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1.  Common Mechanism for RNA Encapsidation by Negative-Strand RNA Viruses 
Journal of Virology  2014;88(7):3766-3775.
ABSTRACT
The nucleocapsid of a negative-strand RNA virus is assembled with a single nucleocapsid protein and the viral genomic RNA. The nucleocapsid protein polymerizes along the length of the single-strand genomic RNA (viral RNA) or its cRNA. This process of encapsidation occurs concomitantly with genomic replication. Structural comparisons of several nucleocapsid-like particles show that the mechanism of RNA encapsidation in negative-strand RNA viruses has many common features. Fundamentally, there is a unifying mechanism to keep the capsid protein protomer monomeric prior to encapsidation of viral RNA. In the nucleocapsid, there is a cavity between two globular domains of the nucleocapsid protein where the viral RNA is sequestered. The viral RNA must be transiently released from the nucleocapsid in order to reveal the template RNA sequence for transcription/replication. There are cross-molecular interactions among the protein subunits linearly along the nucleocapsid to stabilize its structure. Empty capsids can form in the absence of RNA. The common characteristics of RNA encapsidation not only delineate the evolutionary relationship of negative-strand RNA viruses but also provide insights into their mechanism of replication.
IMPORTANCE What separates negative-strand RNA viruses (NSVs) from the rest of the virosphere is that the nucleocapsid of NSVs serves as the template for viral RNA synthesis. Their viral RNA-dependent RNA polymerase can induce local conformational changes in the nucleocapsid to temporarily release the RNA genome so that the viral RNA-dependent RNA polymerase can use it as the template for RNA synthesis during both transcription and replication. After RNA synthesis at the local region is completed, the viral RNA-dependent RNA polymerase processes downstream, and the RNA genome is restored in the nucleocapsid. We found that the nucleocapsid assembly of all NSVs shares three essential elements: a monomeric capsid protein protomer, parallel orientation of subunits in the linear nucleocapsid, and a (5H + 3H) motif that forms a proper cavity for sequestration of the RNA. This observation also suggests that all NSVs evolved from a common ancestor that has this unique nucleocapsid.
doi:10.1128/JVI.03483-13
PMCID: PMC3993539  PMID: 24429372
2.  1.55 Å resolution X-ray crystal structure of Rv3902c from Mycobacterium tuberculosis  
The 1.55 Å resolution X-ray crystal structure of Rv3902c from M. tuberculosis reveals a novel fold.
The crystallographic structure of the Mycobacterium tuberculosis (TB) protein Rv3902c (176 residues; molecular mass of 19.8 kDa) was determined at 1.55 Å resolution. The function of Rv3902c is unknown, although several TB genes involved in bacterial pathogenesis are expressed from the operon containing the Rv3902c gene. The unique structural fold of Rv3902c contains two domains, each consisting of antiparallel β-sheets and α-helices, creating a hand-like binding motif with a small binding pocket in the palm. Structural homology searches reveal that Rv3902c has an overall structure similar to that of the Salmonella virulence-factor chaperone InvB, with an r.m.s.d. for main-chain atoms of 2.3 Å along an aligned domain.
doi:10.1107/S2053230X14003793
PMCID: PMC3976054  PMID: 24699730
Mycobacterium tuberculosis; Rv3902c; virulence-factor chaperone; novel fold
3.  Mutations causing medullary cystic kidney disease type 1 (MCKD1) lie in a large VNTR in MUC1 missed by massively parallel sequencing 
Nature genetics  2013;45(3):299-303.
While genetic lesions responsible for some Mendelian disorders can be rapidly discovered through massively parallel sequencing (MPS) of whole genomes or exomes, not all diseases readily yield to such efforts. We describe the illustrative case of the simple Mendelian disorder medullary cystic kidney disease type 1 (MCKD1), mapped more than a decade ago to a 2-Mb region on chromosome 1. Ultimately, only by cloning, capillary sequencing, and de novo assembly, we found that each of six MCKD1 families harbors an equivalent, but apparently independently arising, mutation in sequence dramatically underrepresented in MPS data: the insertion of a single C in one copy (but a different copy in each family) of the repeat unit comprising the extremely long (~1.5-5 kb), GC-rich (>80%), coding VNTR in the mucin 1 gene. The results provide a cautionary tale about the challenges in identifying genes responsible for Mendelian, let alone more complex, disorders through MPS.
doi:10.1038/ng.2543
PMCID: PMC3901305  PMID: 23396133
4.  Structural and Functional Characterization of the Mumps Virus Phosphoprotein 
Journal of Virology  2013;87(13):7558-7568.
The phosphoprotein (P) is virally encoded by the Rhabdoviridae and Paramyxoviridae in the order Mononegavirales. P is a self-associated oligomer and forms complexes with the large viral polymerase protein (L), the nucleocapsid protein (N), and the assembled nucleocapsid. P from different viruses has shown structural diversities even though their essential functions are the same. We systematically mapped the domains in mumps virus (MuV) P and investigated their interactions with nucleocapsid-like particles (NLPs). Similar to other P proteins, MuV P contains N-terminal, central, and C-terminal domains with flexible linkers between neighboring domains. By pulldown assays, we discovered that in addition to the previously proposed nucleocapsid binding domain (residues 343 to 391), the N-terminal region of MuV P (residues 1 to 194) could also bind NLPs. Further analysis of binding kinetics was conducted using surface plasmon resonance. This is the first observation that both the N- and C-terminal regions of a negative-strand RNA virus P are involved in binding the nucleocapsid. In addition, we defined the oligomerization domain (POD) of MuV P as residues 213 to 277 and determined its crystal structure. The tetrameric MuV POD is formed by one pair of long parallel α-helices with another pair in opposite orientation. Unlike the parallel orientation of each α-helix in the tetramer of Sendai virus POD, this represents a novel orientation of a POD where both the N- and the C-terminal domains are at either end of the tetramer. This is consistent with the observation that both the N- and the C-terminal domains are involved in binding the nucleocapsid.
doi:10.1128/JVI.00653-13
PMCID: PMC3700306  PMID: 23637399
5.  Nucleocapsid protein structures from orthobunyaviruses reveal insight into ribonucleoprotein architecture and RNA polymerization 
Nucleic Acids Research  2013;41(11):5912-5926.
All orthobunyaviruses possess three genome segments of single-stranded negative sense RNA that are encapsidated with the virus-encoded nucleocapsid (N) protein to form a ribonucleoprotein (RNP) complex, which is uncharacterized at high resolution. We report the crystal structure of both the Bunyamwera virus (BUNV) N–RNA complex and the unbound Schmallenberg virus (SBV) N protein, at resolutions of 3.20 and 2.75 Å, respectively. Both N proteins crystallized as ring-like tetramers and exhibit a high degree of structural similarity despite classification into different orthobunyavirus serogroups. The structures represent a new RNA-binding protein fold. BUNV N possesses a positively charged groove into which RNA is deeply sequestered, with the bases facing away from the solvent. This location is highly inaccessible, implying that RNA polymerization and other critical base pairing events in the virus life cycle require RNP disassembly. Mutational analysis of N protein supports a correlation between structure and function. Comparison between these crystal structures and electron microscopy images of both soluble tetramers and authentic RNPs suggests the N protein does not bind RNA as a repeating monomer; thus, it represents a newly described architecture for bunyavirus RNP assembly, with implications for many other segmented negative-strand RNA viruses.
doi:10.1093/nar/gkt268
PMCID: PMC3675483  PMID: 23595147
6.  Chronic Granulomatous Disease Presenting as Aseptic Ascites in a 2-Year-Old Child 
Case Reports in Immunology  2013;2013:927897.
