Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 14 de 14
Filter
1.
PLoS One ; 5(8): e12089, 2010 Aug 09.
Article in English | MEDLINE | ID: mdl-20711500

ABSTRACT

BACKGROUND: Bacillus anthracis, Francisella tularensis, and Yersinia pestis are bacterial pathogens that can cause anthrax, lethal acute pneumonic disease, and bubonic plague, respectively, and are listed as NIAID Category A priority pathogens for possible use as biological weapons. However, the interactions between human proteins and proteins in these bacteria remain poorly characterized leading to an incomplete understanding of their pathogenesis and mechanisms of immune evasion. METHODOLOGY: In this study, we used a high-throughput yeast two-hybrid assay to identify physical interactions between human proteins and proteins from each of these three pathogens. From more than 250,000 screens performed, we identified 3,073 human-B. anthracis, 1,383 human-F. tularensis, and 4,059 human-Y. pestis protein-protein interactions including interactions involving 304 B. anthracis, 52 F. tularensis, and 330 Y. pestis proteins that are uncharacterized. Computational analysis revealed that pathogen proteins preferentially interact with human proteins that are hubs and bottlenecks in the human PPI network. In addition, we computed modules of human-pathogen PPIs that are conserved amongst the three networks. Functionally, such conserved modules reveal commonalities between how the different pathogens interact with crucial host pathways involved in inflammation and immunity. SIGNIFICANCE: These data constitute the first extensive protein interaction networks constructed for bacterial pathogens and their human hosts. This study provides novel insights into host-pathogen interactions.


Subject(s)
Bacillus anthracis/metabolism , Bacterial Proteins/metabolism , Computational Biology , Francisella tularensis/metabolism , Host-Pathogen Interactions , Yersinia pestis/metabolism , Bacillus anthracis/physiology , Francisella tularensis/physiology , Humans , Protein Binding , Yersinia pestis/physiology
3.
Am J Med Genet B Neuropsychiatr Genet ; 153B(1): 321-2, 2010 Jan 05.
Article in English | MEDLINE | ID: mdl-19475634

ABSTRACT

Linkage analysis on Utah pedigrees with strong family histories of major depression including only cases with the SLC6A4 HTTLPR short allele revealed a linkage peak on chromosome 4 (maximum HLOD = 3.5). This evidence suggests epistasis between SLC6A4 and an unknown gene as risk factors for major depression.


Subject(s)
Chromosomes, Human, Pair 4 , Depressive Disorder, Major/genetics , Epistasis, Genetic , Serotonin Plasma Membrane Transport Proteins/genetics , Genetic Linkage , Humans , Risk Factors
4.
J Virol ; 83(23): 12068-83, 2009 Dec.
Article in English | MEDLINE | ID: mdl-19776120

ABSTRACT

Most poxviruses express multiple proteins containing ankyrin (ANK) repeats accounting for a large superfamily of related but unique determinants of poxviral tropism. Recently, select members of this novel family of poxvirus proteins have drawn considerable attention for their potential roles in modulating intracellular signaling networks during viral infection. The rabbit-specific poxvirus, myxoma virus (MYXV), encodes four unique ANK repeat proteins, termed M-T5, M148, M149, and M150, all of which include a carboxy-terminal PRANC domain which closely resembles a cellular protein motif called the F-box domain. Here, we show that each MYXV-encoded ANK repeat protein, including M-T5, interacts directly with the Skp1 component of the host SCF ubiquitin ligase complex, and that the binding of M-T5 to cullin 1 is indirect via binding to Skp1 in the host SCF complex. To understand the significance of these virus-host protein interactions, the various binding domains of M-T5 were mapped. The N-terminal ANK repeats I and II were identified as being important for interaction with Akt, whereas the C-terminal PRANC/F-box-like domain was essential for binding to Skp1. We also report that M-T5 can bind Akt and the host SCF complex (via Skp1) simultaneously in MYXV-infected cells. Finally, we report that M-T5 specifically mediates the relocalization of Akt from the nucleus to the cytoplasm during infection with the wild-type MYXV, but not the M-T5 knockout version of the virus. These results indicate that ANK/PRANC proteins play a critical role in reprogramming disparate cellular signaling cascades to establish a new cellular environment more favorable for virus replication.


