Immunoglobulin Switch Region / Q000235:genetics*; D017404:In Situ Hybridization, Fluorescence; D000072200:Ku Autoantigen; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D014178:Translocation, Genetic / Q000235:genetics* / Q000276:immunology
D034961:Antigens, Nuclear; D017931:DNA Primers; D004268:DNA-Binding Proteins; D007143:Immunoglobulin Heavy Chains; C000606510:Xrcc6 protein, mouse; D000072200:Ku Autoantigen; D011088:DNA Ligases; D000072481:DNA Ligase ATP