癌基因PPO及其同源性基因的结构分析

目的采用PPO(proliferation and phosphorylation oncogene)基因与其同源性基因的结构分析,推测PPO并验证其功能。方法利用PPO的蛋白序列寻找同源基因,比较其结构并利用蛋白免疫杂交方法证实其功能。结果克隆了PPO基因,找到了PPO基因与小鼠、果蝇、蚊子、线虫以及酵母等的同源基因,比较发现PPO在进化上非常保守。人和小鼠的氨基酸序列有许多与磷酸化有关的功能域,并发现其中某些氨基酸在进化上非常保守。进一步研究表明PPO能使ERK2和MEK磷酸化。结论PPO基因在进化上非常保守,与磷酸化功能有关。     

中国广西狂犬病毒野毒株（CGX89－1株）糖蛋白基因核酸序列测定和分析

本文报告了中国广西狂犬病毒野毒株（ＣＧＸ８９－１株）糖蛋白基因ｃＤＮＡ的核苷酸序列及其推导的氨基酸序列。ＣＧＸ８９－１株的糖蛋白基因从起始密码ＡＴＧ到终止密码ＴＡＡ共有１５７５个核苷酸残基，可编码形成５２４个氨基酸残基的多肽链，经修饰后形成具有５０５个氨基酸残基构成的狂犬病毒糖蛋白。ＣＧＸ８９－１株的糖蛋白基因和核苷酸组成分别为：Ａ占２７．１１％，Ｔ占２６．２９％，Ｃ占２１．９７％和Ｇ占２４．６３％。核苷酸序列和巴斯德株（ＰＶ株），国际标准攻击毒株（ＣＶＳ株），中国狂犬病毒疫苗株（３ａＧ株）相比，同源性分别为８４．１％，８３．１％和８４．５％。其推导的氨基酸序列和ＰＶ株、ＣＶＳ株和３ａＧ株相比其同源性分别为９２．４％，８９．７％和８９．５％。ＣＧＸ８９－１株也具有３个Ｎ－糖基化位点，分别位于第３７位、１５７位和３１９位的氨基酸残基上。糖蛋白的膜外区部分重要抗原位点和ＰＶ株、ＣＶＳ株及３ａＧ株具有较高的一致性。     

Characterization of a new subgroup of human Ig V lambda cDNA clone and its expression.

From a human bone marrow cDNA library, we have cloned and sequenced a gene which cross-hybridized with murine pre-B cell-specific gene 8HS-20 cDNA under the low-stringent condition. Sequence analysis predicted that this gene (YM-1) encoded 240 amino acids which had the basic structure of immunoglobulin lambda light chain. The 3' half of the YM-1 sequence was identical to the J lambda 2 C lambda 2 region except for four nucleotides. The 5' part of the gene had 87.6% sequence homology with the reported V lambda gene called T1. Comparison of the deduced amino acid sequences with representative members of the seven other V lambda subgroups showed considerable structural homology, but the maximum homology with these chains was 44%. Therefore, we conclude that YM-1 belongs to a new V lambda subgroup. Interestingly YM-1 showed higher homology with VpreB1 (56%) than with any of the other V lambda subgroups. By Southern blot analysis four to six cross-hybridizing V lambda bands were detected at high stringency. Expression of the V lambda gene was observed in immature as well as mature B cell lines without accompanying V-JC gene joining, suggesting that V lambda of the YM-1 locus is activated at the early stage of maturation.     

Nucleotide sequence,serology and symptomatology suggest that vanilla necrosis potyvirus is a strain of watermelon mosaic virus II

Summary Vanilla necrosis potyvirus (VNV) is the cause of significant losses to the South Pacific islands vanilla crop. The gene for the coat protein of VNV has been cloned and sequenced. Comparison of this gene with other potyviral coat protein sequences revealed 97% nucleotide sequence homology (98% amino acid homology) to a US isolate of watermelon mosaic virus II (WMV-II), 93% nucleotide sequence homology (96% amino acid homology) to an Australian isolate of WMV-II and 81% nucleotide sequence homology (88% amino acid homology) to soybean mosaic virus-N (SMV-N). Serological analysis, by Western blot and ELISA, confirmed the close relationship between VNV and WMV-II. Furthermore, a limited host range determination found VNV and WMV-II able to infect the same series of test plants. However, symptoms differed significantly on three test species demonstrating that VNV and WMV-II are not identical in biological properties. We suggest that VNV be renamed WMV-II (Tonga).     

