Autoimmune diseases are endemic, but the disease mechanisms are poorly understood. A way to better understand these are to find disease-regulating genes. However, this is difficult as the diseases are complex, with several genes as well as environmental factors influencing the development of disease. A way to facilitate the search for genes responsible for the diseases is to use comparative genomic studies. Animal models are relatively easy to analyze since control of environment and breeding are obtained. The University of California at Davies – line 200 (UCD-200) chickens have a hereditary disease that is similar to systemic sclerosis. Using a backcross between UCD-200 chickens and red junglefowl (RJF) chickens we identified three loci linked to the disease. The loci contained immune-regulatory genes suggested to be involved in systemic sclerosis in humans, as well as a previously unidentified linkage between systemic sclerosis in UCD-200 chickens and IGFBP3. The Dark brown (Db) gene enhances red pheomelanin and restricts expression of eumelanin in chickens. The Db phenotype is regulated by an 8 kb deletion upstream of SOX10. Pigmentation studies are potentially useful when trying to identify pathogenic mechanisms and candidate genes in vitiligo The Obese strain (OS) of chickens spontaneously develops an autoimmune thyroiditis which closely resembles human Hashimoto’s thyroiditis. By using an intercross between OS chickens and RJF chickens, we found several disease phenotypes that can be used in an ongoing linkage analysis with the goal to find candidate genes for autoimmune disease. An important phenotype to record and add to the linkage analysis is autoantibodies against thyroid peroxidase, since this phenotype is a key feature in Hashimoto’s thyroiditis. Previous attempts to measure these titres in OS chickens have failed, hence an assay was developed for this purpose.

Magister Scientiae - MSc / Traditionally in human genetic linkage analysis, extended families were only used in the analysis of dichotomous traits, such as Disease/No Disease. For quantitative traits, analyses initially focused on data from family trios (for example, mother, father, and child) or sib-pairs. Recently however, there have been two very important developments in genetics: It became clear that if the disease status of several generations of a family is known and their genetic information is obtained, researchers can pinpoint which pieces of genetic material are linked to the disease or trait. It also became evident that if a trait is quantitative (numerical), as blood pressure or viral loads are, rather than dichotomous, one has much more power for the same sample size. This led to the development of statistical mixed models which could incorporate all the features of the data, including the degree of relationship between each pair of family members. This is necessary because a parent-child pair definitely shares half their genetic material, whereas a pair of cousins share, on average, only an eighth. The statistical methods involved here have however been developed by geneticists, for their specific studies, so there does not seem to be a unified and general description of the theory underlying the methods. The aim of this dissertation is to explain in a unified and statistically comprehensive manner, the theory involved in the analysis of quantitative trait genetic data from extended families. The focus is on linkage analysis: what it is and what it aims to do. There is a step-by-step build up to it, starting with an introduction to genetic epidemiology. This includes an explanation of the relevant genetic terminology. There is also an application section where an appropriate human genetic family dataset is analysed, illustrating the methods explained in the theory sections. / South Africa

