Saint-Jacques E, Séguin C
Centre de recherche en cancérologie de l'Université Laval, L'Hôtel-Dieu de Québec, Canada.
DNA Cell Biol. 1993 May;12(4):329-40. doi: 10.1089/dna.1993.12.329.
A cDNA encoding Xenopus laevis metallothionein (MT) was cloned from a cDNA library constructed using liver poly(A+)RNA of X. laevis adult males treated with CdCl2. The probe used to screen the library was a MT-specific DNA fragment obtained by means of the polymerase chain reaction (PCR) and degenerate oligodeoxynucleotide primers. The cDNA clone encodes a putative protein of 62 amino acids, of which 20 are cysteine residues. The position of all the cysteine residues is conserved with respect to mammalian MT sequences. The amino acid sequence of this X. laevis MT, designated XIMT-A, shares between 60% and 67% identity with various vertebrate MTs. Overall, the structure of XIMT-A is no similar in sequence to MT-1 than it is to MT-2 isoforms of various vertebrates. Ten different X. laevis MT cDNA isolates were partially sequenced and turned out to be identical, suggesting a single species of MT mRNA. Southern blot analysis of X. laevis DNA reveals that the XlMT-A gene is present in at least two copies. This result is consistent with the suggestion that a genome duplication occurred in a X. laevis ancestor. The in vivo response to increasing doses of Cd2+, Zn2+, and Cu2+ metal salts was tested. In the liver, all three metals proved to be potent inducers, raising MT mRNA levels between 50- and 100-fold. The maximum response to Cd2+ was at 12 hr after injection and to Zn2+ at 24 hr after injection. High levels of mRNA were maintained for more than 48 hr. Cd2+ and Zn2+ induced XlMT-A mRNA in all tissues examined (kidney, spleen, heart, intestine, testes, and brain). Dexamethasone did not induce MT mRNA synthesis in the liver.
从用氯化镉处理过的非洲爪蟾成年雄性个体肝脏的聚腺苷酸（poly(A+)）RNA构建的cDNA文库中，克隆出了一个编码非洲爪蟾金属硫蛋白（MT）的cDNA。用于筛选该文库的探针是一个通过聚合酶链反应（PCR）和简并寡脱氧核苷酸引物获得的MT特异性DNA片段。该cDNA克隆编码一个推定的含62个氨基酸的蛋白质，其中20个是半胱氨酸残基。所有半胱氨酸残基的位置相对于哺乳动物MT序列是保守的。这个被命名为XIMT - A的非洲爪蟾MT的氨基酸序列与各种脊椎动物的MT有60%至67%的同一性。总体而言，XIMT - A的结构在序列上与MT - 1的相似性并不高于与各种脊椎动物MT - 2亚型的相似性。对十个不同的非洲爪蟾MT cDNA分离株进行了部分测序，结果显示它们是相同的，这表明存在单一的MT mRNA种类。对非洲爪蟾DNA的Southern印迹分析表明，XlMT - A基因至少以两个拷贝存在。这一结果与在非洲爪蟾祖先中发生过一次基因组加倍的推测一致。测试了对递增剂量的镉离子（Cd2+）、锌离子（Zn2+）和铜离子（Cu2+）金属盐的体内反应。在肝脏中，所有这三种金属都被证明是有效的诱导剂，使MT mRNA水平提高了50至100倍。对Cd2+的最大反应出现在注射后12小时，对Zn2+的最大反应出现在注射后24小时。高水平的mRNA维持了超过48小时。Cd2+和Zn2+在所有检测的组织（肾脏、脾脏、心脏、肠道、睾丸和大脑）中都诱导了XlMT - A mRNA的产生。地塞米松没有在肝脏中诱导MT mRNA的合成。