Occurrence of nuclear-encoded tRNAIle in mitochondria of the liverwort Marchantia polymorpha

 Although 29 tRNA genes have been deduced from the complete nucleotide sequence of the mitochondrial genome from the liverwort Marchantia polymorpha, a tRNA^Ile gene decoding AUU and AUC codons is conspicuously absent. In order to address the question of the possible involvement of nuclear-encoded tRNA, we isolated and identified three variant copies of the nuclear-encoded tRNA^Ile(AAU) gene from the liverwort. Northern analysis showed the presence of nuclear-encoded tRNA^Ile both in the mitochondrion and the cytosol, while both chloroplast DNA-encoded tRNA^Ile and nuclear-encoded tRNA^Tyr were absent in liverwort mitochondria. These results unequivocally establish that import of nuclear tRNA^Ile into mitochondria indeed occurs in one of the most primitive plants, M. polymorpha. Received: 11 January 1996/18 March 1996     

中国基因3型乙型脑炎病毒E基因分子特征

目的 以减毒活疫苗(SA14-14-2株)为对照,分析我国分离的基因3型乙脑病毒E基因区段核苷酸及氨基酸序列分子特征.方法 从GenBank中获取相应乙脑病毒株E基因区段核苷酸序列,通过Clustal X(1.81)、DNAStar、GENEDOC(3.2)等生物学软件进行核苷酸和氨基酸位点差异分析.以蜱传脑炎病毒可溶性蛋白晶体结构为模板进行乙脑病毒E蛋白氨基酸位点分析.结果 我国不同地域、不同宿主分离的基因3型乙脑病毒与SA14-14-2株核苷酸同源性分别在96%和95%以上,氨基酸同源性在95%和94%以上.在同一地域、同一宿主类型分离的毒株之间核苷酸和氨基酸同源性非常高.在E基因区段存在10处共同的氨基酸位点差异,在结构域Ⅰ(E160)、结构域Ⅱ(E123和E227)和两个未在结构域中的氨基酸位点(E441和E487)等5个位点在部分基因3型乙脑病毒中存在差异.结论 我国分离的基因3型乙脑病毒与减毒活疫苗株(SA14-14-2株)E基因区段同源性高,存在5处基因3型乙脑病毒特异的氨基酸位点差异,但现行减毒活疫苗株理论上可以保护我国分离的基因3型乙脑病毒野毒株.     

The evolution of the Calvin cycle from prokaryotic to eukaryotic chromosomes: a case study of functional redundancy in ancient pathways through endosymbiosis

The evolutionary histories of the 12 enzymes that catalyze the reactions of the Calvin cycle in higher-plant chloroplasts are summarized. They are shown to be encoded by a mixture of nuclear genes of cyanobacterial and proteobacterial origin. Moreover, where cytosolic isoforms of these enzymes are found they are almost invariably encoded by genes of clearly endosymbiont origin. We infer that endosymbiosis resulted in functional redundancy that was eliminated through differential gene loss, with intruding eubacterial genes repeatedly replacing pre-existing nuclear counterparts to which they were either functionally or structurally homologous. Our findings fail to support the `product-specificity corollary', which predicts re-targeting of nuclear-encoded gene products to the organelle from whose genome they originated. Rather it would appear that the enzymes of central carbohydrate metabolism have evolved novel targeting possibilities regardless of their origins. Our findings suggest a new hypothesis to explain organelle genome persistence, based on the testable idea that some organelle-encoded gene products might be toxic when present in the cytosol or other inappropriate cellular compartments. Received: 8 January / 20 May 1997     

Intronic deletion affecting a negative regulatory region of TP73 is related to breast and colorectal carcinomas

The TP73 gene encodes a nuclear protein that has high homology with TP53. TP73 is rarely mutated in human cancer. The presence of a 1-kb regulatory fragment within the first intron of TP73 was recently reported. This fragment exerts silencer activity on TP73 mediated by ZEB. We searched for possible mutations in this negative regulatory region in 45 colorectal and 43 breast cancer patients and in 34 healthy donors. The study was carried out using the SSCP method, and the allelic variants detected were sequenced. The expression of TP73 was analyzed by quantitative RT-PCR, and loss of heterozygosity (LOH) was assessed by microsatellite study. In several samples, we identified an allele variant that corresponds to a deletion of 73 bp in tumor tissues and normal counterparts, localized between -489 and -417 from the ATG start site of exon 2. Among the 88 tumor samples, 35 (40%) showed at least 1 allele with the cited deletion, versus 7 of the 34 (21%) healthy donors (P = 0.045). When we classified the patients according to the number of variations into homozygous or heterozygous groups, the significance was clearer (P = 0.03). No LOH was detected in the heterozygous cases. There was a positive quantitative correlation between the expression of TP73 and the presence of the allelic variant (P = 0.029). These data suggest that this allelic variant is common in breast and colorectal cancers and that it could alter the expression of the TP73 gene with an additive effect.     

