The use of methylthioadenosine phosphorylase activity to select for human chromosome 9 in interspecies and intraspecies hybrid cells.
The use of methylthioadenosine phosphorylase activity to select for human chromosome 9 in interspecies and intraspecies hybrid cells.
复制标题
利用甲硫腺苷磷酸化酶活性在种间和种内杂交细胞中选择人类 9 号染色体。
DOI：
10.1007/bf01233252
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Diaz,MO
中科院分区：
文献类型：
--
作者：
Porterfield,BW;Pomykala,H;Maltepe,E;Bohlander,SK;Rowley,JD;Diaz,MO
甲基硫代腺苷磷酸化酶（MTAP）是一种在腺嘌呤合成的补救性途径中起作用的酶。通过对小鼠与人体细胞杂交的分析，已将编码MTAP活性的位点定位到人9号染色体（9q12-9pter）上。具有MTAP活性的细胞将停止增殖，并最终在azaserine（一种新的嘌呤合成抑制剂）的存在下死亡，但可以通过向培养基中添加甲基硫代腺苷（MTA）来挽救。一些小鼠和人肿瘤细胞缺乏MTAP活性，在azazerine和MTA存在下不能生长。我们将MTAP表达的人成纤维细胞与MTAP缺失的小鼠l细胞融合，并在含有azaserine和MTA的培养基中选择含有MTAP活性的体细胞杂交。在另一项实验中，CHO细胞与人成纤维细胞体细胞杂交，包含人类9号染色体的正常拷贝，用于制备微细胞，并将其融合到mtap缺陷的人白血病细胞系CCRF-CEM中。利用荧光原位杂交探针与人类9号染色体（9p21）上的干扰素α和β 1基因以及人类9号染色体的着丝粒杂交，发现体细胞和微细胞杂种保留了人类9号染色体。这是首次利用MTAP活性的互补来选择保留人类9号染色体的体细胞杂交种和微细胞杂交种。
Methylthioadenosine phosphorylase (MTAP) is an enzyme that functions in a salvage pathway for adenine synthesis. The locus that encodes MTAP activity has been mapped to human chromosome 9 (9q12-9pter) by analysis of mouse × human somatic cell hybrids. Cells that have MTAP activity will stop proliferating, and eventually die in the presence of azaserine, an inhibitor of de novo purine synthesis, but can be rescued by the addition of methylthioadenosine (MTA) to the culture medium. Some mouse and human tumor cells lack MTAP activity and can not grow in the presence of azaserine and MTA. We fused MTAP competent human fibroblast cells to MTAP deficient mouse L-cells and selected for somatic cell hybrids, containing MTAP activity, in medium containing azaserine and MTA. In a separate experiment, a CHO cell × human fibroblast somatic cell hybrid, containing a normal copy of human chromosome 9, was used to prepare microcells, which were fused to an MTAP-deficient human leukemic cell line, CCRF-CEM. Somatic cell and microcell hybrids were shown to retain human chromosome 9 by fluorescence in situ hybridization using probes that hybridize to the interferon-alpha and-beta 1 genes on human chromosome 9 (9p21), and the centromere of human chromosome 9. This is the first report of complementation for MTAP activity being used to select for somatic cell hybrids and microcell hybrids that retain a human chromosome 9.