Chronic granulomatous disease (CGD) is a rare inherited immunodeficiency syndrome that results from abnormal nicotinamide adenine dinucleotide phosphate (NADPH) oxidase function. This defect leads to recurrent catalase-positive bacterial and fungal infections as well as associated granuloma formation. We review the case of a 2-year-old boy who presented with ascites and fever of an unknown origin as manifestations of CGD. Cultures were negative for infection throughout his course, and CGD was suspected after identification of granulomas on peritoneal biopsy. Genetic testing revealed a novel mutation in the CYBB gene underlying his condition. This paper highlights the importance of considering CGD in the differential diagnosis of fever of unknown origin and ascites in children.
doi:10.1155/2013/927897
PMCID: PMC4207586  PMID: 25379308
7.  Haplotype structure in Ashkenazi Jewish BRCA1 and BRCA2 mutation carriers 
Im, Kate M. | Kirchhoff, Tomas | Wang, Xianshu | Green, Todd | Chow, Clement Y. | Vijai, Joseph | Korn, Joshua | Gaudet, Mia M. | Fredericksen, Zachary | Pankratz, V. Shane | Guiducci, Candace | Crenshaw, Andrew | McGuffog, Lesley | Kartsonaki, Christiana | Morrison, Jonathan | Healey, Sue | Sinilnikova, Olga M. | Mai, Phuong L. | Greene, Mark H. | Piedmonte, Marion | Rubinstein, Wendy S. | Hogervorst, Frans B. | Rookus, Matti A. | Collée, J. Margriet | Hoogerbrugge, Nicoline | van Asperen, Christi J. | Meijers-Heijboer, Hanne E. J. | Van Roozendaal, Cees E. | Caldes, Trinidad | Perez-Segura, Pedro | Jakubowska, Anna | Lubinski, Jan | Huzarski, Tomasz | Blecharz, Paweł | Nevanlinna, Heli | Aittomäki, Kristiina | Lazaro, Conxi | Blanco, Ignacio | Barkardottir, Rosa B. | Montagna, Marco | D'Andrea, Emma | Devilee, Peter | Olopade, Olufunmilayo I. | Neuhausen, Susan L. | Peissel, Bernard | Bonanni, Bernardo | Peterlongo, Paolo | Singer, Christian F. | Rennert, Gad | Lejbkowicz, Flavio | Andrulis, Irene L. | Glendon, Gord | Ozcelik, Hilmi | Toland, Amanda Ewart | Caligo, Maria Adelaide | Beattie, Mary S. | Chan, Salina | Domchek, Susan M. | Nathanson, Katherine L. | Rebbeck, Timothy R. | Phelan, Catherine | Narod, Steven | John, Esther M. | Hopper, John L. | Buys, Saundra S. | Daly, Mary B. | Southey, Melissa C. | Terry, Mary-Beth | Tung, Nadine | Hansen, Thomas v. O. | Osorio, Ana | Benitez, Javier | Durán, Mercedes | Weitzel, Jeffrey N. | Garber, Judy | Hamann, Ute | Peock, Susan | Cook, Margaret | Oliver, Clare T. | Frost, Debra | Platte, Radka | Evans, D. Gareth | Eeles, Ros | Izatt, Louise | Paterson, Joan | Brewer, Carole | Hodgson, Shirley | Morrison, Patrick J. | Porteous, Mary | Walker, Lisa | Rogers, Mark T. | Side, Lucy E. | Godwin, Andrew K. | Schmutzler, Rita K. | Wappenschmidt, Barbara | Laitman, Yael | Meindl, Alfons | Deissler, Helmut | Varon-Mateeva, Raymonda | Preisler-Adams, Sabine | Kast, Karin | Venat-Bouvet, Laurence | Stoppa-Lyonnet, Dominique | Chenevix-Trench, Georgia | Easton, Douglas F. | Klein, Robert J. | Daly, Mark J. | Friedman, Eitan | Dean, Michael | Clark, Andrew G. | Altshuler, David M. | Antoniou, Antonis C. | Couch, Fergus J. | Offit, Kenneth | Gold, Bert
Human genetics  2011;130(5):685-699.
Abstract Three founder mutations in BRCA1 and BRCA2 contribute to the risk of hereditary breast and ovarian cancer in Ashkenazi Jews (AJ). They are observed at increased frequency in the AJ compared to other BRCA mutations in Caucasian non-Jews (CNJ). Several authors have proposed that elevated allele frequencies in the surrounding genomic regions reflect adaptive or balancing selection. Such proposals predict long-range linkage dis-equilibrium (LD) resulting from a selective sweep, although genetic drift in a founder population may also act to create long-distance LD. To date, few studies have used the tools of statistical genomics to examine the likelihood of long-range LD at a deleterious locus in a population that faced a genetic bottleneck. We studied the genotypes of hundreds of women from a large international consortium of BRCA1 and BRCA2 mutation carriers and found that AJ women exhibited long-range haplotypes compared to CNJ women. More than 50% of the AJ chromosomes with the BRCA1 185delAG mutation share an identical 2.1 Mb haplotype and nearly 16% of AJ chromosomes carrying the BRCA2 6174delT mutation share a 1.4 Mb haplotype. Simulations based on the best inference of Ashkenazi population demography indicate that long-range haplotypes are expected in the context of a genome-wide survey. Our results are consistent with the hypothesis that a local bottleneck effect from population size constriction events could by chance have resulted in the large haplotype blocks observed at high frequency in the BRCA1 and BRCA2 regions of Ashkenazi Jews.
doi:10.1007/s00439-011-1003-z
PMCID: PMC3196382  PMID: 21597964
8.  Deep resequencing of GWAS loci identifies independent rare variants associated with inflammatory bowel disease 
Nature Genetics  2011;43(11):1066-1073.
