Expression of recombination-activating genes and T cell receptor gene recombination in the human T cell leukemia cell line.
BACKGROUND
Recent studies have suggested that mature T cells can change their specificity through reexpression of recombination-activating genes (RAG) and RAG-mediated V(D)J recombination. This process is named receptor revision and has been observed in mature peripheral T cells from transgenic mice and human donors. However, whether thebreceptor revision in mature T cells is a random or orientated process remains poorly understood. Here we used the Jurkathuman T cell line, which represents a mature stage of T cell development, as a model to investigate the regulation of Tcell receptor (TCR) gene recombination.


METHODS
TCR Dbeta-Jbeta signal joint T cell receptor excision DNA circles (sjTRECs) were determined by nested and seminested PCR. Double-strand DNA breaks at recombination signal sequences (RSSs) in the TCRVbeta chain locus were detected by ligation-mediated-PCR. Further analysis of the complementarity-determining region 3 (CDR3) size of the TCRVbeta chain was examined by the TCR GeneScan technique.


RESULTS
RAG1, RAG2, and three crucial components of the nonhomologous DNA end-joining (NHEJ) pathway were readily detected in Jurkat. Characteristics of junctional diversity of Dbeta2-Jbeta2 signal joints and ds RSS breaks associated with the Dbeta2 5' and Dbeta 2 3' sites were detected in DNA from Jurkat cells. CDR3 size and the gene sequences of the TCRVbeta chain did not change during cell proliferation.


CONCLUSIONS
RAG1 and RAG2 and ongoing TCR gene recombination are coexpressed in Jurkat cells, but the ongoing recombination process may not play a role in modification of the TCR repertoire.However, the results suggest that Jurkat could be used as a model for studying the regulation of RAGs and V(D)J recombination and as a "special" model of the coexistence of TCR gene rearrangements and "negative" receptor revision.
Zou
Hong-yun
HY
Institute of Molecular Immunology, School of Biotechnology, Southern Medical University, Guangzhou 510515, China.
Ma
Li
L
Meng
Min-jie
MJ
Yao
Xin-sheng
XS
Lin
Ying
Y
Wu
Zhen-qiang
ZQ
He
Xiao-wei
XW
Wang
Ju-fang
JF
Wang
Xiao-ning
XN
D034961:Antigens, Nuclear / Q000235:genetics; D001483:Base Sequence; D022801:Complementarity Determining Regions; D053960:DNA Breaks; D004268:DNA-Binding Proteins / Q000235:genetics*; D016654:Genes, RAG-1; D019672:Genes, T-Cell Receptor; D006801:Humans; D019169:Jurkat Cells; D000072200:Ku Autoantigen; D015458:Leukemia, T-Cell / Q000235:genetics*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics*; D011995:Recombination, Genetic
D034961:Antigens, Nuclear; D022801:Complementarity Determining Regions; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; C499311:RAG2 protein, human; C000606509:Xrcc6 protein, human; C000606510:Xrcc6 protein, mouse; D000072200:Ku Autoantigen