Human c-ros-1 gene homologous to the v-ros sequence of UR2 sarcoma virus encodes for a transmembrane receptorlike molecule.

We isolated a human gene (designated c-ros-1) homologous to the v-ros sequence of UR2 sarcoma virus. Ten exons, 1,414 base pairs spanning 26 kilobases, contained a tyrosine kinase domain, a transmembrane domain, and a part of an extracellular domain carrying an N glycosylation site which was not acquired by UR2 sarcoma virus. The predicted structure of c-ros-1 is unique among the src family and clearly distinct from the human insulin receptor.     

草鱼和翘嘴鲌fgfrhl-1基因的克隆和表达分析

成纤维细胞生长因子受体同源类似物1(fgfrhl-1)基因是目前仅在鱼类基因组中检测到的fgfr基因家族成员, 该序列在鱼类进化过程中高度保守。为研究fgfrhl-1基因的表达情况和具体的功能, 在亲缘关系较远的草鱼(Ctenopharyngodon idellus)和翘嘴鲌(Culter alburnus Basilewsky)中克隆了fgfrhl-1的cDNA序列, 并通过半定量RT-PCR和冰冻切片原位杂交分析了该基因在成体不同组织中的表达情况。克隆结果的序列分析表明: 草鱼fgfrhl-1 cDNA序列全长为1472 bp, 5′-UTR长213 bp, 3′-UTR长56 bp, 开放阅读框长1203 bp; 翘嘴鲌fgfrhl-1 cDNA序列全长为1886 bp, 5′-UTR长298 bp, 3′-UTR长385 bp, 开放阅读框长1203 bp。在两种鱼类中该基因都编码400个氨基酸, 其预测的氨基酸序列同源性高达95.5%。蛋白二级结构预测表明Fgfrhl-1具有FGFRs家族蛋白的胞内酪氨酸激酶区, 跨膜的螺旋区和胞外配体识别结合区, 但其胞外区比FGFRs缺少了3个免疫球蛋白样结构域。通过RT-PCR方法在两种鱼类的心脏、鳃、肝、脾、尾鳍以及肌肉组织的肌间隔中均检测到了fgfrhl-1表达, 但在肌纤维中均没有检测到其表达。对这两种鱼类的肌肉组织、肝脏和脾脏进行的组织切片原位杂交表明fgfrhl-1只在这些组织和器官的结缔组织及导管中表达, 不在间质细胞结构中表达。这些结果说明: fgfrhl-1的成体组织特异性表达模式在不同鱼类中基本一致, fgfrhl-1在鱼类各组织和器官的结缔组织和导管的细胞中表达, 不在间质细胞中表达。因此, fgfrhl-1可能在鱼类结缔组织及导管分化调控或功能维持中有独特作用。     

对水稻4号染色体一个编码S位点相关的受体样蛋白激酶基因簇的分析

通过对水稻 (OryzasativaL .) 4号染色体一段 32 3kb的序列测定和分析 ,在其中 10 8kb的区域内发现了一个由 14个编码S位点相关的受体样蛋白激酶 (SRK)基因组成的基因簇。RT_PCR实验证明了这 14个基因中有 9个基因表达 ,并且这 9个基因有不同的表达模式 :其中 2个基因主要在生殖器官中表达 ,而另外 7个基因在水稻的营养和生殖器官中均有表达。对这些基因的预测的氨基酸序列进行分析表明他们的细胞外受体部分均和甘蓝的SLG蛋白高度同源 ,而细胞内的激酶区都包含有丝氨酸 /苏氨酸激酶中特异的氨基酸。     

Mutations in the consensus helicase domains of the Werner syndrome gene. Werner's Syndrome Collaborative Group.