More than a thousand disease susceptibility loci have been identified via genome-wide association studies (GWAS) of common variants; however, the specific genes and full allelic spectrum of causal variants underlying these findings generally remain to be defined. We utilize pooled next-generation sequencing to study 56 genes in regions associated to Crohn’s Disease in 350 cases and 350 controls. Follow up genotyping of 70 rare and low-frequency protein-altering variants (MAF ~ .001-.05) in nine independent case-control series (16054 CD patients, 12153 UC patients, 17575 healthy controls) identifies four additional independent risk factors in NOD2, two additional protective variants in IL23R, a highly significant association to a novel, protective splice variant in CARD9 (p < 1e-16, OR ~ 0.29), as well as additional associations to coding variants in IL18RAP, CUL2, C1orf106, PTPN22 and MUC19. We extend the results of successful GWAS by providing novel, rare, and likely functional variants that will empower functional experiments and predictive models.
doi:10.1038/ng.952
PMCID: PMC3378381  PMID: 21983784
9.  Stratifying Type 2 Diabetes Cases by BMI Identifies Genetic Risk Variants in LAMA1 and Enrichment for Risk Variants in Lean Compared to Obese Cases 
Perry, John R. B. | Voight, Benjamin F. | Yengo, Loïc | Amin, Najaf | Dupuis, Josée | Ganser, Martha | Grallert, Harald | Navarro, Pau | Li, Man | Qi, Lu | Steinthorsdottir, Valgerdur | Scott, Robert A. | Almgren, Peter | Arking, Dan E. | Aulchenko, Yurii | Balkau, Beverley | Benediktsson, Rafn | Bergman, Richard N. | Boerwinkle, Eric | Bonnycastle, Lori | Burtt, Noël P. | Campbell, Harry | Charpentier, Guillaume | Collins, Francis S. | Gieger, Christian | Green, Todd | Hadjadj, Samy | Hattersley, Andrew T. | Herder, Christian | Hofman, Albert | Johnson, Andrew D. | Kottgen, Anna | Kraft, Peter | Labrune, Yann | Langenberg, Claudia | Manning, Alisa K. | Mohlke, Karen L. | Morris, Andrew P. | Oostra, Ben | Pankow, James | Petersen, Ann-Kristin | Pramstaller, Peter P. | Prokopenko, Inga | Rathmann, Wolfgang | Rayner, William | Roden, Michael | Rudan, Igor | Rybin, Denis | Scott, Laura J. | Sigurdsson, Gunnar | Sladek, Rob | Thorleifsson, Gudmar | Thorsteinsdottir, Unnur | Tuomilehto, Jaakko | Uitterlinden, Andre G. | Vivequin, Sidonie | Weedon, Michael N. | Wright, Alan F. | Hu, Frank B. | Illig, Thomas | Kao, Linda | Meigs, James B. | Wilson, James F. | Stefansson, Kari | van Duijn, Cornelia | Altschuler, David | Morris, Andrew D. | Boehnke, Michael | McCarthy, Mark I. | Froguel, Philippe | Palmer, Colin N. A. | Wareham, Nicholas J. | Groop, Leif | Frayling, Timothy M. | Cauchi, Stéphane
PLoS Genetics  2012;8(5):e1002741.
Common diseases such as type 2 diabetes are phenotypically heterogeneous. Obesity is a major risk factor for type 2 diabetes, but patients vary appreciably in body mass index. We hypothesized that the genetic predisposition to the disease may be different in lean (BMI<25 Kg/m2) compared to obese cases (BMI≥30 Kg/m2). We performed two case-control genome-wide studies using two accepted cut-offs for defining individuals as overweight or obese. We used 2,112 lean type 2 diabetes cases (BMI<25 kg/m2) or 4,123 obese cases (BMI≥30 kg/m2), and 54,412 un-stratified controls. Replication was performed in 2,881 lean cases or 8,702 obese cases, and 18,957 un-stratified controls. To assess the effects of known signals, we tested the individual and combined effects of SNPs representing 36 type 2 diabetes loci. After combining data from discovery and replication datasets, we identified two signals not previously reported in Europeans. A variant (rs8090011) in the LAMA1 gene was associated with type 2 diabetes in lean cases (P = 8.4×10−9, OR = 1.13 [95% CI 1.09–1.18]), and this association was stronger than that in obese cases (P = 0.04, OR = 1.03 [95% CI 1.00–1.06]). A variant in HMG20A—previously identified in South Asians but not Europeans—was associated with type 2 diabetes in obese cases (P = 1.3×10−8, OR = 1.11 [95% CI 1.07–1.15]), although this association was not significantly stronger than that in lean cases (P = 0.02, OR = 1.09 [95% CI 1.02–1.17]). For 36 known type 2 diabetes loci, 29 had a larger odds ratio in the lean compared to obese (binomial P = 0.0002). In the lean analysis, we observed a weighted per-risk allele OR = 1.13 [95% CI 1.10–1.17], P = 3.2×10−14. This was larger than the same model fitted in the obese analysis where the OR = 1.06 [95% CI 1.05–1.08], P = 2.2×10−16. This study provides evidence that stratification of type 2 diabetes cases by BMI may help identify additional risk variants and that lean cases may have a stronger genetic predisposition to type 2 diabetes.
Author Summary
Individuals with Type 2 diabetes (T2D) can present with variable clinical characteristics. It is well known that obesity is a major risk factor for type 2 diabetes, yet patients can vary considerably—there are many lean diabetes patients and many overweight people without diabetes. We hypothesized that the genetic predisposition to the disease may be different in lean (BMI<25 Kg/m2) compared to obese cases (BMI≥30 Kg/m2). Specifically, as lean T2D patients had lower risk than obese patients, they must have been more genetically susceptible. Using genetic data from multiple genome-wide association studies, we tested genetic markers across the genome in 2,112 lean type 2 diabetes cases (BMI<25 kg/m2), 4,123 obese cases (BMI≥30 kg/m2), and 54,412 healthy controls. We confirmed our results in an additional 2,881 lean cases, 8,702 obese cases, and 18,957 healthy controls. Using these data we found differences in genetic enrichment between lean and obese cases, supporting our original hypothesis. We also searched for genetic variants that may be risk factors only in lean or obese patients and found two novel gene regions not previously reported in European individuals. These findings may influence future study design for type 2 diabetes and provide further insight into the biology of the disease.