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近年來以電子資訊產業掛帥的台灣，傳統產業似乎成了被遺忘一個部分。但透過許多相關產業的調查資料顯示，部分傳統產業在全球化的競爭底下，依舊可在台灣立足，汽車業與船舶業就是其中的代表。由產業群聚的觀點來看，產業與其關聯產業在空間中的關係為何，產業是否集中於某些特定的空間單元，皆為值得討論的課題。 為對汽車業及船舶業進行分析，本研究透過產業關聯分析以及地理資訊系統之熱點分析（hot spot analysis），並結合工商普查資料、產業關聯表、生產者投入係數，進行空間集中指標的計算，從時間序列的變化，瞭解不同時間點運輸製造業的主要關聯產業之差異及其群聚版圖的變遷，並且進一步探討運輸製造業產業群聚之水平連結在空間臨接上所產生之差異。 研究結果顯示在1981年至2001年間，汽車業與船舶業在既有的空間單元中保持穩定的成長，僅北部區域的汽車業集中重心由台北移至桃園，此種情況即代表產業群聚的區位惰性。此外，船舶業在空間單元中有集中於鋼鐵業以及港口周邊的情況；汽車業與其主要關聯產業則都集中在桃園新竹一帶。 / In recent years, Taiwan taking electronics and information industries as main development, the traditional industry seems to become a part forgotten. But show through the survey materials of a lot of relevant industries, some traditional industries that under the competition of globalization, can still base on Taiwan, the automobile industry and shipping industry are representatives among them. In the view point of industry clusters, what are the spatial relationship between industry and its related industries, whether the industry concentrates on some specific space units, all in order to worth discussing. In order to analysis the automobile industry and shipping industry, this research passes the industry linkage analysis, hot spot analysis of geographic information system, and combine the industry, commerce and service census, input-output table, input coefficients table at producers' prices, to make the calculation of spatial concentration index, and from the view point of time series, to find out the difference of main related industries, the change of cluster map, and further more, to discuss the spatial relationship between industry and its main related industries. The result of study shows between 1981 and 2001, the automobile industry and shipping industry keeps steady growth with in the space unit that has already had, only the automobile industry of the northern area concentrates center on being transferred from Taipei to Taoyuan, this kind of situation represents the inert of location of industry clusters. In addition, shipping industry centre in nears the steel industry and port; the automobile industry and its main related industries are mostly concentrated in Taoyuan and Hsinchu.

Nous présentons ici la description clinique et génétique d’un syndrome neurocutané unique. Le laboratoire du Dr Cossette a entrepris la caractérisation clinique et génétique d'une famille canadienne-française qui a été identifiée par les Drs Giroux et Barbeau en 1972 et qui comprend plus de 100 personnes sur six générations. Les membres atteints de cette famille présentent des lésions typiques d'érythrokératodermie (EK) (OMIM 133190, EKV1 et EKV2), associées à une ataxie spinocérébelleuse pure. Dans cette famille, l'ataxie est caractérisée par des troubles de la coordination et de la démarche causés par une dégénérescence du cervelet et de la moelle épinière. Cette ataxie est transmise selon un mode autosomique dominant. Une étude antérieure de cette variante d'EK avec ataxie avait suggéré une liaison sur le chromosome 1p34-p35, soit la même région que les formes EKV de type 1 et 2, causées respectivement par des mutations dans les gènes connexin-31 (GJB3; OMIM 603324) et connexin-30.3 (GJB4; OMIM 605425). Cependant, aucune mutation n'a été retrouvée dans ces gènes pour la famille canadienne-française. Nous avons récemment recontacté la famille et effectué des examens détaillés, incluant une imagerie par résonance magnétique (IRM) et un électromyogramme (EMG). Les manifestations neurologiques des individus atteints sont compatibles avec une nouvelle forme d’ataxie cérébelleuse pure à transmission autosomique dominante (ADCA de type III dans la classification de Harding) que nous avons appelée SCA34. Une cartographie complète du génome nous a permis de localiser le gène SCA34 sur le chromosome 6p12.3-q16.2. Également, en collaboration avec les Drs Alexis Brice (Hôpital Pitié-La Salpêtrière, Paris) et Alfredo Brusco (Hôpital San Giovanni Battista di Torino, Italie), nous avons confirmé que trois autres familles européennes avec SCA inexpliquée étaient également liées au locus SCA34. Notre laboratoire a récemment entrepris la recherche des mutations responsables de SCA34. Les résultats de ce criblage de gènes candidats sont présentés dans le chapitre 3 de cette thèse. / We present here the clinical and genetic description of a unique neuro-cutaneous syndrome. Dr. Cossette’s laboratory began the clinical and genetic characterization