共查询到20条相似文献,搜索用时 15 毫秒
1.
Isolation and molecular characterization of the Arabidopsis TPS1 gene, encoding trehalose-6-phosphate synthase 总被引:1,自引:0,他引:1
MA Blázquez E Santos CL Flores JM Martínez-Zapater J Salinas C Gancedo 《Canadian Metallurgical Quarterly》1998,13(5):685-689
An Arabidopsis thaliana cDNA clone, AtTPS1, that encodes a trehalose-6-phosphate synthase was isolated. The identity of this protein is supported by both structural and functional evidence. On one hand, the predicted sequence of the protein encoded by AtTPS1 showed a high degree of similarity with trehalose-6-phosphate synthases of different organisms. On the other hand, expression of the AtTPS1 cDNA in the yeast tps1 mutant restored its ability to synthesize trehalose and suppressed its growth defect related to the lack of trehalose-6-phosphate. Genomic organization and expression analyses suggest that AtTPS1 is a single-copy gene and is expressed constitutively at very low levels. 相似文献
2.
Glutamine:fructose-6-phosphate amidotransferase (glucosamine-6-phosphate synthase) catalyzes the first step of the hexosamine pathway required for the biosynthesis of cell wall precursors. The Candida albicans GFA1 gene was cloned by complementing a gfa1 mutation of Saccharomyces cerevisiae (previously known as gcn1-1; W. L. Whelan and C. E. Ballou, J. Bacteriol. 124:1545-1557, 1975). GFA1 encodes a predicted protein of 713 amino acids and is homologous to the corresponding gene from S. cerevisiae (72% identity at the nucleotide sequence level) as well as to the genes encoding glucosamine-6-phosphate synthases in bacteria and vertebrates. In cell extracts, the C. albicans enzyme was 4-fold more sensitive than the S. cerevisiae enzyme to UDP-N-acetylglucosamine (an inhibitor of the mammalian enzyme) and 2.5-fold more sensitive to N3-(4-methoxyfumaroyl)-L-2,3-diaminopropanoic acid (a glutamine analog and specific inhibitor of glucosamine-6-phosphate synthase). Cell extracts from the S. cerevisiae gfa1 strain transformed with the C. albicans GFA1 gene exhibited sensitivities to glucosamine-6-phosphate synthase inhibitors that were similar to those shown by the C. albicans enzyme. Southern hybridization indicated that a single GFA1 locus exists in the C. albicans genome. Quantitative Northern (RNA) analysis showed that the expression of GFA1 in C. albicans is regulated during growth: maximum mRNA levels were detected during early log phase. GFA1 mRNA levels increased following induction of the yeast-to-hyphal-form transition, but this was a response to fresh medium rather than to the morphological change. 相似文献
3.
Hyphal growth of Candida albicans was observed when yeast was cultured at 27 degrees C in liquid media containing 1% Tripcasine and 1.8% cyclodextrins (alpha, beta, and gamma respectively). Tripcasine as the sole nitrogen source did not induce the formation of hyphae of C. albicans, but cyclodextrins, especially CD-beta, were able to induce yeast-mycelial transition. In the TCD-beta media 25-30% septate hyphae form was observed. This study indicates the existence of an uptake system for CDs in C. albicans, provided these compounds are linearised in the medium. The CDs are inducers of hyphae and they may enter the C. albicans cells as linear oligosaccharids. 相似文献
4.
