Determination of the genotype of a panel. of human tumor cell lines for the human homologues of yeast cell cycle checkpoint control genes: Identification of cell lines carrying homoallelic missense base substitutions

A number of human homologues of yeast cell cycle checkpoint control genes have been identified recently. In this study, the sequence alterations in six of such novel human genes (hRAD1, hRAD9, hRAD17, hHUS1, CHK1 and CHES1) were analyzed by PCR-single-strand conformational polymorphism (PCR-SSCP) method on a panel of 25 human tumor cell lines in an attempt to search for possible in vivo cases where any of the checkpoint-related genes are altered in human systems. For hRAD9, hHUS1 or CHK1, no SSCP variant was detected in any of the cell lines tested indicating a high stability of these genes in human cancer. Most of the SSCP variants found in the other three genes were due to single nucleotide base substitutions. Two cell lines were found to be homozygous for missense-type base substitutions, i.e., Saos-2 was homoallelic for 1637T --> G in hRAD 17; and COLO320DM for 1189G --> A in CHES1, indicating a possible use of these cell lines for further study. The former nucleotide change in hRAD17, which causes a change of amino acid from arginine to lysine at codon 546, was supposed to be polymorphic. Considering that lysine, but not arginine, is the amino acid that is well conserved among fission yeast, mouse and monkey at the corresponding position, coexistence of both alleles in human may have a functional or selectional implication.