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Bardet-Biedl syndrome (BBS) is an autosomal recessive disorder characterized by obesity, retinitis pigmentosa, polydactyly, mental retardation and hypogonadism. To date, twelve BBS genes have been identified, however, the precise functions of BBS genes are yet to be unraveled. To unravel the pathophysiology of BBS, we developed the zebrafish BBS model. In the zebrafish animal model system, we performed loss-of-function analysis using antisense morpholino oligonucleotides (MO). We observed strikingly overlapping phenotypes in the MO-injected embryos (morphants) including: Kupffer's vesicle (KV) disruption, a premature loss of KV cilia leading to alterations in left-right asymmetry and altered intracellular trafficking of organelles (melanosome). These phenotypes can be rescued by co-injection of in vitro synthesized wild-type RNA demonstrating the specificity of MO knockdown. In contrast to other bbs gene knockdowns, bbs1 knockdown resulted in an additional phenotype: pericardium effusion, which was rescued by co-injection with the bbs1 wild-type RNA and was significantly suppressed by the M390R mutant RNA. In addition, immunohistochemistry revealed that bbs1 wild-type protein localizes to the centrosome and cytoplasm, while bbs1 M390R mutant protein only has cytoplasmic localization. The BBS1 M390R missense mutation is the most common mutation in all BBS cases and the methionine at amino acid position 390 is evolutionarily conserved. Our data suggest that the BBS1 M390R allele is a hypomorphic allele and that the residual function of the mutant protein is necessary for embryonic development. Furthermore, we demonstrated the utility of the zebrafish BBS model in BBS candidate gene identification. Data from linkage analysis, expression profile correlation and mutation screen, suggested that TRIM32 is a BBS gene. Therefore, we identified the zebrafish trim32 gene and performed loss-of-function analysis. MO knockdown of zebrafish trim32 gene expression phenocopied the prototypic zebrafish BBS phenotypes: KV formation defect and a delay in intracellular transport. In addition, these phenotypes were rescued by co-injection of wild-type trim32 and LGMD2H mutant (D487N) RNA, but the RNA with the BBS-associated mutation (P130S) failed to rescue these phenotypes. The functional evidence, combined with data from linkage analysis, expression correlation and mutation screening demonstrate that TRIM32 is a BBS gene (BBS11).

No description available.

Cervical cancer is the second most common cancer in women worldwide. Persistent infection by an oncogenic type of human papillomavirus (HPV) is a necessary but not sufficient cause and there is also a genetic component. This thesis aims to identify host genetic risk factors for cervical cancer based on the hypothesis that susceptibility is affected by genetic variation in the immune response towards HPV infection. Paper I analyzed allergy in sons and cervical cancer in their mothers, and revealed an inverse association between cervical cancer and allergy across generations. Mothers of allergic sons have a lower incidence of cervical cancer, supporting the importance of immunogenetic factors. Paper II investigated the HPV type in 1079 women diagnosed 1965-1993. All women were from families with at least two affected. It appeared that HPV 16 was becoming less common with time. There was no evidence that related women were prone to infection by the same type, indicating that the immunogenetic factors act in a general, rather than an HPV type specific, manner. Paper III and IV analysed the association of candidate genes with susceptibility to cervical cancer in 1306 women with cervical cancer in situ and 288 unrelated controls. Paper III showed the association of variation in the two immune response genes chemokine receptor 2 (CCR-2) and interleukin 4 receptor (IL-4R) with cervical cancer. In paper IV variation at several loci in the MHC region was studied and the importance of the HLA class II locus DQB1 emphasized. This thesis work supports the contribution of genes of the immune system to cervical cancer susceptibility. The genetic risk factors so far identified account for only a part of the genetic susceptibility, which implies that other yet undiscovered variants of importance remain to be identified.

The interferon regulatory factor 5 (IRF5) gene encodes a transcription factor that plays an important role in the innate as well as in the cell-mediated immune response. The IRF5 gene has received considerable attention since it was shown to be associated with two autoimmune diseases; systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). The aim of this thesis was to examine if IRF5 is associated with other autoimmune diseases and to investigate the role of the genetic variation of IRF5. In the first study a set of common polymorphisms in IRF5 were analyzed for their association with two subgroup of inflammatory bowel disease (IBD); Crohn´s diseases (CD) and ulcerative colitis (UC). A strong signal of association of IRF5 with IBD was found. The most strongly associated polymorphism is a 5 base pair (bp) insertion-deletion (indel) in the promoter region of the IRF5 gene. The association was detected within both UC and CD, and appeared to be stronger in UC. In the second study we investigated the association of IRF5 with multiple sclerosis (MS). A similar set of polymorphisms as in the IBD study were genotyped in a cohort of MS patients and controls. The same polymorphisms that were associated with IBD were also found to be associated with MS. In the third study, we performed a comprehensive investigation of the IRF5 gene to detect most of the polymorphisms in the gene, and to determine to what extent they account for the association signals obtained from the gene. IRF5 was sequenced and 34 new polymorphisms were identified. Twenty seven of these, and 20 previously known SNPs in IRF5 were genotyped in an SLE case-control cohort. We found that only two polymorphisms, the 5bp indel and a SNP downstream of IRF5, account for the association signal from all the remaining markers in the IRF5 gene, and that these two polymorphisms are independently associated with SLE. Interestingly, in our studies on IBD and MS, we only observed the signal from the 5bp indel polymorphism as a risk factor for IBDs. In the fourth study the two independent risk alleles in IRF5, were tested for their association with primary Sjögren´s syndrome (pSS). In this study we also included one SNP in the STAT4 gene, since STAT4 had recently been shown to be associated with SLE. Both risk factors in IRF5 and STAT4 were found to be associated with pSS. The regulation of expression of IRF5 was also investigated in the first three studies. We observed allele-specific differences in protein binding as well as increased binding of the transcription factor SP1 to the 5bp risk allele. We also detected increased expression of the IRF5 mRNA from a promoter containing the risk allele. Taken together, the results of our studies suggest a general function for IRF5 as a regulator of the autoimmune response, where the 5bp indel is associated with IBD, MS, SLE and pSS. The additionally polymorphisms, which account for the remaining association signal obtained with SLE and pSS, may contribute to the disease manifestations that are specific for rheumatic diseases. Our studies add to the evidence that there are genes or pathways that are common in multiple autoimmune diseases, and that the type I interferon system is likely to be involved in the development of these diseases.

Thesis (Ph. D.)--York University, 2000. Graduate Programme in History. / Typescript. Includes bibliographical references. Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://wwwlib.umi.com/cr/yorku/fullcit?pNQ56247.

abstract / Paediatrics and Adolescent Medicine / Master / Master of Philosophy

Within the C57BL/Fr inbred mouse strain, microphthalmia, the reduction in eyeball diameter, occurred spontaneously among 14.68% of the newborns. Microphthalmia was commoner and more severe in the right eye and was more frequent among females. Maternal age and parity, month or season of birth and litter size did not influence the microphthalmia frequency. [...]

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Academisch proefschrift--Utrecht. / Bibliography: p. [257]-275.