The physiological role of dihydroxyacetone synthase (DHAS) in Candida boidinii was evaluated at the molecular level. The DAS1 gene, encoding DHAS, was cloned from the host genome, and regulation of its expression by various carbon and nitrogen sources was analyzed. Western and Northern analyses revealed that DAS1 expression was regulated mainly at the mRNA level. The regulatory pattern of DHAS was similar to that of alcohol oxidase but distinct from that of two other enzymes in the formaldehyde dissimilation pathway, glutathione-dependent formaldehyde dehydrogenase and formate dehydrogenase. The DAS1 gene was disrupted in one step in the host genome (das1Delta strain), and the growth of the das1Delta strain in various carbon and nitrogen sources was compared with that of the wild-type strain. The das1Delta strain had completely lost the ability to grow on methanol, while the strain with a disruption of the formate dehydrogenase gene could survive (Y. Sakai et al., J. Bacteriol. 179:4480-4485, 1997). These and other experiments (e.g., those to determine the expression of the gene and the growth ability of the das1Delta strain on media containing methylamine or choline as a nitrogen source) suggested that DAS1 is involved in assimilation rather than dissimilation or detoxification of formaldehyde in the cells. 相似文献
5.
MW Loewus 《Canadian Metallurgical Quarterly》1977,252(20):7221-7223
myo-Inositol-1-phosphate synthase (EC 5.5.1.4) from rat testis, Acer pseudoplatanus L. cell culture and Oryza sativa L. cell culture, converted D-[5-3H]glucose 6-phosphate to myo-[2-3H]inositol 1-phosphate at rates ranging from 0.21 to 0.48 that of unlabeled substrate. D-[3-3H]- and D-[4-3H]glucose 6-phosphate were converted at approximately the same rate as that of unlabeled substrate. In the case of testis enzyme, storage as a frozen solution further lowered the rate with D-[5-3H]glucose 6-phosphate as substrate. When the reaction was run in [3H]water, no 3H appeared in myo-inositol 1-phosphate but a small amount was recovered in substrate isolated from the final reaction mixture. These data support the involvement of carbon 5 of D-glucose 6-phosphate in the mechanism proposed for this conversion. 相似文献
6.
7.
8.
The gene encoding ornithine decarboxylase, SPE1, from the pathogenic yeast Candida albicans has been isolated by complementation of an ornithine decarboxylase-negative (spe1 delta) strain of Saccharomyces cerevisiae. Four transformants, three of which contain plasmids with the SPE1 gene, were isolated by selection on polyamine-free medium. The C. albicans ornithine decarboxylase (ODC) showed high homology with other eukaryotic ODCs at both the amino acid and nucleic acid levels. 相似文献
9.
A previously isolated partial cDNA encoding a cell wall protein antigen found on hyphal surfaces of the opportunistic fungal pathogen, Candida albicans (Staab et al., 1996) was used to clone the complete hyphal wall protein 1 gene (HWP1). Hyphal forms of C. albicans invade mucosal surfaces of immunocompromised patients such as those with AIDS. HWP1 consisted of an open reading frame predicting an acidic protein (pI of 3.37) with a calculated molecular size of 61,122. The antigenic domain was located in the N-terminal third of the protein. The remainder of the protein contained abundant hydroxy amino acids, and terminated with a string of 15 amino acids typical of sequences specifying post-translational modification with glycosylphosphatidylinositol (6PI). The analyses suggested that Hwp1 is a glucan-linked protein with serine/threonine-rich regions that are predicted to function in extending a ligand-binding domain into the extracellular space. 相似文献
10.
PJ Blackshear DJ Stumpo E Carballo JC Lawrence 《Canadian Metallurgical Quarterly》1997,272(50):31510-31514
PHAS-I is the prototype of a group of eIF4E-binding proteins that can regulate mRNA translation in response to hormones and growth factors. To investigate the importance of PHAS-I in the physiology of the intact animal, we disrupted the PHAS-I gene in mice. Tissues and cells derived from the knockout mice contained no detectable PHAS-I protein. A related protein, PHAS-II, and eIF4E were readily detectable in tissues from these animals, but neither appeared to be changed in a compensatory manner. Mice lacking PHAS-I appeared normal at birth. However, male knockout mice weighed approximately 10% less than controls at all ages, whereas female weights were similar to those of controls. Both males and females were fertile. Tissues from adult animals appeared to be normal by routine histological staining techniques, as were routine blood cell counts and chemistries. Fibroblasts derived from PHAS-I-deficient mouse embryos exhibited normal rates of growth and overall protein synthesis, responded normally to serum stimulation of ornithine decarboxylase activity and cell growth, and rapamycin inhibition of cell growth. Under these experimental conditions, PHAS-I is apparently not required for the normal development and reproductive behavior of female mice, but is required for normal body weight in male mice; the mechanisms responsible for this phenotype remain to be determined. 相似文献
11.