Subject(s)
Myxoma virus/pathogenicity , Proto-Oncogene Proteins c-akt/metabolism , S-Phase Kinase-Associated Proteins/metabolism , Signal Transduction , Viral Proteins/physiology , Animals , Cell Line , Cell Nucleus/chemistry , Chlorocebus aethiops , Cullin Proteins/metabolism , Cytoplasm/chemistry , Humans , Protein Binding , Protein Interaction Mapping , SKP Cullin F-Box Protein Ligases/metabolism
5.
J Proteome Res ; 8(9): 4311-8, 2009 Sep.
Article in English | MEDLINE | ID: mdl-19637933

ABSTRACT

Vaccinia virus, a large double-stranded DNA virus, is the prototype of the Orthopoxvirus genus, which includes several pathogenic poxviruses of humans, such as monkeypox virus and variola virus. Here, we report a comprehensive yeast two-hybrid (Y2H) screening for the protein-protein interactions between vaccinia and human proteins. A total of 109 novel vaccinia-human protein interactions were detected among 33 viral proteins. To validate subsets of those interactions, we constructed an ORFeome library of vaccinia virus strain WR using the Gateway plasmid cloning system. By co-expressing selected vaccinia and host proteins in a variety of expression systems, we found that at least 17 of the Y2H hits identified between vaccinia and human proteins can be verified by independent methods using GST pull-down assays, representing a 63% validation rate for the Y2H hits examined (17/27). Because the cloned ORFs are conveniently transferable from the entry vectors to various destination expression vectors, the vaccinia ORFeome library will be a useful resource for future high-throughput functional proteomic experiments.


Subject(s)
Host-Pathogen Interactions/physiology , Two-Hybrid System Techniques , Vaccinia/metabolism , Viral Proteins/metabolism , Humans , Open Reading Frames , Polymerase Chain Reaction , Protein Interaction Mapping/methods , Proteomics/methods , Recombinant Fusion Proteins/metabolism , Reproducibility of Results
6.
Proc Natl Acad Sci U S A ; 106(22): 9045-50, 2009 Jun 02.
Article in English | MEDLINE | ID: mdl-19451633

ABSTRACT

Identification of the binary interactions between viral and host proteins has become a valuable tool for investigating viral tropism and pathogenesis. Here, we present the first systematic protein interaction screening of the unique variola virus proteome by using yeast 2-hybrid screening against a variety of human cDNA libraries. Several protein-protein interactions were identified, including an interaction between variola G1R, an ankryin/F-box containing protein, and human nuclear factor kappa-B1 (NF-kappaB1)/p105. This represents the first direct interaction between a pathogen-encoded protein and NF-kappaB1/p105. Orthologs of G1R are present in a variety of pathogenic orthopoxviruses, but not in vaccinia virus, and expression of any one of these viral proteins blocks NF-kappaB signaling in human cells. Thus, proteomic screening of variola virus has the potential to uncover modulators of the human innate antiviral responses.


Subject(s)
Host-Pathogen Interactions , NF-kappa B p50 Subunit/antagonists & inhibitors , Proteomics , Variola virus/metabolism , Viral Proteins/metabolism , Cell Line , Gene Library , Humans , NF-kappa B p50 Subunit/metabolism , Orthopoxvirus/metabolism , Orthopoxvirus/pathogenicity , Two-Hybrid System Techniques
7.
Biotechniques ; 44(2): 265-73, 2008 Feb.
Article in English | MEDLINE | ID: mdl-18330356

ABSTRACT

The nuclear yeast two-hybrid (Y2H) system is the most widely used technology for detecting interactions between proteins. A common approach is to screen specific test proteins (baits) against large compilations of randomly cloned proteins (prey libraries). For eukaryotic organisms, libraries have traditionally been generated using messenger RNA (mRNA) extracted from various tissues and cells. Here we present a library construction strategy made possible by ongoing public efforts to establish collections of full-length protein encoding clones. Our approach generates libraries that are essentially normalized and contain both randomly fragmented as well as full-length inserts. We refer to this type of protein-coding clone-derived library as random and full-length (RAFL) Y2H library. The library described here is based on clones from the Mammalian Gene Collection, but our strategy is compatible with the use of any protein-coding clone collection from any organism in any vector and does not require inserts to be devoid of untranslated regions. We tested our prototype human RAFL library against a set of baits that had previously been searched against multiple cDNA libraries. These Y2H searches yielded a combination of novel as well as expected interactions, indicating that the RAFL library constitutes a valuable complement to Y2H cDNA libraries.


