Modulation of terminal deoxynucleotidyltransferase activity by the DNA-dependent protein kinase.
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.
Mickelsen
S
S
Harold C. Simmons Arthritis Research Center, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas 75235, USA.
Snyder
C
C
Trujillo
K
K
Bogue
M
M
Roth
D B
DB
Meek
K
K
D000818:Animals; D002384:Catalysis; D004253:DNA Nucleotidylexotransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D003838:Deoxyadenine Nucleotides / Q000378:metabolism; D003845:Deoxycytosine Nucleotides / Q000378:metabolism; D003848:Deoxyguanine Nucleotides / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D009687:Nuclear Proteins; D009841:Oligonucleotides / Q000737:chemistry / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D013379:Substrate Specificity; D013942:Thymine Nucleotides / Q000378:metabolism; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D003838:Deoxyadenine Nucleotides; D003845:Deoxycytosine Nucleotides; D003848:Deoxyguanine Nucleotides; D009687:Nuclear Proteins; D009841:Oligonucleotides; D013942:Thymine Nucleotides; C029603:deoxyguanosine triphosphate; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; D004253:DNA Nucleotidylexotransferase; C026600:2'-deoxyadenosine triphosphate; C024157:thymidine 5'-triphosphate