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At least 1,099 records · Page 61Linked to original sources
PubMed2004

Y chromosomal haplogroup J as a signature of the post-neolithic colonization of Europe.

In order to attain a finer reconstruction of the peopling of southern and central-eastern Europe from the Levant, we determined the frequencies of eight lineages internal to the Y chromosomal haplogroup J, defined by biallelic markers, in 22 population samples obtained with a fine-grained sampling scheme. Our results partially resolve a major multifurcation of lineages within the haplogroup. Analyses of molecular variance show that the area covered by haplogroup J dispersal is characterized by a significant degree of molecular radiation for unique event polymorphisms within the haplogroup, with a higher incidence of the most derived sub-haplogroups on the northern Mediterranean coast, from Turkey westward; here, J diversity is not simply a subset of that present in the area in which this haplogroup first originated. Dating estimates, based on simple tandem repeat loci (STR) diversity within each lineage, confirmed the presence of a major population structuring at the time of spread of haplogroup J in Europe and a punctuation in the peopling of this continent in the post-Neolithic, compatible with the expansion of the Greek world. We also present here, for the first time, a novel method for comparative dating of lineages, free of assumptions of STR mutation rates.

Africa, Northern↗
PubMed1999

Identification and molecular characterization of a novel Salmonella enteritidis pathogenicity islet encoding an ABC transporter.

Using a newly constructed minitransposon with a phoA reporter gene in a Salmonella enteritidis phoN mutant, we have identified an iron- and pH-inducible lipoprotein gene sfbA, which is a component of a novel ABC-type transporter system required for virulence. This gene is located on a 4 kb Salmonella-specific chromosomal segment, which constitutes a new pathogenicity islet. This islet encodes an outer membrane protein, OmpX, and contains the operon designated sfbABC (Salmonella ferric binding) encoding a putative periplasmic iron-binding lipoprotein SfbA, a nucleotide-binding ATPase SfbB and a cytoplasmic permease SfbC, as predicted by their characteristic signature sequences. Inactivation of the sfbA gene resulted in a mutant that is avirulent and induces protective immunity in BALB/c mice. The wild-type phenotype could be restored by in vivo complementation with the sfbABC operon. This novel transporter might be involved in iron uptake in Salmonella.

ATP-Binding Cassette Transporters↗
PubMed2004

Imparting temperature sensitivity and attenuation in ferrets to A/Puerto Rico/8/34 influenza virus by transferring the genetic signature for temperature sensitivity from cold-adapted A/Ann Arbor/6/60.

The four temperature-sensitive (ts) loci identified in the PB1 and PB2 gene segments of cold-adapted A/Ann Arbor/6/60 influenza virus, the master donor virus for influenza A virus (MDV-A) FluMist vaccines, were introduced into a divergent A/Puerto Rico/8/34 influenza virus strain. Recombinant A/Puerto Rico/8/34 virus with these four introduced ts loci exhibited both ts and att phenotypes similar to those of MDV-A, which could be used as a donor virus for manufacturing large quantities of inactivated influenza virus vaccine against potential pandemic strains.

Animals↗
PubMed2005

Histologic type, organ of origin, and Wnt pathway status: effect on gene expression in ovarian and uterine carcinomas.

PURPOSE: Ovarian and uterine carcinomas manifest several differentiation patterns resembling those seen in nonneoplastic epithelia of the gynecologic tract. Specific oncogene and tumor suppressor gene defects have been associated with particular differentiation patterns in carcinomas arising in either the uterus or ovary. For instance, ovarian and uterine carcinomas with endometrioid differentiation frequently show beta-catenin mutations. Whereas type of differentiation is considered in the treatment of uterine carcinomas, it does not presently contribute to decisions about treatment of ovarian carcinomas. A widely accepted view is that the accumulation of specific gene defects and gene expression changes underlies phenotypic traits of cancers, including their response to treatment. EXPERIMENTAL DESIGN: Using oligonucleotide microarrays to assess gene expression in 103 primary ovarian and uterine carcinomas, we sought to address whether organ of origin or type of differentiation (histotype; endometrioid versus serous) had a more substantial effect on gene expression patterns. RESULTS: We found that effects on gene expression due to organ of origin and histotype are similar in magnitude and are parallel in that organ effects are similar in the two histotypes and histotype effects are similar in the two organs. In addition, ovarian and uterine endometrioid adenocarcinomas with beta-catenin defects show a common gene expression signature largely distinct from that seen in tumors lacking such defects. CONCLUSIONS: Our results illustrate how organ of origin, type of differentiation, and specific molecular defects all contribute to gene expression in the most common types of ovarian and uterine cancers. The findings also imply gene expression data will be of value for stratifying ovarian cancer patients for new treatment approaches.