A cosmid carrying the orlA gene from Aspergillus nidulans was identified by complementation of an orlA1 mutant strain with DNA from the pKBY2 cosmid library. An orlA1 complementing fragment from the cosmid was sequenced. orlA encodes a predicted polypeptide of 227 amino acids (26360 Da) that is homologous to a 211-amino-acid domain from the polypeptide encoded by the Saccharomyces cerevisiae TPS2 gene and to almost the entire Escherichia coli otsB-encoded polypeptide. TPS2 and otsB each specify a trehalose-6-phosphate phosphatase, an enzyme that is necessary for trehalose synthesis. orlA disruptants accumulate trehalose-6-phosphate and have reduced trehalose-6-phosphatate phosphatase levels, indicating that the gene encodes a trehalose-6-phosphatate phosphatase. Disruptants have a nearly-wild-type morphology at 32 degrees C. When germinated at 42 degrees C, the conidia and hyphae from disruptants are chitin deficient, swell excessively, and lyse. The lysis is almost completely remedied by osmotic stabilizers and is partially remedied by N-acetylglucosamine (GlcNAc). The activity of glutamine:fructose-6-phosphate amido-transferase (GFAT), the first enzyme unique to aminosugar synthesis, is reduced and is labile in orlA disruption strains. The findings are consistent with the hypothesis that trehalose-6-phosphate reduces the temperature stability of GFAT and other enzymes of chitin metabolism at elevated temperatures. The results extend to filamentous organisms the observation that mutations in fungal trehalose synthesis are highly pleiotropic and affect aspects of carbohydrate metabolism that are not directly related to trehalose synthesis. 相似文献
12.
An experiment was conducted to compare the functional performance of younger and older adults on familiar and unfamiliar tasks under 2 conditions of perceived control. Specifically, the relation between age and motor and process skills was examined. The familiar tasks were simple cooking tasks, whereas the unfamiliar tasks were contrived, meaningless tasks developed for this study. Younger and older adults did not differ in the ratings of the familiarity of the tasks, but results from 2 Age x Task x Choice analyses of variance demonstrated a significant age difference for motor and process skills under all conditions. This suggests that older adults demonstrate age-related decline, even with activities that take motivational, experiential, and ecological validity components into account. For the process skills scale, there was also a significant main effect for choice. These results support the concept that perceived control may improve performance, but not differentially for older adults; that is, younger and older adults both demonstrated improved process performance when given their choice of tasks. 相似文献
13.
SD Leidich AS Ibrahim Y Fu A Koul C Jessup J Vitullo W Fonzi F Mirbod S Nakashima Y Nozawa MA Ghannoum 《Canadian Metallurgical Quarterly》1998,273(40):26078-26086
The Candida albicans PLB1 gene was cloned using a polymerase chain reaction-based approach relying on degenerate oligonucleotide primers designed according to the amino acid sequences of two peptide fragments obtained from a purified candidal enzyme displaying phospholipase activity (Mirbod, F., Banno, Y., Ghannoum, M. A., Ibrahim, A. S., Nakashima, S., Yasuo, K., Cole, G. T., and Nozawa, Y. (1995) Biochim. Biophys. Acta 1257, 181-188). Sequence analysis of a 6.7-kilobase pair EcoRI-ClaI genomic clone revealed a single open reading frame of 1818 base pairs that predicts for a pre-protein of 605 residues. Comparison of the putative candidal phospholipase with those of other proteins in data base revealed significant homology to known fungal phospholipase Bs from Saccharomyces cerevisiae (45%), Penicillium notatum (42%), Torulaspora delbrueckii (48%), and Schizosaccharomyces pombe (38%). Thus, we have cloned the gene encoding a C. albicans phospholipase B homolog. This gene, designated caPLB1, was mapped to chromosome 6. Disruption experiments revealed that the caplb1 null mutant is viable and displays no obvious phenotype. However, the virulence of strains deleted for caPLB1, as assessed in a murine model for hematogenously disseminated candidiasis, was significantly attenuated compared with the isogenic wild-type parental strain. Although deletion of caPLB1 did not produce any detectable effects on candidal adherence to human endothelial or epithelial cells, the ability of the caplb1 null mutant to penetrate host cells was dramatically reduced. Thus, phospholipase B may well contribute to the pathogenicity of C. albicans by abetting the fungus in damaging and traversing host cell membranes, processes which likely increase the rapidity of disseminated infection. 相似文献
14.