Werner syndrome (WS) is an autosomal recessive disease with a complex phenotype that is suggestive of accelerated aging. WS is caused by mutations in a gene, WRN, that encodes a predicted 1,432-amino-acid protein with homology to DNA and RNA helicases. Previous work identified four WS mutations in the 3' end of the gene, which resulted in predicted truncated protein products of 1,060-1,247 amino acids but did not disrupt the helicase domain region (amino acids 569-859). Here, additional WS subjects were screened for mutations, and the intron-exon structure of the gene was determined. A total of 35 exons were defined, with the coding sequences beginning in the second exon. Five new WS mutations were identified: two nonsense mutations at codons 369 and 889; a mutation at a splice-junction site, resulting in a predicted truncated protein of 760 amino acids; a 1-bp deletion causing a frameshift; and a predicted truncated protein of 391 amino acids. Another deletion is >15 kb of genomic DNA, including exons 19-23; the predicted protein is 1,186 amino acids long. Four of these new mutations either partially disrupt the helicase domain region or result in predicted protein products completely missing the helicase region. These results confirm that mutations in the WRN gene are responsible for WS. Also, the location of the mutations indicates that the presence or absence of the helicase domain does not influence the WS phenotype and suggests that WS is the result of complete loss of function of the WRN gene product.     

Molecular cloning and predicted full-length amino acid sequence of the type I beta isozyme of cGMP-dependent protein kinase from human placenta. Tissue distribution and developmental changes in rat

In this study we report the isolation and characterization of three overlapping cDNA clones for the type I beta isozyme of cGMP-dependent protein kinase (cGK) from human placenta libraries. The composite sequence was 3740 nucleotides long and contained 58 nucleotides from the 5'-noncoding region, an open reading frame of 2061 bases including the stop codon, and a 3'-noncoding region of 1621 nucleotides. The predicted full-length human type I beta cGK protein contained 686 amino acids including the initiator methionine, and had an estimated molecular mass of 77,803 Da. On comparison to the published amino acid sequence of bovine lung I alpha, human placenta I beta cGK differed by only two amino acids in the carboxyl-terminal region (amino acids 105-686). In contrast, the amino-terminal region of the two proteins was markedly different (only 36% similarity), and human I beta cGK was 16 amino acids longer. In a specific region in the amino-terminus (amino acids 63-75), 12 out of 13 amino acids of the human I beta cGK were identical to the partial amino acid sequence recently published for a new I beta isoform of cGK from bovine aorta. Northern blot analysis demonstrated a human I beta cGK mRNA, 7 kb in size, in human uterus and weakly in placenta. An mRNA of 7 kb was also observed in rat cerebellum, cerebrum, lung, kidney, and adrenal, whereas an mRNA doublet of 7.5 and 6.5 kb were observed in rat heart. Comparison of Northern and Western blot analyses demonstrated that the mRNA and protein for cerebellar cGK increased during the development of rats from 5 to 30 days old, whereas the 6.5 kb mRNA in rat heart declined.     

Molecular cloning and sequencing of human palmitoyl-CoA ligase and its tissue specific expression

A complimentary DNA clone encoding the entire human palmitoyl-CoA ligase has been isolated from a liver cDNA library and sequenced in it's entirety. The predicted product is a 699 amino acid protein. Southern analysis utilizing the human palmitoyl-CoA ligase gene as a probe revealed varying degrees of similarity amongst various mammalian species. The palmitoyl-CoA ligase gene is highly expressed in liver, heart, skeletal muscle and kidney, and to a lesser extent in brain, lung, placenta and pancreas. The expression of palmitoyl-CoA ligase in various tissue parallels the function of this enzyme in the metabolism of fatty acids in these tissues.     