Cell Differentiation↗
PubMed2002

Large-scale identification of serotype 4 Streptococcus pneumoniae virulence factors.

Streptococcus pneumoniae (the pneumococcus) is carried in the nasopharynx of healthy individuals, but can spread to other host sites and lead to pneumonia, bacteraemia, otitis media and meningitis. Although it is logical to think a priori that differential gene expression would contribute to the ability of this pathogen to colonize different sites, in fact very few genes have been demonstrated to play tissue-specific roles in virulence or carriage. Using signature-tagged mutagenesis to screen 6149 mariner-transposon insertion strains, we identified 387 mutants attenuated for infection in a murine model of pneumonia. Among these mutants are ones with disruptions in a number of putative tissue-specific transcriptional regulators, surface proteins, metabolic proteins and proteins of unknown function, most of which had not previously been associated with virulence. A subset of these, including most of those with insertions in putative transcriptional regulators,was examined for phenotypes in murine models of bacteraemia and nasopharyngeal carriage. Four classes of mutants defective in infection models of the: (I) lung, (II) lung and blood, (III) lung and nasopharynx,and (IV) all three tissues were identified, thus demonstrating the existence of tissue-specific pneumococcal virulence factors. Included in these strains were two with disruptions in a genetic locus that putatively codes for a transcriptional regulator, three surface proteins and three sortase homologues. Mutation analysis revealed that three of the seven genes in this locus are virulence factors that are specific to mucosal surfaces.

Amino Acid Sequence↗
PubMed2004

Identification of Mycobacterium marinum virulence genes using signature-tagged mutagenesis and the goldfish model of mycobacterial pathogenesis.

Mycobacterium marinum, a causative agent of fish tuberculosis, is one of the most closely related Mycobacterium species (outside the M. tuberculosis complex) to M. tuberculosis, the etiologic agent of human tuberculosis. Signature-tagged mutagenesis was used to identify genes of M. marinum required for in vivo survival in a goldfish model of mycobacterial pathogenesis. Screening the first 1008 M. marinum mutants led to the identification of 40 putative virulence mutants. DNA sequence analysis of these 40 mutants identified transposon insertions in 35 unique loci. Twenty-eight out of 33 (85%) loci encoding putative virulence genes have homologous genes in M. tuberculosis.

Animals↗
PubMed2004

Polycystin-1L2 is a novel G-protein-binding protein.

Mutations in genes encoding polycystin-1 (PC1) and polycystin-2 cause autosomal dominant polycystic kidney disease. The polycystin protein family is composed of Ca2+-permeable pore-forming subunits and receptor-like integral membrane proteins. Here we describe a novel member of the polycystin-1-like subfamily, polycystin-1L2 (PC1L2), encoded by PKD1L2, which has various alternative splicing forms with two translation initiation sites. PC1L2 short form starts in exon 12 of the long form. The longest open reading frame of PKD1L2 short form, determined from human testis cDNA, encodes a 1775-amino-acid protein and 32 exons, whereas the long form is predicted to encode a 2460-residue protein. Both forms have a small receptor for egg jelly domain, a G-protein-coupled receptor proteolytic site, an LH2/PLAT, and 11 putative transmembrane domains, as well as a number of rhodopsin-like G-protein-coupled receptor signatures. RT-PCR analysis shows that the short form, but not the long form, of human PKD1L2 is expressed in the developing and adult heart and kidney. Furthermore, by GST pull-down assay we observed that PC1L2 and polycystin-1L1 are able to bind to specific G-protein subunits. We also show that PC1 C-terminal cytosolic domain binds to Galpha12, Galphas, and Galphai1, while it weakly interacts with Galphai2. Our results indicate that both PC1-like molecules may act as G-protein-coupled receptors.

Alternative Splicing↗
PubMed2002

Sequence analysis of bacteriophage T4 DNA packaging/terminase genes 16 and 17 reveals a common ATPase center in the large subunit of viral terminases.