Subject(s)
Cloning, Molecular/methods , Gene Library , Proteins/metabolism , Saccharomyces cerevisiae/metabolism , Two-Hybrid System Techniques , Humans , Protein Binding , Proteins/genetics , Saccharomyces cerevisiae/genetics
8.
Hum Mol Genet ; 15(18): 2709-20, 2006 Sep 15.
Article in English | MEDLINE | ID: mdl-16893906

ABSTRACT

The molecular etiology of obesity predisposition is largely unknown. Here, we present evidence that genetic variation in TBC1D1 confers risk for severe obesity in females. We identified a coding variant (R125W) in TBC1D1 that segregated with the disease in 4p15-14-linked obesity pedigrees. In cases derived from pedigrees with the strongest linkage evidence, the variant was significantly associated with obesity (P=0.000007) and chromosomes carrying R125W accounted for the majority of the evidence that originally linked 4p15-14 with the disease. In addition, by selecting families that segregated R125W with obesity, we were able to generate highly significant linkage evidence for an obesity predisposition locus at 4q34-35. This result provides additional and confirming evidence that R125W affects obesity susceptibility, delimits the location of an obesity gene at 4q34-35 and identifies a gene/gene interaction that influences the risk for obesity predisposition. Finally, although the function of TBC1D1 is unknown, the protein is structurally similar to a known regulator of insulin-mediated Glut4 translocation.


Subject(s)
Endopeptidases/genetics , Obesity/genetics , Oncogene Proteins/genetics , Chromosomes, Human, Pair 4/genetics , Female , Gene Expression , Genetic Variation , Haplotypes , Humans , Linkage Disequilibrium , Lod Score , Male , Obesity/etiology , Pedigree , Phenotype , Polymorphism, Single Nucleotide , Proto-Oncogene Proteins , RNA, Messenger/genetics , RNA, Messenger/metabolism , Tissue Distribution , Ubiquitin Thiolesterase
9.
BMC Mol Biol ; 6: 12, 2005 May 13.
Article in English | MEDLINE | ID: mdl-15892892

ABSTRACT

BACKGROUND: In humans, mutation of ELAC2 is associated with an increased risk of prostate cancer. ELAC2 has been shown to have tRNase Z activity and is associated with the gamma-tubulin complex. RESULTS: In this work, we show that the yeast homolog of ELAC2, encoded by TRZ1 (tRNase Z 1), is involved genetically in RNA processing. The temperature sensitivity of a trz1 mutant can be rescued by multiple copies of REX2, which encodes a protein with RNA 3' processing activity, suggesting a role of Trz1p in RNA processing in vivo. Trz1p has two putative nucleotide triphosphate-binding motifs (P-loop) and a conserved histidine motif. The histidine motif and the putative nucleotide binding motif at the C-domain are important for Trz1p function because mutant proteins bearing changes to the critical residues in these motifs are unable to rescue deletion of TRZ1. The growth defect exhibited by trz1 yeast is not complemented by the heterologous ELAC2, suggesting that Trz1p may have additional functions in yeast. CONCLUSION: Our results provide genetic evidence that prostate cancer susceptibility gene ELAC2 may be involved in RNA processing, especially rRNA processing and mitochondrial function.


Subject(s)
Endoribonucleases/physiology , Neoplasm Proteins/physiology , Prostatic Neoplasms/genetics , RNA Processing, Post-Transcriptional/genetics , Saccharomyces cerevisiae Proteins/chemistry , Saccharomyces cerevisiae/genetics , Amino Acid Motifs/physiology , Amino Acid Sequence , DNA, Recombinant/genetics , Endoribonucleases/genetics , Gene Expression Regulation, Neoplastic , Genetic Complementation Test/methods , Humans , Male , Mitochondria/metabolism , Molecular Sequence Data , Mutation , Neoplasm Proteins/genetics , Prostatic Neoplasms/pathology , RNA Splicing , RNA, Messenger/metabolism , RNA, Ribosomal/metabolism , RNA, Transfer/metabolism , Saccharomyces cerevisiae/growth & development , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/physiology , Sequence Analysis, Protein , Sequence Homology, Amino Acid , Temperature
10.
Am J Med Genet B Neuropsychiatr Genet ; 135B(1): 85-93, 2005 May 05.
Article in English | MEDLINE | ID: mdl-15806581