Mitochondrial DNA of Chlamydomonas reinhardtii: the DNA sequence of a region showing homology with mammalian URF2

Summary A 1.27 kb DNA fragment of the 15 kb DNA of Chlamydomonas reinhardtii has been cloned and sequenced. A 906 bp long open reading frame was found showing homology with the URF2 genes of mammals and insects. This homology is functional evidence for Chlamydomonas reinhardtii 15 kb DNA representing indeed mitochondrial DNA. This is the first report of an URF2 gene in mitochondria of a photosynthetic organism. The absence of a TGA codon within the gene suggests that it is used as stop codon like in higher plants and not as tryptophan like in animal and fungal mitochondria.     

A plastid-targeted heat shock cognate 70 kDa protein interacts with the Abutilon mosaic virus movement protein

The movement protein (MP) of bipartite geminiviruses facilitates cell-to-cell as well as long-distance transport within plants and influences viral pathogenicity. Yeast two-hybrid assays identified a chaperone, the nuclear-encoded and plastid-targeted heat shock cognate 70 kDa protein (cpHSC70-1) of Arabidopsis thaliana, as a potential binding partner for the Abutilon mosaic virus (AbMV) MP. In planta, bimolecular fluorescence complementation (BiFC) analysis showed cpHSC70-1/MP complexes and MP homooligomers at the cell periphery and co-localized with chloroplasts. BiFC revealed cpHSC70-1 oligomers associated with chloroplasts, but also distributed at the cellular margin and in filaments arising from plastids reminiscent of stromules. Silencing the cpHSC70 gene of Nicotiana benthamiana using an AbMV DNA A-derived gene silencing vector induced minute white leaf areas, which indicate an effect on chloroplast stability. Although AbMV DNA accumulated within chlorotic spots, a spatial restriction of these occurred, suggesting a functional relevance of the MP-chaperone interaction for viral transport and symptom induction.     

烟台沿海地区人源和猪源戊型肝炎病毒基因型别的研究

目的调查烟台市沿海地区人源与猪源戊型肝炎病毒（HEV）基因型别的相关性。方法应用逆转录一巢式聚合酶联反应（RT—nPcR）方法对当地急性散发戊型肝炎患者、正常人群中抗HEV-IgM阳性者和当地养猪场的猪进行HEVRNA检测,并对HEVRNA阳性标本进行克隆测序和序列分析。结果16例急性散发戊型肝炎患者中有7例粪便标本HEVRNA阳性;51份IgM阳性正常人群血清标本中有1份HEVRNA阳性;34份猪胆汁标本中有1份HEVRNA阳性。序列分析发现该地区HEV人株与猪株在ORF2部分区域的核苷酸序列同源性为87％～98．1％。7株患者的戊肝病毒基因型和1株猪的戊肝病毒基因型均为Ⅳ型,基因序列同源性在87％～98．1％之间;其中有6例患者和猪的基因序列同源性在93．9％～98．1％之间,为Ⅳ型a亚型;1例患者和猪的基因序列同源性为87％,为Ⅳ型d亚型。正常人群的1例戊肝病毒基因型为Ⅰ型d亚型。该地区人与猪HEV的ORF2的部分基因片段与HEVⅠ～Ⅳ型的代表株进行比较,核苷酸序列同源性分别是82．5％～100％,81．7％～92．9％,81．4％～93．9％,84．9％～100％。结论该地区人群中流行的HEV存在2个基因型3个亚型,主要以基因Ⅳa型为主,与猪群中流行的HEV基因Ⅳa型同源性较高;HEVI型在人群中散在存在。     