Mouse cathepsin F: cDNA cloning, genomic organization and chromosomal assignment of the gene

A murine cysteine protease of the papain family was identified by dbEST-database search. A 1.87kb full-length cDNA encoding a predicted polypeptide of 462 amino acids was sequenced. Since the encoded polypeptide shows more than 80% sequence identity with human cathepsin F, it is most likely that this cDNA represents the murine homologue of cathepsin F, and it was therefore named accordingly. Murine cathepsin F exhibits a domain structure typical for papain-like cysteine proteases, a 20 amino acid N-terminal hydrophobic signal sequence followed by an extraordinarily long propeptide of 228 amino acids and the domain of the mature protease comprising 214 amino acids. The mature region contains all features characteristic of a papain-like cysteine protease, including the highly conserved cysteine, histidine and asparagine residues of the 'catalytic triad'. Genomic clones covering the murine cathepsin F gene were isolated. The mouse cathepsin F gene consists of 14 exons and 13 introns and spans 5.8kb. Murine cathepsin F was mapped to chromosome 19, a region with synteny homology to a region of human chromosome 11 to which human cathepsin F has been mapped previously. Northern blot analysis of RNA from multiple tissues revealed a ubiquitous expression of cathepsin F in mouse and man.     

Molecular cloning and characterization of cathepsin F gene from olive flounder Paralichthys Olivaceus

We isolated a homologue of cathepsin F from cDNA library of olive flounder liver. A 2,077 kb full-length cDNA encoding a predicted polypeptide of 474 amino acids was sequenced. The flounder cathepsin F exhibits a domain structure typical for papain-like cysteine proteases, a 17 amino acid N-terminal hydrophobic signal sequence followed by an extraordinarily long propeptide of 244 amino acids and the domain of the mature protease comprising 213 amino acids. The mature region contains all features characteristic of a papain-like cysteine protease, including the highly conserved cysteine, histidine and asparagine residues of the ‘catalytic triad’. The cathepsin F protein showed 49–99% amino acid sequence identity with other known cathepsin F sequences. An in vivo expression study showed that cathepsin F mRNA was expressed predominantly in brain, liver, eye and heart, and moderately in other tissues. The accumulation of cathepsin F mRNA in early stage of development increased with development. This expression pattern suggests that flounder cathepsin F has been implicated in the growth and reproduction regulation.     

Expression cloning of an activin receptor, a predicted transmembrane serine kinase.

Activins are involved in the regulation of multiple biological events, ranging from early development to pituitary function. To characterize the cellular mechanisms involved in these processes, cDNAs coding for an activin receptor were cloned from AtT20 mouse corticotropic cells by screening COS cell transfectants for binding of 125I-activin A. The cDNAs code for a protein of 494 amino acids comprising a ligand-binding extracellular domain, a single membrane-spanning domain, and an intracellular kinase domain with predicted serine/threonine specificity. 125I-activin A binds to transfected COS cells with an affinity of 180 pM and can be competed by activin A, activin B, and inhibin A, but not by transforming growth factor beta 1. The kinase domain, but not the extracellular sequence, of the activin receptor is most closely related to the C. elegans daf-1 gene product, a putative transmembrane serine/threonine-specific protein kinase for which the ligand is not known.     

Molecular cloning and characterization of an alkalophilic Bacillus sp. C125 gene homologous to Bacillus subtilis sec Y.

A 1.8 kb HindIII DNA fragment containing the secY gene of alkalophilic Bacillus sp. C125 has been cloned into plasmid pUC119 using the B. subtilis secY gene as a probe. The complete nucleotide sequence of the cloned DNA indicated that it contained one complete ORF and parts of two other ORFs. The similarity of these ORFs to the sequences of the B. subtilis proteins indicated that they were the genes for ribosomal protein L15-SecY-adenylate kinase, in that order. The gene product of the alkalophilic Bacillus sp. C125 secY homologue was composed of 431 amino acids and its M(r) value has been calculated to be 47,100. The distribution of hydrophobic amino acids in the gene product suggested that the protein was a membrane integrated protein with ten transmembrane segments. The total amino acid sequence of alkalophilic Bacillus sp. C125 secY homologue showed 69.7% homology with that of B. subtilis secY. Regions of remarkably high homology (78% identity) were present in transmembrane regions, and cytoplasmic domains (73% identity) with less homologous regions present in extracellular domains (43% identity).     