of a French-Canadian family who was identified by Drs. Giroux and Barbeau in 1972 and includes more than 100 people over six generations. The affected members of this family have typical lesions of erythrokeratodermia (EK) (OMIM 133190, and EKV1 EKV2), associated with pure spinocerebellar ataxia. In this family, the clinical phenotype is characterized by gait ataxia caused by degeneration of the cerebellum and spinal cord and the pattern of inheritance is compatible with an autosomal dominant trait. In a previous study of this variant of ataxia with EK, putative linkage was found on chromosome 1p34-p35, the same chromosomal region of EKV1 and EKV2 that are respectively caused by mutations in the connexin-31 gene (GJB3, OMIM 603324) and connexin -30.3 (GJB4, OMIM 605425). However, no mutations have been found in these latter genes for the French-Canadian family. We recently contacted the family and carried out detailed examinations, including a magnetic resonance imaging (MRI) and electromyography (EMG). Neurological manifestations of affected individuals are consistent with a new form of pure autosomal dominant cerebellar ataxia, (ADCA type III in the classification of Harding) that we named SCA34. A whole genome scan allowed us to map the gene on chromosome 6p12.3-q16.2. Interestingly, in collaboration with Dr. Alexis Brice (Hôpital Pitié-La Salpêtrière, Paris), and Alfredo Brusco (San Giovanni Battista Hospital, Turin, Italy), we found that three additional European families with unexplained SCA were also linked to the SCA34 locus. Our laboratory has recently begun the search for mutations causing SCA34. The results of this screening of candidate genes are presented in Chapter 3 of this thesis.

Thèse réalisée en cotutelle avec l'Université Pierre et Marie Curie, Paris 6(UPMC, Paris, France). / Bien que les bases physiopathologiques de beaucoup de maladies musculaires soient dorénavant connues, les myopathies congénitales à cores (MCs), maladies génétiques qui se présentent dès la naissance avec un retard du développement moteur, une faiblesse musculaire et des complications respiratoires et/ou cardiaques parfois mortelles, demeurent mal comprises. Des mutations dans RYR1, SEPN1, TTN, ACTA1, CFL2 et MEGF10 ont été associées aux MCs, pourtant, dans plus de 50% des cas, le gène responsable reste à identifier. L’objectif de ma thèse a été de clarifier les mécanismes physiopathologiques des MCs par l’identification de nouveaux gènes ou de nouvelles mutations. Cette thèse a eu une dimension internationale concrétisée par la mise en place d’une cotutelle UPMC (France) et UdeM (Québec). J’ai développé deux axes de recherche complémentaires. D’une part j’ai étudié 21 familles informatives avec MC récessive, scoliose et atteinte respiratoire, en combinant clonage positionnel et étude de gènes candidats et en utilisant des outils variés allant du génotypage au séquençage de nouvelle génération (NGS). En parallèle, j’ai étudié 24 familles avec une MC autosomique récessive affectant les muscles cardiaque et squelettiques et dont le phénotype était semblable à celui observé chez des patients avec des délétions dans les 6 derniers exons de TTN. Ainsi pour l'analyse de cette deuxième cohorte, nous avons appliqué une stratégie de séquençage de gène candidat ciblée sur ces exons et de NGS pour le reste du gène. Pendant mon doctorat j'ai identifié les défauts génétiques de 8 des 45 familles étudiées (18 %), et caractérisé 3 nouvelles entités médicales, dont deux MCs dues à des nouvelles mutations de TTN. Ces résultats ont servi à l’identification de nouvelles interactions protéiques de la titine et contribuent à définir TTN comme une cause majeure de pathologies musculaires cardiaques et/ou squelettiques. Une troisième nouvelle forme de MC est provoquée par une mutation d'un coactivateur transcriptionnel peu connu et jamais associé à une maladie. Ces résultats ont révélé un nouvel acteur clef et une nouvelle voie de signalisation dans la physiopathologie du muscle, ont eu un bénéfice direct en termes de conseil génétique et ouvrent la voie pour le développement de thérapies. / While the pathophysiological bases