Le Guen L, Thomas M, Kreis M
Université de Paris-Sud, IBP, URA CNRS 1128, Orsay, France.
Mol Gen Genet. 1994 Nov 1;245(3):390-6. doi: 10.1007/BF00290120.
We have characterized a 6.4 kb genomic fragment from Arabidopsis thaliana ecotype Columbia overlapping the 5' end of the AKin10 gene which encodes a protein Ser/Thr kinase. Using, as probes, various restriction fragments located upstream of AKin10, two cDNA clones have been isolated from a cDNA library prepared from young shoot tissue. A comparison between the cDNA and the above genomic sequences allowed us to locate two novel genes, Atcys1 and Athyp1 (for Arabidopsis thaliana cystathionine gamma-synthase 1 and hypothetical protein 1). The coding sequences of both genes are interrupted by introns and the exons match the sequences of the corresponding cDNAs. Further analysis of the genomic fragment revealed the presence of an open reading frame (ORF) of 609 nucleotides situated between the two genes. Atcys1, Athyp1, AKin10 and the ORF are very close to each other and organized in the same polarity; hence, the intergenic regions probably contain, within less than 0.5 kb, all the regulatory elements necessary to govern initiation and termination of transcription. The deduced protein sequence of Atcys1 shows a high degree of similarity with the cystathionine gamma-synthase from Escherichia coli. The putative product of the Athyp1 gene contains seven hydrophobic regions flanked by hydrophilic domains, reminiscent of membrane-spanning proteins. Southern blot hybridization experiments suggest the presence of one copy of Atcys1, Athyp1 and AKin10 per haploid genome, and Northern blot analysis demonstrates that the three genes are differentially expressed in roots, shoots and leaves.
我们鉴定了来自拟南芥生态型哥伦比亚的一个6.4 kb基因组片段,该片段与编码蛋白丝氨酸/苏氨酸激酶的AKin10基因的5'端重叠。使用位于AKin10上游的各种限制性片段作为探针,从幼嫩茎组织制备的cDNA文库中分离出两个cDNA克隆。通过比较cDNA与上述基因组序列,我们定位了两个新基因,Atcys1和Athyp1(分别代表拟南芥胱硫醚γ-合酶1和假定蛋白1)。这两个基因的编码序列都被内含子打断,外显子与相应cDNA的序列匹配。对该基因组片段的进一步分析揭示了在这两个基因之间存在一个609个核苷酸的开放阅读框(ORF)。Atcys1、Athyp1、AKin10和ORF彼此非常接近且极性相同;因此,基因间区域可能在不到0.5 kb的范围内包含调控转录起始和终止所需的所有调控元件。Atcys1推导的蛋白质序列与大肠杆菌的胱硫醚γ-合酶具有高度相似性。Athyp1基因的推定产物包含七个被亲水区侧翼的疏水区,类似于跨膜蛋白。Southern杂交实验表明每个单倍体基因组中存在一个Atcys1、Athyp1和AKin10拷贝,Northern印迹分析表明这三个基因在根、茎和叶中差异表达。