doi:10.1371/journal.pgen.1002741
PMCID: PMC3364960  PMID: 22693455
10.  Meta-Analysis Increases to 71 the Tally of Confirmed Crohn’s Disease Susceptibility Loci 
Franke, Andre | McGovern, Dermot P.B. | Barrett, Jeffrey C. | Wang, Kai | Radford-Smith, Graham L. | Ahmad, Tariq | Lees, Charlie W. | Balschun, Tobias | Lee, James | Roberts, Rebecca | Anderson, Carl A. | Bis, Joshua C. | Bumpstead, Suzanne | Ellinghaus, David | Festen, Eleonora M. | Georges, Michel | Haritunians, Talin | Jostins, Luke | Latiano, Anna | Mathew, Christopher G. | Montgomery, Grant W. | Prescott, Natalie J. | Rotter, Jerome I. | Schumm, Philip | Sharma, Yashoda | Simms, Lisa A. | Taylor, Kent D. | Whiteman, David | Wijmenga, Cisca | Baldassano, Robert N. | Barclay, Murray | Bayless, Theodore M. | Brand, Stephan | Buning, Carsten | Cohen, Albert | Colombel, Jean-Frederick | Cottone, Mario | Stronati, Laura | Denson, Ted | De Vos, Martine | D’Inca, Renata | Dubinsky, Marla | Edwards, Cathryn | Florin, Tim | Franchimont, Denis | Gearry, Richard | Glas, Jürgen | Van Gossum, Andre | Guthery, Stephen L. | Halfvarson, Jonas | Hommes, Daan | Hugot, Jean-Pierre | Laukens, Debby | Lawrance, Ian | Lemann, Marc | Levine, Arie | Libioulle, Cecile | Louis, Edouard | Mowat, Craig | Newman, William | Panés, Julián | Phillips, Anne | Proctor, Deborah D. | Regueiro, Miguel | Rutgeerts, Paul | Sanderson, Jeremy | Sans, Miquel | Seibold, Frank | Steinhart, A. Hillary | Stokkers, Pieter C.F. | Torkvist, Leif | Ublick, Gerd Kullak | Raychaudhuri, Soumya | Green, Todd | Walters, Thomas | Targan, Stephan R. | Brant, Steven R. | Rioux, John D. | D’Amato, Mauro | Weersma, Rinse | Kugathasan, Subra | Griffiths, Anne M. | Mansfield, John C. | Vermeire, Severine | Duerr, Richard H. | Silverberg, Mark S. | Satsangi, Jack | Schreiber, Stefan | Cho, Judy H. | Annese, Vito | Hakonarson, Hakon | Daly, Mark J. | Parkes, Miles
Nature genetics  2010;42(12):1118-1125.
doi:10.1038/ng.717
PMCID: PMC3299551  PMID: 21102463
11.  Candidate Gene Association Study for Diabetic Retinopathy in Persons with Type 2 Diabetes: The Candidate Gene Association Resource (CARe) 
Cardiovascular disease candidate genes, including genes previously associated with type 2 diabetes and diabetic nephropathy, were not associated with diabetic retinopathy, although a limited number of variants merit further investigation in larger cohorts.
Purpose.
To investigate whether variants in cardiovascular candidate genes, some of which have been previously associated with type 2 diabetes (T2D), diabetic retinopathy (DR), and diabetic nephropathy (DN), are associated with DR in the Candidate gene Association Resource (CARe).
Methods.
Persons with T2D who were enrolled in the study (n = 2691) had fundus photography and genotyping of single nucleotide polymorphisms (SNPs) in 2000 candidate genes. Two case definitions were investigated: Early Treatment Diabetic Retinopathy Study (ETDRS) grades ≥14 and ≥30. The χ2 analyses for each CARe cohort were combined by Cochran-Mantel-Haenszel (CMH) pooling of odds ratios (ORs) and corrected for multiple hypothesis testing. Logistic regression was performed with adjustment for other DR risk factors. Results from replication in independent cohorts were analyzed with CMH meta-analysis methods.
Results.
Among 39 genes previously associated with DR, DN, or T2D, three SNPs in P-selectin (SELP) were associated with DR. The strongest association was to rs6128 (OR = 0.43, P = 0.0001, after Bonferroni correction). These associations remained significant after adjustment for DR risk factors. Among other genes examined, several variants were associated with DR with significant P values, including rs6856425 tagging α-l-iduronidase (IDUA) (P = 2.1 × 10−5, after Bonferroni correction). However, replication in independent cohorts did not reveal study-wide significant effects. The P values after replication were 0.55 and 0.10 for rs6128 and rs6856425, respectively.
Conclusions.
Genes associated with DN, T2D, and vascular diseases do not appear to be consistently associated with DR. A few genetic variants associated with DR, particularly those in SELP and near IDUA, should be investigated in additional DR cohorts.
doi:10.1167/iovs.11-7510
PMCID: PMC3183981  PMID: 21873659
12.  Access to RNA Encapsidated in the Nucleocapsid of Vesicular Stomatitis Virus▿  
Journal of Virology  2010;85(6):2714-2722.
The genomic RNA of negative-strand RNA viruses, such as vesicular stomatitis virus (VSV), is completely enwrapped by the nucleocapsid protein (N) in every stage of virus infection. During viral transcription/replication, however, the genomic RNA in the nucleocapsid must be accessible by the virus-encoded RNA-dependent RNA polymerase in order to serve as the template for RNA synthesis. With the VSV nucleocapsid and a nucleocapsid-like particle (NLP) produced in Escherichia coli, we have found that the RNA in the VSV nucleocapsid can be removed by RNase A, in contrast to what was previously reported. Removal of the RNA did not disrupt the assembly of the N protein, resulting in an empty capsid. Polyribonucleotides were reencapsidated into the empty NLP, and the crystal structures were determined. The crystal structures revealed variable degrees of association of the N protein with a specific RNA sequence.
doi:10.1128/JVI.01927-10
PMCID: PMC3067934  PMID: 21177817
13.  Twelve type 2 diabetes susceptibility loci identified through large-scale association analysis 
Voight, Benjamin F | Scott, Laura J | Steinthorsdottir, Valgerdur | Morris, Andrew P | Dina, Christian | Welch, Ryan P | Zeggini, Eleftheria | Huth, Cornelia | Aulchenko, Yurii S | Thorleifsson, Gudmar | McCulloch, Laura J | Ferreira, Teresa | Grallert, Harald | Amin, Najaf | Wu, Guanming | Willer, Cristen J | Raychaudhuri, Soumya | McCarroll, Steve A | Langenberg, Claudia | Hofmann, Oliver M | Dupuis, Josée | Qi, Lu | Segrè, Ayellet V | van Hoek, Mandy | Navarro, Pau | Ardlie, Kristin | Balkau, Beverley | Benediktsson, Rafn | Bennett, Amanda J | Blagieva, Roza | Boerwinkle, Eric | Bonnycastle, Lori L | Boström, Kristina Bengtsson | Bravenboer, Bert | Bumpstead, Suzannah | Burtt, Noisël P | Charpentier, Guillaume | Chines, Peter S | Cornelis, Marilyn | Couper, David J | Crawford, Gabe | Doney, Alex S F | Elliott, Katherine S | Elliott, Amanda L | Erdos, Michael R | Fox, Caroline S | Franklin, Christopher S | Ganser, Martha | Gieger, Christian | Grarup, Niels | Green, Todd | Griffin, Simon | Groves, Christopher J | Guiducci, Candace | Hadjadj, Samy | Hassanali, Neelam | Herder, Christian | Isomaa, Bo | Jackson, Anne U | Johnson, Paul R V | Jørgensen, Torben | Kao, Wen H L | Klopp, Norman | Kong, Augustine | Kraft, Peter | Kuusisto, Johanna | Lauritzen, Torsten | Li, Man | Lieverse, Aloysius | Lindgren, Cecilia M | Lyssenko, Valeriya | Marre, Michel | Meitinger, Thomas | Midthjell, Kristian | Morken, Mario A | Narisu, Narisu | Nilsson, Peter | Owen, Katharine R | Payne, Felicity | Perry, John R B | Petersen, Ann-Kristin | Platou, Carl | Proença, Christine | Prokopenko, Inga | Rathmann, Wolfgang | Rayner, N William | Robertson, Neil R | Rocheleau, Ghislain | Roden, Michael | Sampson, Michael J | Saxena, Richa | Shields, Beverley M | Shrader, Peter | Sigurdsson, Gunnar | Sparsø, Thomas | Strassburger, Klaus | Stringham, Heather M | Sun, Qi | Swift, Amy J | Thorand, Barbara | Tichet, Jean | Tuomi, Tiinamaija | van Dam, Rob M | van Haeften, Timon W | van Herpt, Thijs | van Vliet-Ostaptchouk, Jana V | Walters, G Bragi | Weedon, Michael N | Wijmenga, Cisca | Witteman, Jacqueline | Bergman, Richard N | Cauchi, Stephane | Collins, Francis S | Gloyn, Anna L | Gyllensten, Ulf | Hansen, Torben | Hide, Winston A | Hitman, Graham A | Hofman, Albert | Hunter, David J | Hveem, Kristian | Laakso, Markku | Mohlke, Karen L | Morris, Andrew D | Palmer, Colin N A | Pramstaller, Peter P | Rudan, Igor | Sijbrands, Eric | Stein, Lincoln D | Tuomilehto, Jaakko | Uitterlinden, Andre | Walker, Mark | Wareham, Nicholas J | Watanabe, Richard M | Abecasis, Gonçalo R | Boehm, Bernhard O | Campbell, Harry | Daly, Mark J | Hattersley, Andrew T | Hu, Frank B | Meigs, James B | Pankow, James S | Pedersen, Oluf | Wichmann, H-Erich | Barroso, Inês | Florez, Jose C | Frayling, Timothy M | Groop, Leif | Sladek, Rob | Thorsteinsdottir, Unnur | Wilson, James F | Illig, Thomas | Froguel, Philippe | van Duijn, Cornelia M | Stefansson, Kari | Altshuler, David | Boehnke, Michael | McCarthy, Mark I
Nature genetics  2010;42(7):579-589.