of many muscular diseases are nowadays well known, congenital core myopathies (CMs) remain poorly understood. CMs are genetic diseases which generally present at birth with delayed motor development, muscle weakness, and sometimes fatal respiratory or cardiological complications. Mutations in RYR1, SEPN1, ACTA1, TTN and MEGF10 have been associated with various CMs, yet for about 50% of CM cases the responsible gene has not been identified. The objective of my thesis was to clarify the pathophysiological mechanisms of new forms of CM through the identification of new genes or new mutations in known genes. This thesis had an international dimension as manifested by a UPMC (France) and UdeM (Québec) joint direction. I developed two complementary axes of research. First, I studied 21 informative families with a recessive CM with scoliosis and respiratory failure, for which I combined positional cloning and candidate gene studies, using various tools from genotyping to next generation sequencing (NGS). The second part of this work consisted on the analysis of 24 families with recessive CM affecting both cardiac and skeletal muscles. Their phenotype was similar to that previously observed in cases with deletions in the last 6 exons of the giant gene TTN. Thus we applied a candidate gene strategy through direct Sanger sequencing coupled with NGS for the analysis of this second cohort. During my PhD work I identified the molecular defect in 8 out of the 45 families included (18%), which led to the identification and characterization of 3 novel medical entities, including two new CMs due to novel defects of TTN. These results served to identify new titin protein interactions, and participate in the definition of TTN defects as a major cause of both cardiac and skeletal muscle conditions. A third new form of CM is due to mutations of a poorly-known transcriptional coactivator whose role in striated muscle physiology was unknown and which had never been associated to a human condition. Globally, these results unveiled a novel important protein and pathway in muscle pathophysiology, have direct health benefits (molecular diagnosis) and open the way for therapeutic investigations.

Die Linksventrikuläre Noncompaction des Myokards (LVNC) ist eine seltene primäre Herzmuskelerkrankung. Es wird angenommen, dass es sich um eine embryonale Entwicklungsstörung des Myokards handelt. Mutationen in dem X-chromosomalen Gen TAZ sind verantwortlich für Fälle von frühkindlicher LVNC während die genetische Ursache autosomal-dominant vererbter adulter LVNC weitgehend unbekannt ist. In dieser Arbeit wurde die genetische Ursache der LVNC in der Familie LVNC-105 untersucht. Weiterhin wurden in einem großen Kollektiv von LVNC-Indexpatienten Kandidatengenanalysen durchgeführt. Bei der Familie LVNC-105 zeigte die genomweite Kopplungsanalyse nur signifikant hohe 2-Punkt-LOD-Werte auf Chromosom 11p15. Der maximale 2-Punkt-LOD-Wert betrug 5,06 bei D11S902 und der Lokus konnte auf 3,2 Mb (4,9 cM) eingeengt werden. Unter den 40 Genen des Erkrankungslokus war das Kandidatengen CSRP3, das bereits für 2 andere Kardiomyopathien, die dilatative und die hypertrophe Kardiomyopathie (DCM und HCM), als Krankheitsgen beschrieben wurde. Die Sequenzierung des genomischen Bereichs von CSRP3 zeigte keine Mutation bei den betroffenen Familienmitgliedern. Auch die Analyse von weiteren, im Lokus enthaltenen Gene ergab keine Mutation in kodierenden Exons. Auch Untersuchungen auf Transkriptebene offenbarten keine genetische Veränderung. Bei der Sequenzierung der LVNC-Kandidatengene LDB3, LMNA, Nkx2.5 und\linebreak BMP10 bei 63 erwachsenen Indexpatienten mit isolierter LVNC wurde nur eine Mutation in LDB3 gefunden. Erstmals wurden auch 7 Gene, die für sarkomere Proteine kodieren und als Krankheitsgene für HCM und DCM bekannt sind, mittels DHPLC untersucht. Es wurden Mutationen in einem großen Anteil der LVNC-Indexpatienten (19%) in MYH7, ACTC, TPM1 und TNNT2 identifiziert. Klinische Untersuchungen zeigten bei 7 von 12 Patienten mit Mutationen das Vorliegen einer familiären LVNC. In 4 autosomal-dominanten LVNC-Familien kosegregierten die MYH7 Mutationen