Squamous cell carcinoma (SCC) antigen (Ag) present and expressed in normal epithelium and epithelial tissues is used primarily as tumor marker of SCC of the uterine cervix. In this review, we considered factors interfering in vitro with the collection of samples and assay procedures, benign and malignant nongynecological diseases which may be the cause of elevated serum levels of SCC Ag. Contamination with skin or saliva strongly influences SCC Ag levels. SCC Ag is elevated in several benign lesions, including pulmonary (tuberculosis, adult respiratory distress syndrome, pulmonary infiltration with eosinophilia, sarcoidosis, bronchogenic cyst) and skin (eczema, pemphigus, erythroderma epidermitis, psoriasis) diseases. Elevations are observed in SCC malignancies of the head and neck, esophagus, skin, lung, urothelium, anal canal and vulva. 相似文献
15.
16.
Elevated blood glucose concentrations result in increased intracellular levels of glucose 6-phosphate in liver, skeletal muscle, and adipose tissue. In liver, blood glucose concentrations are the main factor in control of the synthesis of glycogen; insulin has only a potentiating effect. In skeletal muscle and adipocytes, glucose alone has little effect on the activity of glycogen synthase, the limiting enzyme in glycogen synthesis. However, insulin released as a result of elevated blood glucose stimulates the translocation of specific glucose transporters to the cell membrane, increases the uptake of glucose, and causes the covalent, dephosphorylation-mediated activation of glycogen synthase. We present evidence that elevated intracellular contents of glucose 6-phosphate provoke the activation of glycogen synthase in liver, muscle, and adipose tissue. In addition, glucose 6-phosphate may inhibit the phosphorylation of glycogen synthase by cyclic AMP-stimulated protein kinase. We show that the stimulated glucose uptake and phosphorylation appear to play a major role in the control by insulin of the enzymes involved in glycogen synthesis. 相似文献
17.
We have cloned and analysed the sequence of a putative histidine kinase, two-component gene (CaHK1) from Candida albicans. This gene encodes a 2471 amino acid protein (Cahk1p) with an estimated molecular mass of 281.8 kDa. A homology search of Cahk1p with other proteins in the databases showed that Cahk1p exhibits the greatest homology at its C-terminus with both the sensor and regulator components of prokaryotic and eukaryotic two-component histidine kinases. A further analysis of this homology showed that the Cahk1p possessed both sensor and regulator domains in the same polypeptide. Also, Cahk1p is likely to be a soluble protein. The sensor kinase domain of Cahk1p contains conserved motifs that are characteristic of all histidine kinase proteins, including the putative histidine which is believed to be autophosphorylated during activation, ATP binding motifs and others (F- and N-motifs), with unknown function. The Cahk1p regulator domain also contains conserved aspartate and lysine residues and the putative aspartate, which is secondarily phosphorylated by the autophosphorylated histidine. Finally, according to the codon usage frequency of the CaHK1 gene in comparison with other genes from C. albicans, there would appear to be a low level of expression of the gene. 相似文献
18.