Phage DNA packaging is believed to be driven by a rotary device coupled to an ATPase 'motor'. Recent evidence suggests that the phage DNA packaging motor is one of the strongest force-generating molecular motors reported to date. However, the ATPase center that is responsible for generating this force is unknown. In order to identify the DNA translocating ATPase, the sequences of the packaging/terminase genes of coliphages T4 and RB49 and vibriophages KVP40 and KVP20 have been analyzed. Alignment of the terminase polypeptide sequences revealed a number of functional signatures in the terminase genes 16 and 17. Most importantly, the data provide compelling evidence for an ATPase catalytic center in the N-terminal half of the large terminase subunit gp17. An analogous ATPase domain consisting of conserved functional signatures is also identified in the large terminase subunit of other bacteriophages and herpesviruses. Interestingly, the putative terminase ATPase domain exhibits some of the common features found in the ATPase domain of DEAD box helicases. Residues that would be critical for ATPase catalysis and its coupling to DNA packaging are identified. Com binatorial mutagenesis shows that the predicted threonine residues in the putative ATPase coupling motif are indeed critical for function.

Adenosine Triphosphatases↗
PubMed2003

Leaf vitamin C contents modulate plant defense transcripts and regulate genes that control development through hormone signaling.

Vitamin C deficiency in the Arabidopsis mutant vtc1 causes slow growth and late flowering. This is not attributable to changes in photosynthesis or increased oxidative stress. We have used the vtc1 mutant to provide a molecular signature for vitamin C deficiency in plants. Using statistical analysis, we show that 171 genes are expressed differentially in vtc1 compared with the wild type. Many defense genes are activated, particularly those that encode pathogenesis-related proteins. Furthermore, transcript changes indicate that growth and development are constrained in vtc1 by the modulation of abscisic acid signaling. Abscisic acid contents are significantly higher in vtc1 than in the wild type. Key features of the molecular signature of ascorbate deficiency can be reversed by incubating vtc1 leaf discs in ascorbate. This finding provides evidence that many of the observed effects on transcript abundance in vtc1 result from ascorbate deficiency. Hence, through modifying gene expression, vitamin C contents not only act to regulate defense and survival but also act via phytohormones to modulate plant growth under optimal conditions.

Abscisic Acid↗
PubMed1995

Proteus mirabilis urease: operon fusion and linker insertion analysis of ure gene organization, regulation, and function.

Urease is an inducible virulence factor of uropathogenic Proteus mirabilis. Although eight contiguous genes necessary for urease activity have been cloned and sequenced, the transcriptional organization and regulation of specific genes within the Proteus gene cluster has not been investigated in detail. The first gene, ureR, is located 400 bp upstream and is oriented in the direction opposite the other seven genes, ureDABCEFG. The structural subunits of urease are encoded by ureABC. Previously, UreR was shown to contain a putative helix-turn-helix DNA-binding motif 30 residues upstream of a consensus sequence which is a signature for the AraC family of positive regulators; this polypeptide is homologous to other DNA-binding regulatory proteins. Nested deletions of ureR linked to either ureD-lacZ or ureA-lacZ operon fusions demonstrated that an intact ureR is required for urea-induced synthesis of LacZ from either ureA or ureD and identified a urea-regulated promoter in the ureR-ureD intergenic region. However, lacZ operon fusions to fragments encompassing putative promoter regions upstream of ureA and ureF demonstrated that no urea-regulated promoters occur upstream of these open reading frames; regions upstream of ureR, ureE, and ureG were not tested. These data suggest that UreR acts as a positive regulator in the presence of urea, activating transcription of urease structural and accessory genes via sequences upstream of ureD. To address the role of the nonstructural regulatory and accessory genes, we constructed deletion, cassette, and linker insertion mutations throughout the ure gene cluster and determined the effect of these mutations on production and regulation of urease activity in Escherichia coli. Mutations were obtained, with locations determine by DNA sequencing, in all genes except ureA and ureE. In each case, the mutation resulted in a urease-negative phenotype.

Bacterial Proteins↗
PubMed2002

The sequence, expression, and chromosomal localization of a novel polycystic kidney disease 1-like gene, PKD1L1, in human.

