ISOLATION OF THE GENE ENCODING ADENYLATE-CYCLASE IN SACCHAROMYCES-CEREVISIAE
ISOLATION OF THE GENE ENCODING ADENYLATE-CYCLASE IN SACCHAROMYCES-CEREVISIAE
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DOI：
10.1073/pnas.82.15.5060
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发表时间：
1985-01-01
影响因子：
11.1
通讯作者：
BOURNE, HR
中科院分区：
文献类型：
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作者：
CASPERSON, GF;WALKER, N;BOURNE, HR
通过对酿酒酵母cyr1-1突变的互补，分离出含有腺苷酸环化酶催化单元编码结构基因的酵母基因组DNA。分离的DNA恢复了cry1-1突变体的腺苷酸环化酶活性，并指导了CYR1位点的整合。用CYR1 DNA在高拷贝数载体YEp24上转化的野生型菌株，其腺苷酸环化酶活性比携带无插入质粒的菌株高4- 6倍。这一结果表明，CYR1基因产物的表达，而不是其他多肽组成的腺苷酸环化酶系统，限制了酿酒酵母总腺苷酸环化酶的活性。含有CYR1-的质粒还补充了细胞分裂周期突变cdc35-1的温度敏感生长缺陷，该缺陷在限制性条件下具有与CYR1- 1相似的表型，并且定位于相同的位点。此外，cdc35-1 cam突变体在外源cAMP存在的限制性温度下生长，该突变体包含使其能够从培养基中吸收cAMP的突变。这些观察结果支持了CDC35和CYR1是等位基因的观点，并证实了细胞通过细胞分裂周期起始位置需要cAMP合成的假设。
By complementation of the cyr1-1 mutation in S. cerevisiae, yeast genomic DNA was isolated containing the structural gene that encodes the catalytic unit of adenylate cyclase. The isolated DNA restored adenylate cyclase activity to cry1-1 mutants and directed integration at the CYR1 locus. Wild-type strains transformed with CYR1 DNA on the high copy number vector YEp24 contained 4- to 6-fold more adenylate cyclase activity than strains carrying the plasmid with no insert. This result suggests that expression of the CYR1 gene product, rather than that of other polypeptide components of the adenylate cyclase system, limits total adenylate cyclase activity in S. cerevisiae. CYR1- containing plasmids also complemented the temperature-sensitive growth defect of the cell division cycle mutation cdc35-1, which confers a phenotype under restrictive conditions similar to that of cyr1-1 and maps to the same locus. Further, cdc35-1 cam mutants, which contain mutations that enable them to take up cAMP from the medium, grew at the restrictive temperature in the presence of exogenous cAMP. These observations support the view that CDC35 and CYR1 are allelic and confirm the hypothesis that cAMP synthesis is required for cells to pass through the start position of the cell division cycle.