By combining genome-wide association data from 8,130 individuals with type 2 diabetes (T2D) and 38,987 controls of European descent and following up previously unidentified meta-analysis signals in a further 34,412 cases and 59,925 controls, we identified 12 new T2D association signals with combinedP < 5 × 10−8. These include a second independent signal at the KCNQ1 locus; the first report, to our knowledge, of an X-chromosomal association (near DUSP9); and a further instance of overlap between loci implicated in monogenic and multifactorial forms of diabetes (at HNF1A). The identified loci affect both beta-cell function and insulin action, and, overall, T2D association signals show evidence of enrichment for genes involved in cell cycle regulation. We also show that a high proportion of T2D susceptibility loci harbor independent association signals influencing apparently unrelated complex traits.
doi:10.1038/ng.609
PMCID: PMC3080658  PMID: 20581827
14.  A Meta-Analysis of Genome-Wide Association Scans Identifies IL18RAP, PTPN2, TAGAP, and PUS10 As Shared Risk Loci for Crohn's Disease and Celiac Disease 
PLoS Genetics  2011;7(1):e1001283.
Crohn's disease (CD) and celiac disease (CelD) are chronic intestinal inflammatory diseases, involving genetic and environmental factors in their pathogenesis. The two diseases can co-occur within families, and studies suggest that CelD patients have a higher risk to develop CD than the general population. These observations suggest that CD and CelD may share common genetic risk loci. Two such shared loci, IL18RAP and PTPN2, have already been identified independently in these two diseases. The aim of our study was to explicitly identify shared risk loci for these diseases by combining results from genome-wide association study (GWAS) datasets of CD and CelD. Specifically, GWAS results from CelD (768 cases, 1,422 controls) and CD (3,230 cases, 4,829 controls) were combined in a meta-analysis. Nine independent regions had nominal association p-value <1.0×10−5 in this meta-analysis and showed evidence of association to the individual diseases in the original scans (p-value <1×10−2 in CelD and <1×10−3 in CD). These include the two previously reported shared loci, IL18RAP and PTPN2, with p-values of 3.37×10−8 and 6.39×10−9, respectively, in the meta-analysis. The other seven had not been reported as shared loci and thus were tested in additional CelD (3,149 cases and 4,714 controls) and CD (1,835 cases and 1,669 controls) cohorts. Two of these loci, TAGAP and PUS10, showed significant evidence of replication (Bonferroni corrected p-values <0.0071) in the combined CelD and CD replication cohorts and were firmly established as shared risk loci of genome-wide significance, with overall combined p-values of 1.55×10−10 and 1.38×10−11 respectively. Through a meta-analysis of GWAS data from CD and CelD, we have identified four shared risk loci: PTPN2, IL18RAP, TAGAP, and PUS10. The combined analysis of the two datasets provided the power, lacking in the individual GWAS for single diseases, to detect shared loci with a relatively small effect.
Author Summary
Celiac disease and Crohn's disease are both chronic inflammatory diseases of the digestive tract. Both of these diseases are complex genetic traits with multiple genetic and non-genetic risk factors. Recent genome-wide association (GWA) studies have identified some of the genetic risk factors for these diseases. Interestingly, in addition to some similarities in phenotype, these studies have shown that CelD and CD share some genetic risk factors. Specifically, by comparing the results of independent GWA studies of CD and CelD, two genetic risk loci were found in common: the PTPN2 locus and the IL18RAP locus. Therefore, in order to directly test for additional shared genetic risk factors, we combined the GWA results from two large studies of CelD and CD, essentially creating a combined phenotype with anyone with CD or CelD being coded as affected. Association results were then replicated in additional cohorts of CelD and CD. It is expected that shared risk loci should show association in this analysis, whereas the signal of risk loci specific to either of the two diseases should be diluted. With this method of meta-analysis, we identified next to PTPN2 and IL18 RAP two loci harbouring TAGAP and PUS10 as shared risk loci for Crohn's disease and celiac disease at genome-wide significance.