Polycystin-1 and polycystin-2 are the products of PKD1 and PKD2, genes that are mutated in most cases of autosomal dominant polycystic kidney disease. Since the first two polycystins were cloned, three new members, polycystin-L, -2L2, and -REJ, have been identified. In this study, we describe a sixth member of the family, polycystin-1L1, encoded by PKD1L1 in human. The full-length cDNA sequence of PKD1L1, determined from human testis cDNA, encodes a 2849-amino-acid protein and 58 exons in a 187-kb genomic region. The deduced amino acid sequence of polycystin-1L1 has significant homology with all known polycystins, but the longest stretches of homology were found with polycystin-1 and -REJ over the 1453- and 932-amino-acid residues, respectively. Polycystin-1L1 is predicted to have two Ig-like PKD, a REJ, a GPS, a LH2/PLAT, a coiled-coil, and 11 putative transmembrane domains. Several rhodopsin-like G-protein-coupled receptor (GPCR) signatures are also found in polycystin-1L1. Dot-blot analysis and RT-PCR revealed that human PKD1L1 is expressed in testis and in fetal and adult heart. In situ hybridization analysis showed that the most abundant and specific expression of Pkd1l1 was found in Leydig cells, a known source of testosterone production, in mouse testis. We have assigned PKD1L1 to the short arm of human chromosome 7 in bands p12--p13 and Pkd1l1 to mouse chromosome 11 in band A2 by fluorescence in situ hybridization. We hypothesize a role for polycystin-1L1 in the heart and in the male reproductive system.

Amino Acid Sequence↗
PubMed2003

Identification of a novel type IV pilus gene cluster required for gastrointestinal colonization of Citrobacter rodentium.

Citrobacter rodentium is used as an in vivo model system for clinically significant enteric pathogens such as enterohaemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC). These pathogens all colonize the lumen side of the host gastrointestinal tract via attaching and effacing (A/E) lesion formation. In order to identify genes required for the colonization of A/E-forming pathogens, a library of signature-tagged transposon mutants of C. rodentium was constructed and screened in mice. Of the 576 mutants tested, 14 were attenuated in their ability to colonize the descending colon. Of these, eight mapped to the locus of enterocyte effacement (LEE), which is required for the formation of A/E lesions, underlying the importance of this mechanism for pathogenesis. Another mutant, P5H2, was found to have a transposon insertion in an open reading frame that has strong similarity to type IV pilus nucleotide-binding proteins. The region flanking the transposon insertion was sequenced, identifying a cluster of 12 genes that encode the first described pilus of C. rodentium (named colonization factor Citrobacter, CFC). The proteins encoded by cfc genes have identity to proteins of the type IV COF pilus of enterotoxigenic E. coli (ETEC), the toxin co-regulated pilus of Vibrio cholerae and the bundle-forming pilus of EPEC. A non-polar mutation in cfcI, complementation of this strain with wild-type cfcI and complementation of strain P5H2 with wild-type cfcH confirmed that these genes are required for colonization of the gastrointestinal tract by C. rodentium. Thus, CFC provides a convenient model to study type IV pilus-mediated pathogen-host interactions under physiological conditions in the natural colonic environment.

Amino Acid Sequence↗
PubMed2006

GeneChips in stem cell research.

An understanding of the genes and signaling networks responsible for stem cell growth and differentiation will be essential for their ultimate therapeutic application. GeneChips are miniature platforms of nucleotides capable of monitoring the expression levels of almost every known and unknown gene. Performing a GeneChip experiment is like snapping a picture of a cell's mRNA (transcripts), thus giving a static view and measurement of gene expression inside the cell. Taking multiple "pictures" of stem cells as they grow and differentiate will provide insight into the genetic mechanisms of "stemness" or can be used to create "transcriptional signatures" to assess differentiation and variability between stem cell lines. The first half of this chapter covers the many components involved in a GeneChip experiment, illustrating the many variables at each step and describing a protocol for analysis that is inexpensive and requires minimal computer skills. The chapter then describes how researchers are currently applying GeneChips to stem cell biology. We conclude that the true potential of GeneChip technology lies in the in silico analysis-their integration and comparison of diverse data sets, where the biological questions are the driving force in the analysis.

Animals↗
PubMed1997

Bifunctional structure of two adenylyl cyclases from the myxobacterium Stigmatella aurantiaca.

Two adenylyl cyclase genes (cyaA and cyaB) from the myxobacterium Stigmatella aurantiaca were cloned by complementation of Escherichia coli mutants defective in the cya gene. cyaA codes for a protein of 424 amino acid residues (AC1), while cyaB encodes a protein of 352 residues (AC2). Both cyclases are sensitive to adenosine: cAMP production was strongly inhibited in E coli cells and cell extracts expressing these genes. AC1 comprises a hydrophobic domain of six transmembrane helices coupled to a cytoplasmic catalytic domain endowed with adenylyl cyclase activity. A 17 amino acid residue sequence, which is a signature of G-protein coupled receptors, as well as of slime mold Dictyostelium discoideum cyclic AMP receptors, was found in the membrane domain. AC2 displays features also indicating that it is a bifunctional enzyme. The domain located upstream from the catalytic adenylyl cyclase domain shows strong similarity to receiver modules of response regulators of two-component bacterial signaling systems. In vitro mutagenesis of conserved aspartate residues in this domain was shown to interfere with cAMP synthesis.

