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Harold C. Simmons Arthritis Research Center and Departments of Internal Medicine and Microbiology, University of Texas Southwestern Medical Center, Dallas, TX 75235;
Department of Molecular Medicine, Institute for Biotechnology, University of Texas Health Science Center, San Antonio, TX 78245; and
Department of Microbiology and Immunology and
§
Howard Hughes Medical Institute, Baylor College of Medicine, Houston, TX 77030
Rare Ig and TCR coding joints can be isolated from mice that have a targeted deletion in the gene encoding the 86-kDa subunit of the Ku heterodimer, the regulatory subunit of the DNA-dependent protein kinase (DNA-PK). However in the coding joints isolated from Ku86-/- animals, there is an extreme paucity of N regions (the random nucleotides added during V(D)J recombination by the enzyme TdT). This finding is consistent with a decreased frequency of coding joints containing N regions isolated from C.B-17 SCID mice that express a truncated form of the catalytic subunit of the DNA-PK (DNA-PKCS). This finding suggests an unexpected role for DNA-PK in addition of N nucleotides to coding ends during V(D)J recombination. In this report, we establish that TdT forms a stable complex with DNA-PK. Furthermore, we show that DNA-PK modulates TdT activity in vitro by limiting both the length and composition of nucleotide additions.
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