V(D)J recombination intermediates and non-standard products in XRCC4-deficient cells.
V(D)J recombination assembles immunoglobulin (Ig) and T cell receptor (TCR) gene segments during lymphocyte development. Recombination is initiated by the RAG-1 and RAG-2 proteins, which introduce double-stranded DNA breaks (DSB) adjacent to the Ig and TCR gene segments. The broken ends are joined by the DSB repair machinery, which includes the XRCC4 protein. While XRCC4 is essential for both DSB repair and V(D)J recombination, the functions of this protein remain enigmatic. Because the rare V(D)J recombination products isolated from XRCC4-deficient cells generally show evidence of excessive nucleotide loss, it was hypothesized that XRCC4 may function to protect broken DNA ends. Here we report the first examination of V(D)J recombination intermediates in XRCC4-deficient cells. We found that both types of intermediates, signal ends and coding ends, are abundant in the absence of XRCC4. Furthermore, the signal ends are full length. We also showed that alternative V(D)J recombination products, hybrid joints, form with normal efficiency and without excessive deletion in XRCC4-deficient cells. These data indicate that impaired formation of V(D)J recombination products in XRCC4-deficient cells does not result from excessive degradation of recombination intermediates. Potential roles of XRCC4 in the joining reaction are discussed.
The Department of Microbiology and Immunology, Baylor College of Medicine and The Howard Hughes Medical Institute, Baylor College of Medicine, Houston, TX 77030, USA.
J O
JO
Han
L A
LA
Erskine
M M
MM
Purugganan
T D
TD
Stamato
D B
DB
Roth
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D006224:Cricetinae; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000378:metabolism; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015321:Gene Rearrangement; D015345:Oligonucleotide Probes / Q000235:genetics; D011995:Recombination, Genetic; D014162:Transfection
D004268:DNA-Binding Proteins; D015345:Oligonucleotide Probes; C101467:XRCC4 protein, human; D011088:DNA Ligases; D000072481:DNA Ligase ATP