doi:10.1371/journal.pgen.1001283
PMCID: PMC3029251  PMID: 21298027
15.  Correction: Common Genetic Variants and Modification of Penetrance of BRCA2-Associated Breast Cancer 
Gaudet, Mia M. | Kirchhoff, Tomas | Green, Todd | Vijai, Joseph | Korn, Joshua M. | Guiducci, Candace | Segrè, Ayellet V. | McGee, Kate | McGuffog, Lesley | Kartsonaki, Christiana | Morrison, Jonathan | Healey, Sue | Sinilnikova, Olga M. | Stoppa-Lyonnet, Dominique | Mazoyer, Sylvie | Gauthier-Villars, Marion | Sobol, Hagay | Longy, Michel | Frenay, Marc | GEMO Study Collaborators,  | Hogervorst, Frans B. L. | Rookus, Matti A. | Collée, J. Margriet | Hoogerbrugge, Nicoline | van Roozendaal, Kees E. P. | Piedmonte, Marion | Rubinstein, Wendy | Nerenstone, Stacy | Van Le, Linda | Blank, Stephanie V. | Caldés, Trinidad | de la Hoya, Miguel | Nevanlinna, Heli | Aittomäki, Kristiina | Lazaro, Conxi | Blanco, Ignacio | Arason, Adalgeir | Johannsson, Oskar T. | Barkardottir, Rosa B. | Devilee, Peter | Olopade, Olofunmilayo I. | Neuhausen, Susan L. | Wang, Xianshu | Fredericksen, Zachary S. | Peterlongo, Paolo | Manoukian, Siranoush | Barile, Monica | Viel, Alessandra | Radice, Paolo | Phelan, Catherine M. | Narod, Steven | Rennert, Gad | Lejbkowicz, Flavio | Flugelman, Anath | Andrulis, Irene L. | Glendon, Gord | Ozcelik, Hilmi | Toland, Amanda E. | Montagna, Marco | D'Andrea, Emma | Friedman, Eitan | Laitman, Yael | Borg, Ake | Beattie, Mary | Ramus, Susan J. | Domchek, Susan M. | Nathanson, Katherine L. | Rebbeck, Tim | Spurdle, Amanda B. | Chen, Xiaoqing | Holland, Helene | John, Esther M. | Hopper, John L. | Buys, Saundra S. | Daly, Mary B. | Southey, Melissa C. | Terry, Mary Beth | Tung, Nadine | Overeem Hansen, Thomas V. | Nielsen, Finn C. | Greene, Mark H. | Mai, Phuong L. | Osorio, Ana | Durán, Mercedes | Andres, Raquel | Benítez, Javier | Weitzel, Jeffrey N. | Garber, Judy | Hamann, Ute | Peock, Susan | Cook, Margaret | Oliver, Clare | Frost, Debra | Platte, Radka | Evans, D. Gareth | Lalloo, Fiona | Eeles, Ros | Izatt, Louise | Walker, Lisa | Eason, Jacqueline | Barwell, Julian | Godwin, Andrew K. | Schmutzler, Rita K. | Wappenschmidt, Barbara | Engert, Stefanie | Arnold, Norbert | Gadzicki, Dorothea | Dean, Michael | Gold, Bert | Klein, Robert J. | Couch, Fergus J. | Chenevix-Trench, Georgia | Easton, Douglas F. | Daly, Mark J. | Antoniou, Antonis C. | Altshuler, David M. | Offit, Kenneth
PLoS Genetics  2010;6(11):10.1371/annotation/59ea8540-4e63-4f4a-a79e-f68765fdeac7.
doi:10.1371/annotation/59ea8540-4e63-4f4a-a79e-f68765fdeac7
PMCID: PMC2988688
16.  Correction: Common Genetic Variants and Modification of Penetrance of BRCA2-Associated Breast Cancer 
Gaudet, Mia M. | Kirchhoff, Tomas | Green, Todd | Vijai, Joseph | Korn, Joshua M. | Guiducci, Candace | Segrè, Ayellet V. | McGee, Kate | McGuffog, Lesley | Kartsonaki, Christiana | Morrison, Jonathan | Healey, Sue | Sinilnikova, Olga M. | Stoppa-Lyonnet, Dominique | Mazoyer, Sylvie | Gauthier-Villars, Marion | Sobol, Hagay | Longy, Michel | Frenay, Marc | GEMO Study Collaborators,  | Hogervorst, Frans B. L. | Rookus, Matti A. | Collée, J. Margriet | Hoogerbrugge, Nicoline | van Roozendaal, Kees E. P. | Piedmonte, Marion | Rubinstein, Wendy | Nerenstone, Stacy | Van Le, Linda | Blank, Stephanie V. | Caldés, Trinidad | de la Hoya, Miguel | Nevanlinna, Heli | Aittomäki, Kristiina | Lazaro, Conxi | Blanco, Ignacio | Arason, Adalgeir | Johannsson, Oskar T. | Barkardottir, Rosa B. | Devilee, Peter | Olopade, Olofunmilayo I. | Neuhausen, Susan L. | Wang, Xianshu | Fredericksen, Zachary S. | Peterlongo, Paolo | Manoukian, Siranoush | Barile, Monica | Viel, Alessandra | Radice, Paolo | Phelan, Catherine M. | Narod, Steven | Rennert, Gad | Lejbkowicz, Flavio | Flugelman, Anath | Andrulis, Irene L. | Glendon, Gord | Ozcelik, Hilmi | Toland, Amanda E. | Montagna, Marco | D'Andrea, Emma | Friedman, Eitan | Laitman, Yael | Borg, Ake | Beattie, Mary | Ramus, Susan J. | Domchek, Susan M. | Nathanson, Katherine L. | Rebbeck, Tim | Spurdle, Amanda B. | Chen, Xiaoqing | Holland, Helene | John, Esther M. | Hopper, John L. | Buys, Saundra S. | Daly, Mary B. | Southey, Melissa C. | Terry, Mary Beth | Tung, Nadine | Overeem Hansen, Thomas V. | Nielsen, Finn C. | Greene, Mark I. | Mai, Phuong L. | Osorio, Ana | Durán, Mercedes | Andres, Raquel | Benítez, Javier | Weitzel, Jeffrey N. | Garber, Judy | Hamann, Ute | Peock, Susan | Cook, Margaret | Oliver, Clare | Frost, Debra | Platte, Radka | Evans, D. Gareth | Lalloo, Fiona | Eeles, Ros | Izatt, Louise | Walker, Lisa | Eason, Jacqueline | Barwell, Julian | Godwin, Andrew K. | Schmutzler, Rita K. | Wappenschmidt, Barbara | Engert, Stefanie | Arnold, Norbert | Gadzicki, Dorothea | Dean, Michael | Gold, Bert | Klein, Robert J. | Couch, Fergus J. | Chenevix-Trench, Georgia | Easton, Douglas F. | Daly, Mark J. | Antoniou, Antonis C. | Altshuler, David M. | Offit, Kenneth
PLoS Genetics  2010;6(11):10.1371/annotation/b28cf02d-7196-4a16-8b36-6562a0b84f75.