广西眼镜王蛇毒杂合性酸性磷脂酶A2的基因克隆与序列分析

目的：研究广西眼镜王蛇毒杂合性酸性磷脂酶A2（APLA2）的基因克隆与序列分析。方法：从广西眼镜王蛇毒腺中提取总RNA，根据种属关系接近的台湾眼镜蛇毒APLA2两端非翻译区的保守序列设计引物，利用RT－PCR进行体外扩增，获得磷脂酶A2基因，克隆至pMD18-T载体，经测序筛选出APLA2－1，APLA2－2及一个新的酸性磷脂酶A2（命名为APLA2－3）。根据测序结果确定了APLA2－3基因的全序列，并由此推导编码的氨基酸序列。结果：利用Antheprot 4.3c蛋白质序列分析软件包计算它的等电点为4．885，相对分子质量为16．543kD，并预测了它的二级结构和部分理化性质，同源性比较表明，APLA2－3的1－39位氨基酸残基序列与APLA2－2相同，其同源性为64％，而40－108位氨基酸残基序列为APLA2－1相同，其同源性为69％，10位氨基酸残基序列皆不同于这两种APLA2。结论：揭示了APLA2具有基因多样性，可能与物种进化和生物活性有关，并为进一步研究PLA2的结构与功能的关系提供更多的信息。     

Lack of a functional plastid tRNACys gene is associated with loss of photosynthesis in a lineage of parasitic plants

Summary We recently reported that the gene for chloroplast tRNA^Cys(GCA) is a pseudogene in the plastid DNA of Epifagus virginiana, a non-photosynthetic parastic flowering plant in the family Orobanchaceae. Since this is the only tRNA^Cys gene in the plastid genome, and since Epifagus appears to possess a functional plastid translational apparatus, it seems probable that nuclear-encoded tRNAs are imported into plastids to effect translation. In this study we have surveyed species closely related to Epifagus to establish how widespread the loss of this tRNA gene has been. We find that Conopholis americana, another non-photosynthetic parasite, lacks the gene altogether, but that seven closely-related photosythetic plants (both parasitic and free-living) maintain an intact chloroplast tRNA^Cys gene. Thus, the tRNA^Cys gene appears to have become non-functional at the same time that photosynthetic ability was lost. This may be because the levels of putatively imported tRNAs are sufficient to meet the demands of plastid gene expression under nonphotosynthetic conditions only.     

The virulence-determining genomic BamHI fragment 4 of pseudorabies virus contains genes corresponding to the UL15 (partial), UL18, UL19, UL20, and UL21 genes of herpes simplex virus and a putative origin of replication.

The genomic BamHI-fragment 4 of pseudorabies virus (PrV) has previously been shown to encode functions necessary for expression of PrV neurovirulence (B. Lomniczi, S. Watanabe, T. Ben-Porat, and A. S. Kaplan, 1984, J. Virol. 52, 198-205). To identify proteins that might be involved in the neurotropism of PrV we sequenced the complete 9382-bp fragment BamHI-4, the longest contiguous sequence determined in the UL region of PrV so far, and analyzed its coding capacity. In an arrangement similar to that found in herpes simplex virus type 1 we identified complete open reading frames encoding proteins with strong homology to the UL18 (50% homology), UL19 (60% homology), UL20 (33% homology), and UL21 (36% homology) polypeptides and the 3'-part of a gene homologous to UL15 (67% homology) of HSV-1. In addition, a consensus sequence for an alphaherpesviral origin of replication was found at the left terminus of the fragment.     

Identification and characterization of human FHOD3 gene in silico

Formin homology proteins are actin regulators with scaffold function, which are implicated in organogenesis, normal tissue homeostasis, and cancer-cell invasion. FHOD1/FHOS, GRID2IP, Fmn1 and Fmn2 are non-FDD-type Formin homology proteins, while FMNL1, FMNL2/FHOD2, FMNL3, DAAM1, DAAM2, DIAPH1, DIAPH2 and DIAPH3 are FDD-type Formin homology proteins. Here, we identified and characterized FHOD3 (also known as FHOS2), a novel gene homologous to FHOD1, by using bioinformatics. Because FLJ46173, FLJ22297, KIAA1695 and FLJ34580 were partial FHOD3 cDNAs, complete coding sequence of FHOD3 cDNA was determined by assembling nucleotide sequences of FLJ46173 and FLJ22297. FHOD3 gene at human chromosome 18q12.2 was found consisting of at least 25 exons. Exon 11 of FHOD3 gene was spliced out in KIAA1695 cDNA and BF116064 EST, while exon 13 of FHOD3 gene was spliced out in FLJ46173 cDNA. FHOD3 gene encodes at least three isoforms due to alternative splicing of the exon skipping type. FHOD3 and FHOD1 showed 52.1% total-amino-acid identity. Drosophila CG32030 showed 43.9% total-amino-acid identity with human FHOD3, and 39.1% total-amino-acid identity with human FHOD1. FHDHN domain (codon 1-327 of FHOD3) and FHDHC domain (codon 1377-1421 of FHOD3) were identified as the N-terminal conserved region and the juxta C-terminal conserved region, respectively. Human FHOD3, FHOD1 and Drosophila CG32030 were found to share the conserved domain structure consisting of FHDHN, FH1, FH2, and FHDHC domains. This is the first report on the FHOD3 gene as well as on the novel FHDHN and FHDHC domains.     