ABSTRACT

Major depressive disorder (MDD) is a common, clinically heterogeneous disorder often found comorbid with other disorders. We studied recurrent, early-onset MDD (MDD-RE) and anxiety disorders in combination to define powerful phenotypes for genetic study. We used 87 large, extended Utah pedigrees to investigate linkage to 3 phenotypes: "MDD-RE;" "MDD-RE or anxiety;" and "MDD-RE and anxiety;" where in the latter definition the disorders must appear comorbid within an individual. Pedigrees ranged in size from 2 to 6 generations and contained 3 to 42 individuals affected with MDD or anxiety (718 total). In primary analyses, we identified three regions with at least suggestive genome-wide evidence for linkage on chromosomes 3centr, 7p, and 18q. Both 7p and 18q are replication findings for related phenotypes. The best linkage evidence was for a novel locus at 3p12.3-q12.3 (LOD = 3.88, "MDD-RE or anxiety") and 18q21.33-q22.2 (LOD = 3.75, "MDD-RE and anxiety"), a well-established susceptibility locus for bipolar disorder. In our secondary sex-specific analyses, we identified two further regions of interest on chromosomes 4q and 15q. Using linked pedigrees, we localized 3centr and 18q to 9.8 and 12.2 cM, respectively, with potential for further localization with the addition of markers in specific pedigrees. Our success in replication and novel locus identification illustrates the utility of large extended pedigrees for common disorders, such as MDD. Further, it supports the hypothesis that MDD and anxiety disorders have over-lapping genetic etiologies and suggests that comorbid diagnoses may be useful in defining more genetically homogeneous forms of MDD for linkage mapping.


Subject(s)
Anxiety Disorders/genetics , Depressive Disorder/genetics , Genetic Linkage/genetics , Genetic Predisposition to Disease/genetics , Genome, Human , Anxiety Disorders/psychology , Chromosomes, Human, Pair 18/genetics , Chromosomes, Human, Pair 3/genetics , Chromosomes, Human, Pair 7/genetics , Depressive Disorder/psychology , Family Health , Female , Genotype , Humans , Lod Score , Male , Microsatellite Repeats , Pedigree , Recurrence , Sex Factors , Utah
11.
Mamm Genome ; 15(6): 460-71, 2004 Jun.
Article in English | MEDLINE | ID: mdl-15181538

ABSTRACT

Our primary objective was to discover simplified mouse models corresponding to human obesity linkages. We used the B10.UW- H3(b) we Pax1(un) a(t)/Sn (B10.UW) congenic strain, a subcongenic strain with a reduced UW strain donor region, and their C57BL/10SnJ background strain. The congenic and subcongenic UW strain donor regions are on mouse Chr 2. We measured body length [anal-nasal (AN) length], summed fat depot weights normalized for body weight (Adiposity Index, AI), and percentage of body weight that is lipid. The B10.UW congenic and subcongenic strains have significantly smaller AN lengths ( p < 0.0001) and have a significantly lower AI and percentage of body weight as fat than the background strain ( p < 0.0001). In an F(2) intercross of the congenic and background strains, AN and AI were both linked to the distal half of the donor region with LOD scores greater than 19 and 5, respectively. F(2) haplotypes identified a minimal region for AN linkage of 0.8 megabases (Mb) that is estimated to express four genes in the current Celera mouse genome assembly. We narrowed the most likely location of the obesity gene to 15 Mb whose homologous genes are all located on human Chr 20 in the region surrounding the centromere. Since a previous study identified human obesity linkage peaking near the centromere, then the B10.UW mice may exhibit obesity due to the homologous gene.