mit der Erkrankung. MYH7 war mit einem Anteil von 13% das häufigste Krankheitsgen. Die Mutationen in MYH7 lagen vorwiegend in der ATP-Bindungsstelle. LVNC gehört damit zum Spektrum der Kardiomyopathien, die durch Mutationen in sarkomeren Proteinen hervorgerufen werden können. / Left ventricular noncompaction of the myocardium (LVNC) constitutes a rare primary cardiomyopathy. The mechanistic basis is assumed to be an arrest in embryonic cardiac development. Mutations in the X-linked TAZ gene are responsible for cases of infantile LVNC whereas the genetic base of late-onset LVNC in most patients is still unresolved. The objectives of this dissertation were to investigate the genetic defect in family LVNC-105 with autosomal dominant inherited LVNC and to screen a large cohort of patients with isolated LVNC for mutations in candidate genes. In kindred LVNC-105 genome wide linkage analysis revealed significant two-point LOD scores only at chromosome 11p15. A peak 2-point LOD score of 5.06 was obtained with marker D11S902 and a critical interval of 3.2 Mb (4.9 cM) was determined. Among the 40 genes within the disease region one candidate gene was CSRP3, a disease gene for hypertrophic (HCM) and dilated (DCM) cardiomyopathy. Sequence analysis of the genomic CSRP3 region did not reveal mutations in affected family members. Also, analysis of the coding region of further candidate genes contained within the disease locus did not show mutations. Investigations of the genes on transcript level did not detect alterations. Candidate gene analysis of LDB3, LMNA, Nkx2.5 and BMP10 in 63 index patients with isolated LVNC only one mutation was detected in LDB3. For the first time 7 genes encoding sarcomere proteins, known as disease genes for HCM and DCM, were screened for mutations by DHPLC in LVNC patients. Mutations were found in a significant proportion of the cohort of LVNC index patients (19%) in MYH7, ACTC, TPM1 and TNNT2. Clinical evaluations demonstrated familial disease in 7 of 12 probands with sarcomere gene mutations. MYH7 mutations segregated with the disease in 4 autosomal dominant LVNC kindreds. MYH7 was identified as the most prevalent LVNC disease gene (13%) in this cohort. Modified residues in MYH7 were mainly located within the ATP binding site. In conclusion, LVNC belongs to the spectrum of cardiomyopathies originating in molecular defects of the sarcomere.

<p>Systemic lupus erythematosus (SLE) is a complex autoimmune disease with unknown etiology. The aim of this thesis was to identify susceptibility regions through genetic mapping, using model-based linkage analysis on nuclear and extended SLE multicase families.</p><p>In the first paper we performed a genome scan on 19 genetically homogenous Icelandic and Swedish families. One region at 2q37 was identified with a significant linkage with contribution from both populations (Z=4.24). Five other regions 2q11, 4p13, 9p22, 9p13 and 9q13 showed suggestive linkage (Z>2.0).</p><p>In the second paper, 87 families from 10 different countries were analysed only for chromosome 1. One region at 1q31 showed significant linkage (Z=3.79) with contribution from families from all populations, including Mexicans and Europeans. Four other regions 1p36, 1p21, 1q23, and 1q25, showed levels of suggestive linkage. Linkage for most regions was highly dependent on what population was used, which indicated strong genetic heterogeneity in the genetic susceptibility for SLE.</p><p>In the two last papers, we used the positional candidate gene strategy, in order to investigate candidate genes in two regions linked to SLE. For the Bcl-2 gene (at 18q21) we could not detect any association with SLE using three different markers. However, when we investigated the tightly linked low-affinity family of FcγR genes (at 1q23), we could find association for two risk alleles in the FcγRIIA and FcγRIIIA genes. The risk alleles were transmitted to SLE patients on one specific haplotype and therefore are not independent risk alleles.</p><p>The results show that model-based linkage analysis is a strong approach in the search for susceptibility genes behind complex diseases like SLE.</p>