Adenosine↗
PubMed2001

Identification of bacterial genes required for in-vivo survival.

Genetic approaches used for in-vivo studies of bacterial pathogenesis are providing insights into how bacteria disrupt host defences and exploit host molecules for their own advantage. Signature tagged mutagenesis (STM) provides a means of identifying the genes involved in the process of infection, particularly those genes that are important for bacterial proliferation in-vivo. In this review, the application of STM to the understanding of bacterial pathogenesis and findings from work on three human pathogens, Salmonella typhimurium, Mycobacterium tuberculosis and Neisseria meningitidis, are discussed. The next challenge is to understand how these and other genes influence the infective process at the molecular and cellular levels and to design novel interventions to block the progression of disease.

Bacteria↗
PubMed2005

Cloning and characterization of TMPRSS6, a novel type 2 transmembrane serine protease.

We have identified TMPRSS6, a novel type 2 transmembrane serine protease. TMPRSS6 possesses all the signature motifs of the family of transmembrane serine proteases (TMPRSSs), including a transmembrane domain, an LDL receptor class A (LDLRA) domain, a scavenger receptor cysteine-rich (SRCR) domain, and a serine protease domain. The substrate specificity of TMPRSS6 is slightly different from those of other TMPRSS family members. Combined with the finding that TMPRSS6 is expressed strongly in the thyroid and weakly in the trachea, this may indicate that TMPRSS6 has a specialized role.

Amino Acid Sequence↗
PubMed2004

Amplification refractory mutation system PCR assays for the detection of variola and Orthopoxvirus.

PCR assays that can identify the presence of variola virus (VARV) sequences in an unknown DNA sample were developed using principles established for the amplification refractory mutation system (ARMS). The assay's specificity utilised unique single nucleotide polymorphisms (SNP) identified among Orthopoxvirus (OPV) orthologs of the vaccinia virus Copenhagen strain A13L and A36R genes. When a variola virus specific primer was used with a consensus primer in an ARMS assay with different Orthopoxvirus genomes, a PCR product was only amplified from variola virus DNA. Incorporating a second consensus primer into the assay produced a multiplex PCR that provided Orthopoxvirus generic and variola-specific products with variola virus DNA. We tested two single nucleotide polymorphisms with a panel of 43 variola virus strains, collected over 40 years from countries across the world, and have shown that they provide reliable markers for variola virus identification. The variola virus specific primers did not produce amplicons with either assay format when tested with 50 other Orthopoxvirus DNA samples. Our analysis shows that these two polymorphisms were conserved in variola virus genomes and provide a reliable signature of Orthopoxvirus species identification.

Animals↗
PubMed1997

The WW domain of neural protein FE65 interacts with proline-rich motifs in Mena, the mammalian homolog of Drosophila enabled.

The neural protein FE65 contains two types of protein-protein interaction modules: one WW binding domain and two phosphotyrosine binding domains. The carboxyl-terminal phosphotyrosine binding domain of FE65 interacts in vivo with the beta-amyloid precursor protein, which is implicated in Alzheimer disease. To understand the function of this adapter protein, we identified binding partners for the FE65 WW domain. Proline-rich sequences sharing a proline-proline-leucine-proline core motif were recovered by screening expression libraries for ligands of the FE65 WW domain. Five proteins of molecular masses 60, 75, 80, 140, and 200 kDa could be purified from mouse brain lysates by affinity to the FE65 WW domain. We identified two of these five proteins as the 80- and 140-kDa isoforms encoded by Mena, the mammalian homolog of the Drosophila Enabled gene. Using the SPOTs technique of peptide synthesis, we identified the sequences in Mena that interact with the FE65 WW domain and found that they contain the signature proline-proline-leucine-proline motif. Finally, we demonstrated that Mena binds to FE65 in vivo by coimmunoprecipitation assay from COS cell extracts. The specificity of the Mena-FE65 WW domain association was confirmed by competition assays. Further characterization of the FE65-Mena complex may identify a physiological role for these proteins in beta-amyloid precursor protein biogenesis and may help in understanding the mechanism of molecular changes that underlie Alzheimer disease.

Amino Acid Sequence↗