ANALYSIS OF DOMINANT-NEGATIVE MUTATIONS OF THE CAENORHABDITIS-ELEGANS LET-60 RAS GENE
ANALYSIS OF DOMINANT-NEGATIVE MUTATIONS OF THE CAENORHABDITIS-ELEGANS LET-60 RAS GENE
复制标题
DOI：
10.1101/gad.5.12a.2188
复制
发表时间：
1991-12-01
影响因子：
10.5
通讯作者：
STERNBERG, PW
中科院分区：
文献类型：
--
作者：
HAN, M;STERNBERG, PW
秀丽隐杆线虫的let-60基因根据性腺的诱导信号控制外阴和皮下分化的选择。Let-60编码一种ras蛋白，在信号转导途径中作用于let-23受体酪氨酸激酶的下游。let-60(Vet-60(dn))]的显性负突变导致let-60(dn)/+杂合子基因活性降低和外阴少突变表型。我们发现9个let-60（dn）突变导致保守残基的替换。其中四个处于两个新位置；其他基因位于已知的位置，会在哺乳动物细胞中引起显性负突变。这些病变的位置表明它们破坏了ras蛋白结合鸟嘌呤核苷酸的能力。将4个let-60（dn）突变基因以染色体外阵列的形式引入野生型动物，发现根据基因剂量和等位基因的不同，产生三种显性表型——致死性、无外阴或多外阴。高剂量转基因let-60（dn）基因引起的显性致死表明这些突变基因在早期发育中具有毒性作用。杂合子中这些突变的显性负效应可能是由let-60（dn）和let-60（+）蛋白之间争夺一个正调节因子引起的。所有的let-60（dn）突变都会干扰let-60（+）的活性，但一些等位基因具有部分构成活性，这表明与激活子相互作用的能力与发挥生理作用（刺激外阴分化）的能力是可分离的。这些dn突变可能有助于干扰其他多细胞生物中ras介导的信号转导途径。
The let-60 gene of Caenorhabditis elegans controls the choice between vulval and hypodermal differentiation in response to an inductive signal from the gonad. let-60 encodes a ras protein that acts downstream of the let-23 receptor tyrosine kinase in a signal transduction pathway. Dominant-negative mutations of let-60 Vet-60(dn)] cause a reduction of the gene activity in let-60(dn)/+ heterozygotes and a vulva-less mutant phenotype. We have found that nine let-60(dn) mutations cause replacements of conserved residues. Four are in two novel positions; others are in positions known previously to cause dominant-negative mutations in mammalian cells. The locations of these lesions suggest that they disrupt the ability of the ras protein to bind guanine nucleotides. Four let-60(dn) mutant genes were introduced into wild-type animals in the form of extrachromosomal arrays and were found to generate three dominant phenotypes-lethality, vulva-less, or multivulva-depending on gene dose and alleles. The dominant lethality caused by high-dose transgenic let-60(dn) genes suggests a toxic effect of these mutant genes in early development. The dominant-negative effects of these mutations in heterozygotes are likely to be caused by competition between let-60(dn) and let-60(+) protein for a positive regulator. All let-60(dn) mutations interfere with let-60(+) activity, but some alleles have partial constitutive activity, suggesting that the ability to interact with the activator is separable from the ability to exert a physiological effect (stimulation of vulval differentiation). These dn mutations might be useful for interfering with ras-mediated signal transduction pathways in other multicellular organisms.