doi:10.1371/annotation/b28cf02d-7196-4a16-8b36-6562a0b84f75
PMCID: PMC2999983
17.  Common Genetic Variants and Modification of Penetrance of BRCA2-Associated Breast Cancer 
Gaudet, Mia M. | Kirchhoff, Tomas | Green, Todd | Vijai, Joseph | Korn, Joshua M. | Guiducci, Candace | Segrè, Ayellet V. | McGee, Kate | McGuffog, Lesley | Kartsonaki, Christiana | Morrison, Jonathan | Healey, Sue | Sinilnikova, Olga M. | Stoppa-Lyonnet, Dominique | Mazoyer, Sylvie | Gauthier-Villars, Marion | Sobol, Hagay | Longy, Michel | Frenay, Marc | GEMO Study Collaborators,  | Hogervorst, Frans B. L. | Rookus, Matti A. | Collée, J. Margriet | Hoogerbrugge, Nicoline | van Roozendaal, Kees E. P. | Piedmonte, Marion | Rubinstein, Wendy | Nerenstone, Stacy | Van Le, Linda | Blank, Stephanie V. | Caldés, Trinidad | de la Hoya, Miguel | Nevanlinna, Heli | Aittomäki, Kristiina | Lazaro, Conxi | Blanco, Ignacio | Arason, Adalgeir | Johannsson, Oskar T. | Barkardottir, Rosa B. | Devilee, Peter | Olopade, Olofunmilayo I. | Neuhausen, Susan L. | Wang, Xianshu | Fredericksen, Zachary S. | Peterlongo, Paolo | Manoukian, Siranoush | Barile, Monica | Viel, Alessandra | Radice, Paolo | Phelan, Catherine M. | Narod, Steven | Rennert, Gad | Lejbkowicz, Flavio | Flugelman, Anath | Andrulis, Irene L. | Glendon, Gord | Ozcelik, Hilmi | Toland, Amanda E. | Montagna, Marco | D'Andrea, Emma | Friedman, Eitan | Laitman, Yael | Borg, Ake | Beattie, Mary | Ramus, Susan J. | Domchek, Susan M. | Nathanson, Katherine L. | Rebbeck, Tim | Spurdle, Amanda B. | Chen, Xiaoqing | Holland, Helene | John, Esther M. | Hopper, John L. | Buys, Saundra S. | Daly, Mary B. | Southey, Melissa C. | Terry, Mary Beth | Tung, Nadine | Overeem Hansen, Thomas V. | Nielsen, Finn C. | Greene, Mark I. | Mai, Phuong L. | Osorio, Ana | Durán, Mercedes | Andres, Raquel | Benítez, Javier | Weitzel, Jeffrey N. | Garber, Judy | Hamann, Ute | Peock, Susan | Cook, Margaret | Oliver, Clare | Frost, Debra | Platte, Radka | Evans, D. Gareth | Lalloo, Fiona | Eeles, Ros | Izatt, Louise | Walker, Lisa | Eason, Jacqueline | Barwell, Julian | Godwin, Andrew K. | Schmutzler, Rita K. | Wappenschmidt, Barbara | Engert, Stefanie | Arnold, Norbert | Gadzicki, Dorothea | Dean, Michael | Gold, Bert | Klein, Robert J. | Couch, Fergus J. | Chenevix-Trench, Georgia | Easton, Douglas F. | Daly, Mark J. | Antoniou, Antonis C. | Altshuler, David M. | Offit, Kenneth
PLoS Genetics  2010;6(10):e1001183.
The considerable uncertainty regarding cancer risks associated with inherited mutations of BRCA2 is due to unknown factors. To investigate whether common genetic variants modify penetrance for BRCA2 mutation carriers, we undertook a two-staged genome-wide association study in BRCA2 mutation carriers. In stage 1 using the Affymetrix 6.0 platform, 592,163 filtered SNPs genotyped were available on 899 young (<40 years) affected and 804 unaffected carriers of European ancestry. Associations were evaluated using a survival-based score test adjusted for familial correlations and stratified by country of the study and BRCA2*6174delT mutation status. The genomic inflation factor (λ) was 1.011. The stage 1 association analysis revealed multiple variants associated with breast cancer risk: 3 SNPs had p-values<10−5 and 39 SNPs had p-values<10−4. These variants included several previously associated with sporadic breast cancer risk and two novel loci on chromosome 20 (rs311499) and chromosome 10 (rs16917302). The chromosome 10 locus was in ZNF365, which contains another variant that has recently been associated with breast cancer in an independent study of unselected cases. In stage 2, the top 85 loci from stage 1 were genotyped in 1,264 cases and 1,222 controls. Hazard ratios (HR) and 95% confidence intervals (CI) for stage 1 and 2 were combined and estimated using a retrospective likelihood approach, stratified by country of residence and the most common mutation, BRCA2*6174delT. The combined per allele HR of the minor allele for the novel loci rs16917302 was 0.75 (95% CI 0.66–0.86, ) and for rs311499 was 0.72 (95% CI 0.61–0.85, ). FGFR2 rs2981575 had the strongest association with breast cancer risk (per allele HR = 1.28, 95% CI 1.18–1.39, ). These results indicate that SNPs that modify BRCA2 penetrance identified by an agnostic approach thus far are limited to variants that also modify risk of sporadic BRCA2 wild-type breast cancer.
Author Summary
The risk of breast cancer associated with BRCA2 mutations varies widely. To determine whether common genetic variants modify the penetrance of BRCA2 mutations, we conducted the first genome-wide association study of breast cancer among women with BRCA2 mutations using a two-stage approach. The major finding of the study is that only those loci known to be associated with breast cancer risk in the general population, including FGFR2 (rs2981575), modified BRCA2-associated risk in our high-risk population. Two novel loci, on chromosomes 10 in ZNF365 (rs16917302) and chromosome 20 (rs311499), were shown to modify risk in BRCA2 mutation carriers, although not at a genome-wide level of significance. However, the ZNF365 locus has recently independently been associated with breast cancer risk in sporadic tumors, highlighting the potential significance of this zinc finger-containing gene in breast cancer pathogenesis. Our results indicate that it is unlikely that other common variants have a strong modifying effect on BRCA2 penetrance.
doi:10.1371/journal.pgen.1001183
PMCID: PMC2965747  PMID: 21060860
18.  CryoEM Model of the Bullet-Shaped Vesicular Stomatitis Virus 
Science (New York, N.Y.)  2010;327(5966):689-693.
Vesicular stomatitis virus (VSV) is a bullet-shaped rhabdovirus and a model system of negative-strand RNA viruses. Based on direct visualization by cryo-electron microscopy, we show that each virion contains two nested, left-handed helices, an outer helix of matrix protein M and an inner helix of nucleoprotein N and RNA. M has a hub domain with four contact sites that link to neighboring M and N subunits, providing rigidity by clamping adjacent turns of the nucleocapsid. Side-by-side interactions between neighboring N subunits are critical for the nucleocapsid to form a bullet shape, and structure-based mutagenesis results support this description. Together, our data suggest a mechanism of VSV assembly in which the nucleocapsid spirals from the tip to become the helical trunk, both subsequently framed and rigidified by the M layer.
doi:10.1126/science.1181766
PMCID: PMC2892700  PMID: 20133572
19.  Characterization of a Mumps Virus Nucleocapsidlike Particle ▿  
Journal of Virology  2009;83(21):11402-11406.
The nucleocapsid protein (NP) of mumps virus (MuV), a paramyxovirus, was coexpressed with the phosphoprotein (P) in Escherichia coli. The NP and P proteins form a soluble complex containing RNA. Under a transmission electron microscope, the NP-RNA complex appears as a nucleocapsidlike ring that has a diameter of approximately 20 nm with 13 subunits. There is a piece of single-stranded RNA with a length of 78 nucleotides in the NP-RNA ring. Shorter RNA pieces are also visible. The MuV NP protein may provide weaker protection of the RNA than the NP protein of some other negative-strand RNA viruses, reflecting the degree of NP protein association.
doi:10.1128/JVI.00504-09
PMCID: PMC2772791  PMID: 19692473
20.  Genome-wide association study identifies five novel susceptibility loci for Crohn's disease and implicates a role for autophagy in disease pathogenesis. 
