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.
Harold C. Simmons Arthritis Research Center, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas 75235, USA.
S
S
Mickelsen
C
C
Snyder
K
K
Trujillo
M
M
Bogue
D B
DB
Roth
K
K
Meek
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