Potential gene products of bean golden mosaic virus have higher sequence homologies to those of tomato golden mosaic virus than to those of cassava latent virus

Comparison of the nucleotide sequences of the DNAs of bean golden mosaic virus (BGMV), tomato golden mosaic virus (TGMV) and cassava latent virus (CLV) revealed a fairly close relationship between BGMV DNA1, TGMV DNA1, and CLV DNA1 and a comparatively distant relationship between BGMV DNA2, TGMV DNA2, and CLV DNA2. The 200-base region common to the two DNAs of each virus had little sequence homology, except for a highly conserved 33–36 base sequence potentially capable of forming a stable hairpin structure. All the potential coding regions in the BGMV DNAs had counterparts in the TGMV and CLV DNAs suggesting an overall similarity in genome organization but two potential coding regions in the BGMV DNAs had no counterparts in the TGMV DNAs. The most highly conserved ORFs, BGMV 1R1, TGMV 1R1, and CLV 1R1, are the putative genes for the coat proteins of BGMV, TGMV, and CLV. BGMV 1R1 has 91.9% and 71.6% homology with respect to TGMV and CLV. The ORFs (BGMV 1L1; CLV 1L1; TGMV 1L1) and the two smaller overlapping ORFs (BGMV 1L2, 1L3; TGMV 1L2, 1L3; CLV 1L5, 1L3) are conserved in the three viruses. BGMV 2R1 and BGMV 2L1 have higher homology with respect to TGMV but not with respect to 2R1 and 2L1 in CLV. From this study we conclude that BGMV is more closely related to TGMV than CLV.     

Expression of an inducible nitric oxide synthase gene in rainbow trout Oncorhynchus mykiss

Using oligonucleotide primers based on mammalian nitric oxide synthases (NOS), expression of an inducible NOS (iNOS) gene was detected in head kidney and gill tissue of bacterially-challenged rainbow trout. Three overlapping fragments were amplified by RT-PCR and used to construct a contiguous sequence of 1410bp, with high nucleotide homology to iNOS in birds (61%) and mammals (57-59%). The nucleotide sequence translated in one reading frame to produce a partial peptide containing 470 amino acids, with 69-71% amino acid homology with mammalian iNOS, 81% homology with chicken iNOS and 85% homology with a partial (492bp) goldfish iNOS sequence. In vitro stimulation of head kidney macrophages with LPS also induced expression of the trout iNOS RNA, with optimal expression seen using 20-50 microg/ml LPS at 2h to 6h post-stimulation. The evolutionary and functional significance of the trout iNOS sequence are discussed.     

Molecular cloning of a ligand for the inducible T cell gene 4-1BB: a member of an emerging family of cytokines with homology to tumor necrosis factor

4-1BB is an inducible T cell antigen that shows sequence homology to members of an emerging family of cytokine receptors, including those for tumor necrosis factor and nerve growth factor. To aid in the analysis of the function of 4-1BB we have utilized a soluble form of the molecule as a probe to identify and clone the gene which encodes its ligand. The ligand for 4-1BB is a type II membrane glycoprotein that has homology to tumor necrosis factor, lymphotoxin, and the ligands for CD40 and CD27, all of which are themselves ligands to receptors in this superfamily. The gene for 4-IBB is on mouse chromosome 4 and maps close to the p80 form of the tumor necrosis factor receptor as well as the gene for CD30. The gene for 4-IBB ligand maps to mouse chromosome 17, but considerably distal to the tumor necrosis factor and lymphotoxin genes. Interaction of 4-1BB with its ligand induces the proliferation of activated thymocytes and splenic T cells, a response which is mimicked on similar cell populations stimulated with an antibody to 4-1BB.