Nature genetics  2007;39(5):596-604.
We present a genome-wide association study of ileal Crohn's disease (CD) and two independent replication studies that identify five novel regions of association to CD. Specifically, in addition to the previously established CARD15 and IL23R associations, we report strong associations with independent replication to variation in the genomic regions encoding the PHOX2B, NCF4 and ATG16L1 genes, as well as a predicted gene on 16q24.1 (FAM92B) and an intergenic region on 10q21.1. We further demonstrate that the ATG16L1 gene is expressed in intestinal epithelial cell lines and that functional knock down of this gene abrogates autophagy of Salmonella typhimurium. Together these findings suggest that autophagy and host cell responses to intra-cellular microbes are involved in the pathogenesis of CD.
doi:10.1038/ng2032
PMCID: PMC2757939  PMID: 17435756
21.  Genome-wide association defines more than thirty distinct susceptibility loci for Crohn's disease 
Nature genetics  2008;40(8):955-962.
Several new risk factors for Crohn's disease have been identified in recent genome-wide association studies. To advance gene discovery further we have combined the data from three studies (a total of 3,230 cases and 4,829 controls) and performed replication in 3,664 independent cases with a mixture of population-based and family-based controls. The results strongly confirm 11 previously reported loci and provide genome-wide significant evidence for 21 new loci, including the regions containing STAT3, JAK2, ICOSLG, CDKAL1, and ITLN1. The expanded molecular understanding of the basis of disease offers promise for informed therapeutic development.
doi:10.1038/NG.175
PMCID: PMC2574810  PMID: 18587394
22.  A high resolution HLA and SNP haplotype map for disease association studies in the extended human MHC 
Nature genetics  2006;38(10):1166-1172.
The proteins encoded by the classical HLA class I and class II genes in the major histocompatibility complex (MHC) are highly polymorphic and play an essential role in self/non-self immune recognition. HLA variation is a crucial determinant of transplant rejection and susceptibility to a large number of infectious and autoimmune disease1. Yet identification of causal variants is problematic due to linkage disequilibrium (LD) that extends across multiple HLA and non-HLA genes in the MHC2,3. We therefore set out to characterize the LD patterns between the highly polymorphic HLA genes and background variation by typing the classical HLA genes and >7,500 common single nucleotide polymorphisms (SNPs) and deletion/insertion polymorphisms (DIPs) across four population samples. The analysis provides informative tag SNPs that capture some of the variation in the MHC region and that could be used in initial disease association studies, and provides new insight into the evolutionary dynamics and ancestral origins of the HLA loci and their haplotypes.
doi:10.1038/ng1885
PMCID: PMC2670196  PMID: 16998491
23.  MAST3: a Novel IBD Risk Factor that Modulates TLR4 Signaling 
Genes and immunity  2008;9(7):602-612.
Inflammatory bowel disease (IBD) is a chronic disorder caused by multiple factors in a genetically susceptible host. Significant advances in the study of genetic susceptibility have highlighted the importance of the innate immune system in this disease. We previously completed a genomewide linkage study and found a significant locus (IBD6) on chromosome 19p. We were interested in identifying the causal variant in IBD6. We performed a two-stage association mapping study. In stage one, 1530 SNPs were selected from the HapMap database and genotyped in 761 patients with IBD. Among the SNPs that passed the threshold for replication, 26 were successfully genotyped in 754 additional patients (stage two). One intronic variant, rs273506 located in the MAST3 gene was found to be associated in both stages (pooled P=2×10−4). We identified four MAST3 coding variants, including a non-synonymous SNP rs8108738, correlated to rs273506 and associated to IBD. To test whether MAST3 was expressed in cells of interest, we performed expression assays which showed abundant expression of MAST3 in antigen presenting cells and in lymphocytes. The knockdown of MAST3 specifically decreased TLR4 dependent NF-κB activity. Our findings are additional proof of the pivotal role played by modulators of NF-κB activity in IBD pathogenesis.
doi:10.1038/gene.2008.57
PMCID: PMC2574831  PMID: 18650832
MAST3; Inflammatory Bowel Disease; TLR4; NF-κB
24.  Conserved characteristics of the rhabdovirus nucleoprotein 
Virus research  2007;129(2):246-251.
Rhabdovirus is a negative strand RNA virus that packages a ribonucleorptein (RNP) complex. The RNP is composed of a genome that is encapsidated completely by the nucleoprotein (N). Structural comparisons of the RNA-nucleoprotein complexes from two members, vesicular stomatitis virus (VSV) and rabies virus (RABV), revealed highly conserved characteristics of folding, RNA binding, and assembly despite their lack of significant homology in amino acid sequence. The RNA binding cavity is located between two conserved domains formed by α–helices, but the positively charged residues that coordinate with the phosphate groups are at different sites. The intermolecular interactions among N molecules have a conserved pattern that is rendered, however, by different residues. The curvature of the RABV N-RNA complex in the crystal structure is larger than that of the VSV N-RNA complex. The more relaxed curvature allows the bases in the RNA to stack more tightly, and at the same time, the helices near the C-terminus move into the created space in order to cover the bound RNA. This may explain how the RNP can adopt different conformations from being packed as a superhelix in the virion to a relaxed linear structure once being delivered into the cytoplasm.
doi:10.1016/j.virusres.2007.07.011
PMCID: PMC2082134  PMID: 17764775
Negative strand RNA virus; ribonucleoprotein; assembly; RNA encapsidation; nucleoprotein; conserved structure
25.  Role of Intermolecular Interactions of Vesicular Stomatitis Virus Nucleoprotein in RNA Encapsidation▿  
Journal of Virology  2007;82(2):674-682.
The crystal structure of the vesicular stomatitis virus nucleoprotein (N) in complex with RNA reveals extensive and specific intermolecular interactions among the N molecules in the 10-member oligomer. What roles these interactions play in encapsidating RNA was studied by mutagenesis of the N protein. Three N mutants intended for disruption of the intermolecular interactions were designed and coexpressed with the phosphoprotein (P) in an Escherichia coli system previously described (T. J. Green et al., J. Virol. 74:9515-9524, 2000). Mutants N (Δ1-22), N (Δ347-352), and N (320-324, (Ala)5) lost RNA encapsidation and oligomerization but still bound with P. Another mutant, N (Ser290→Trp), was able to form a stable ring-like N oligomer and bind with the P protein but was no longer able to encapsidate RNA. The crystal structure of N (Ser290→Trp) at 2.8 Å resolution showed that this mutant can maintain all the same intermolecular interactions as the wild-type N except for a slight unwinding of the N-terminal lobe. These results suggest that the intermolecular contacts among the N molecules are required for encapsidation of the viral RNA.
doi:10.1128/JVI.00935-07
PMCID: PMC2224587  PMID: 18003727

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