We have isolated CaKRE9, a gene from Candida albicans, that is a functional homologue of the Saccharomyces cerevisiae KRE9 gene involved in beta-1,6-glucan synthesis. Disruption of the CaKRE9 gene in C. albicans shows that CaKre9p is required for the synthesis or assembly of this fungal polymer. Homozygous null disruptants of CaKRE9 grow poorly on galactose and fail to form hyphae in serum, and, in growth medium containing glucose, the gene is essential. Thus, the CaKRE9 gene product is a potentially useful candidate as a target for fungal-specific drugs. 相似文献
19.
F De Bernardis FA Mühlschlegel A Cassone WA Fonzi 《Canadian Metallurgical Quarterly》1998,66(7):3317-3325
Little is known of the biological attributes conferring pathogenicity on the opportunistic fungal pathogen Candida albicans. Infection by this pathogen, as for bacterial pathogens, may rely upon environmental signals within the host niche to regulate the expression of virulence determinants. To determine if C. albicans responds to the pH of the host niche, we tested the virulence of strains with mutations in either of two pH-regulated genes, PHR1 and PHR2. In vitro, PHR1 is expressed when the ambient pH is at 5.5 or higher and deletion of the gene results in growth and morphological defects at neutral to alkaline pHs. Conversely, PHR2 is expressed at an ambient pH below 5.5, and the growth and morphology of the null mutant is compromised below this pH. A PHR1 null mutant was avirulent in a mouse model of systemic infection but uncompromised in its ability to cause vaginal infection in rats. Since systemic pH is near neutrality and vaginal pH is around 4.5, the virulence phenotype paralleled the pH dependence of the in vitro phenotypes. The virulence phenotype of a PHR2 null mutant was the inverse. The mutant was virulent in a systemic-infection model but avirulent in a vaginal-infection model. Heterozygous mutants exhibited partial reductions in their pathogenic potential, suggesting a gene dosage effect. Unexpectedly, deletion of PHR2 did not prevent hyphal development in vaginal tissue, suggesting that it is not essential for hyphal development in this host niche. The results suggest that the pH of the infection site regulates the expression of genes essential to survival within that niche. This implies that the study of environmentally regulated genes may provide a rationale for understanding the pathobiology of C. albicans. 相似文献
20.
Virulence and hyphal formation of Candida albicans require the Ste20p-like protein kinase CaCla4p 总被引:1,自引:0,他引:1
E Leberer K Ziegelbauer A Schmidt D Harcus D Dignard J Ash L Johnson DY Thomas 《Canadian Metallurgical Quarterly》1997,7(8):539-546
BACKGROUND: The pathogenic fungus Candida albicans is capable of a morphological transition from a unicellular budding yeast to a filamentous form. Extensive filamentous growth leads to the formation of mycelia displaying hyphae with branches and lateral buds. Hyphae have been observed to adhere to and invade host tissues more readily than the yeast form, suggesting that filamentous growth may contribute to the virulence of this major human pathogen. A molecular and genetic understanding of the potential role of morphological switching in the pathogenicity of C. albicans would be of significant benefit in view of the increasing incidence of candidiasis. RESULTS: The CaCLA4 gene of C. albicans was cloned by functional complementation of the growth defect of cells of the budding yeast Saccharomyces cerevisiae deleted for the STE20 gene and the CLA4 gene. CaCLA4 encodes a member of the Ste20p family of serine/threonine protein kinases and is characterized by a pleckstrin homology domain and a Cdc42p-binding domain in its amino-terminal non-catalytic region. Deletion of both alleles of CaCLA4 in C. albicans caused defects in hyphal formation in vitro, in both synthetic liquid and solid media, and in vivo in a mouse model for systemic candidiasis. The gene deletions reduced colonization of the kidneys in infected mice and suppressed C. albicans virulence in the mouse model. CONCLUSIONS: Our results demonstrate that the function of the CaCla4p protein kinase is essential for virulence and morphological switching of C. albicans in a mouse model. Thus, hyphal formation of C. albicans mediated by CaCla4p may contribute to the pathogenicity of this dimorphic fungus, suggesting that regulators of morphological switching may be useful targets for antifungal drugs. 相似文献