两个鼻咽癌负相关新基因的分离与特性

8个通过 c DNA代表差异分析法 ( c DNA representational difference analysis,c DNA RDA)分离的新 c DNA序列中 ,经 RT- PCR验证 ,发现其中一 c DNA序列 (登录号 :AF0 91 51 7)在 40 %的鼻咽癌活检组织中存在表达缺失和下调 .Northern杂交显示 ,AF0 91 51 7代表转录本为 1 .1 kb和 1 .4kb大小的两个基因 ,进而采用文库筛选 ,成功分离出 3′端完全不同的两个基因 ,命名为 NAG1 1和 NAG 1 2 (登录号分别为 AF 1 70 30 7和 AF 1 94971 ) .经过计算机预测 ,NAG 1 1编码 87个氨基酸组成的跨膜蛋白 ,NAG1 2编码 1 36个氨基酸组成的可溶性的核蛋白 ,两者无任何同源性 .NAG 1 1蛋白含有 3个 ATP结合区、两个蛋白激酶 C磷酸化位点和两个 N-肉豆寇酸化位点 ,NAG 1 2含有POU结构域和多个功能位点 .结果说明 NAG1 1和 NAG1 2的表达的缺失与下调可能参与了鼻咽癌的进程 ;NAG1 1基因产物可能与 ATP的跨膜转运有关 ;NAG1 2基因产物可能与转录翻译有关 .     

The region of the IncN plasmid R46 coding for resistance to beta-lactam antibiotics, streptomycin/spectinomycin and sulphonamides is closely related to antibiotic resistance segments found in IncW plasmids and in Tn21-like transposons.

The nucleotide sequence of a 2.5 kb segment of the pKM101 (R46) genome has been determined. The 1.3 kb from a BamHI site at 153 to base 1440 differs by only 2 bases from a part of the published sequence of the aadB (gentamicin resistance) gene region including the coding region for the N-terminal 70 amino acids of the predicted aadB product. The same sequence has been found 5'-to the dhfrII gene of R388 and to the aadA gene of Tn21 (R538-1). Three open reading frames are located in this region, two on the same strand as the resistance genes and one on the complementary strand. The latter predicts a polypeptide of 337 amino acids, whose N-terminal segment is 40% homologous to the predicted product of an open reading frame of 179 amino acids located next to the dhfrI gene of Tn7. The oxa2 (oxacillin resistance) gene predicts a long polypeptide commencing with (the N-terminal) 70 amino acids of the aadB product. A similar arrangement is found in the aadA gene of R538-1. The N-terminal segment of an aadA gene is located 3'- to oxa2, separated by 36 bases. Sequences surrounding the BamHI site are identical to sequences 5'- to the tnpM gene of Tn21 and homology ceases where homology between Tn21 and Tn501 commences. The possibility that this antibiotic resistance segment is a discrete mobile DNA element is discussed.     

TTID: A novel gene at 5q31 encoding a protein with titin-like features.

A deletion of the long arm of chromosome 5 is a recurring abnormality in malignant myeloid disorders. In previous studies, we identified an approximately 1-Mb segment in 5q31 that was deleted in all patients examined. As part of a positional cloning project to identify transcribed sequences in this region, we identified and characterized the TTID gene. This gene contains 10 exons that extend over 19 kb. The composite cDNA is approximately 2.3 kb and encodes a protein of 498 amino acids, with a predicted molecular mass of 55 kDa. The C-terminal half of this putative protein contains an internally repeated domain of 43 amino acids, which resembles the N-terminal half of an immunoglobulin domain from the immense skeletal muscle protein titin. The TTID gene is expressed in multiple muscle tissue types as well as in thyroid gland and bone marrow. We evaluated the gene as a candidate tumor suppressor gene by searching for mutations in malignant myeloid disorders with abnormalities of chromosome 5. However, we detected no inactivating mutations. A single nucleotide change (G to A) was identified at nucleotide position 1889 in the untranslated region of the mRNA, which may represent a polymorphism. Therefore, TTID is unlikely to be the candidate tumor suppressor gene involved in malignant myeloid disorders.