Subject(s)
Mice/anatomy & histology , Obesity/genetics , Animals , Biometry , Chromosome Mapping , Female , Male , Mice/genetics , Mice, Congenic , Mice, Inbred Strains , Phenotype
12.
Hum Genet ; 114(4): 349-53, 2004 Mar.
Article in English | MEDLINE | ID: mdl-14727179

ABSTRACT

Familial hypercholesterolemia results from mutations in the low-density lipoprotein (LDL) receptor or apolipoprotein B genes. We have previously reported the identification of a Utah autosomal-dominant hypercholesterolemia pedigree (kindred 1173) that did not show linkage to either of these loci (Hunt et al. 2000). Expansion of the pedigree and increased marker density within the region of interest have resulted in a multipoint LOD score of 9.6 and enabled us to decrease the size of the linked region to approximately 7.5 Mbp. In addition, we were able to identify additional families sharing the same microsatellite haplotype. While all haplotype carriers in kindred 1173 (K1173) are affected, the haplotype carriers within the newly identified families are unaffected, suggesting that the causal mutation in K1173 had occurred after divergence of these pedigrees from a common ancestor. Mutation screening of genes in the region identified a single nucleotide variant (G-->T) present on the K1173 haplotype that was not present on the same haplotype in the other kindreds. This variant results in a D374Y missense change in the gene PCSK9.


Subject(s)
Hyperlipoproteinemia Type II/genetics , Point Mutation , Serine Endopeptidases/genetics , Female , Genes, Dominant , Haplotypes , Heterozygote , Humans , Lod Score , Male , Mutation, Missense , Pedigree , Phenotype , Proprotein Convertase 9 , Proprotein Convertases , Utah
13.
Am J Hum Genet ; 73(6): 1271-81, 2003 Dec.
Article in English | MEDLINE | ID: mdl-14606042

ABSTRACT

Major depression disorder is a common psychiatric disease with a major economic impact on society. In many cases, no effective treatment is available. The etiology of major depression is complex, but it is clear that the disease is, to a large extent, determined genetically, especially among individuals with a familial history of major depression, presumably through the involvement of multiple predisposition genes in addition to an environmental component. As a first step toward identification of chromosomal loci contributing to genetic predisposition to major depression, we have conducted a genomewide scan by using 628 microsatellite markers on 1,890 individuals from 110 Utah pedigrees with a strong family history of major depression. We identified significant linkage to major depression in males at marker D12S1300 (multipoint heterogeneity LOD score 4.6; P=.00003 after adjustment for multiple testing). With additional markers, the linkage evidence became highly significant, with the multipoint heterogeneity LOD score at marker D12S1706 increasing to 6.1 (P=.0000007 after adjustment for multiple testing). This study confirms the presence of one or more genes involved in psychiatric diseases on the q arm of chromosome 12 and provides strong evidence for the existence of a sex-specific predisposition gene to major depression at 12q22-q23.2.


Subject(s)
Chromosomes, Human, Pair 12/genetics , Depressive Disorder, Major/genetics , Genetic Linkage/genetics , Chromosome Mapping , Genetic Testing , Genome, Human , Humans , Microsatellite Repeats/genetics , Pedigree , Utah
14.
Am J Hum Genet ; 70(6): 1459-68, 2002 Jun.
Article in English | MEDLINE | ID: mdl-11957135

ABSTRACT

Although the predisposition to morbid obesity is heritable, the identities of the disease-causing genes are largely unknown. Therefore, we have conducted a genomewide search with 628 markers, using multigenerational Utah pedigrees to identify genes involved in predisposition to obesity. In the genomewide search, we identified a highly significant linkage to high body-mass index in female patients, at D4S2632, with a multipoint heterogeneity LOD (HLOD) score of 6.1 and a nonparametric linkage (NPL) score of 5.3. To further delineate the linkage, we increased both the marker density around D4S2632 and the size of our pedigree data set. As a result, the linkage evidence increased to a multipoint HLOD score of 9.2 (at D4S3350) and an NPL score of 11.3. Evidence from almost half of the families in this analysis support this linkage, and therefore the gene in this region might account for a significant percentage of the genetic predisposition to severe obesity in females. However, further studies are necessary to clarify the effect that this gene has in males and in the general population.


Subject(s)
Chromosomes, Human, Pair 4/genetics , Genetic Predisposition to Disease/genetics , Obesity/genetics , Body Mass Index , Chromosome Mapping , Female , Genes, Dominant , Genes, Recessive , Genetic Markers/genetics , Genome, Human , Genotype , Humans , Lod Score , Male , Pedigree , Phenotype , Sex Characteristics , Utah
SELECTION OF CITATIONS
SEARCH DETAIL
...