Southern blot analysis of T-cell receptor gene rearrangements in cynomolgus monkeys, and identification of a progenitor cell HPRT mutation.
Increases in peripheral blood T-lymphocyte HPRT mutant frequency may reflect either a number of independent HPRT gene mutational events or clonal proliferation of a single HPRT mutant. Sequence analysis of HPRT mutations in conjunction with T-cell receptor (TCR) gene rearrangement pattern analysis can distinguish these possibilities. Our laboratory previously characterized a nonhuman primate model for in vivo mutation studies using the clonal HPRT mutation assay. In the present study we report the use of probes for human TCR beta and gamma genes to characterize TCR rearrangements in cynomolgus monkeys. Together, these methods were used to examine a monkey which exhibited a mean spontaneous HPRT mutant frequency (MF) of 16.4 x 10(-6), compared to the normal mean MF of 3.03 x 10(-6). The elevated MF resulted from the occurrence of a single HPRT mutation in a lymphocyte progenitor cell or stem cell, since T-cell clones isolated from the monkey exhibited a G to T transversion at base pair 539 in the HPRT coding region, and had unique rearrangements of TCR gamma along with an apparent germline TCR beta configuration. In a preliminary in vivo mutation study, the animal was treated with the investigational potent mutagen and antitumor agent adozelesin (U-73975). No increase in HPRT mutant frequency was observed. The HPRT mutant clones isolated after treatment showed rearrangement of both TCR gamma and beta genes. Possible explanations for these findings are discussed.
Mattano
S S
SS
Upjohn Company, Kalamazoo, Michigan 49001, USA.
Zimmer
D M
DM
Harbach
P R
PR
Hunter
T C
TC
Aaron
C S
CS
D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000633:toxicity; D001482:Base Composition; D001483:Base Sequence; D001572:Benzofurans; D015139:Blotting, Southern; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003509:Cyclohexanecarboxylic Acids / Q000633:toxicity; D053138:Cyclohexenes; D017931:DNA Primers / Q000737:chemistry; D015507:Drugs, Investigational; D000080890:Duocarmycins; D015329:Gene Rearrangement, T-Lymphocyte / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000187:drug effects / Q000235:genetics*; D007211:Indoles; D008252:Macaca fascicularis; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009693:Nucleic Acid Hybridization; D011948:Receptors, Antigen, T-Cell / Q000187:drug effects / Q000235:genetics*; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects
D018906:Antineoplastic Agents, Alkylating; D001572:Benzofurans; D003509:Cyclohexanecarboxylic Acids; D053138:Cyclohexenes; D017931:DNA Primers; D015507:Drugs, Investigational; D000080890:Duocarmycins; D007211:Indoles; D011948:Receptors, Antigen, T-Cell; C060090:adozelesin; D007041:Hypoxanthine Phosphoribosyltransferase