INVESTIGATING THE ROLE OF HUMAN GENOME- WIDE HETEROZYGOSITY AS A HEALTH RISK FACTOR Ozren Polasek PhD Thesis The University of Edinburgh 2009 i TABLE OF CONTENTS ABSTRACT...............................................................................................................iv FOREWORD.............................................................................................................. v DECLARATION......................................................................................................vii NOTES .....................................................................................................................viii GLOSSARY...............................................................................................................ix 1. INTRODUCTION.................................................................................................. 1 1.1. Prologue ............................................................................................................ 1 1.2. Background ...................................................................................................... 3 1.2.1. Inbreeding depression ................................................................................ 4 1.2.2. Inbreeding vigour ....................................................................................... 6 1.2.3. Outbreeding depression ............................................................................. 7 1.2.4. Heterosis .................................................................................................... 8 1.3. Measuring autozygosity, homozygosity and heterozygosity .......................... 10 1.3.1. Pedigree-based estimates ......................................................................... 10 1.3.2. Genetic marker based heterozygosity estimates ...................................... 12 1.4. Suggested molecular mechanisms of inbreeding depression and heterosis 21 1.5. Heterozygosity – fitness correlations ............................................................. 25 1.5.1. Effects of genome-wide heterozygosity on animal fitness and survival ... 28 1.5.2. Effects of genome-wide heterozygosity on human health ........................ 31 1.5.2.1. Serum lipid levels.............................................................................. 38 1.5.2.2. Bone mineral density and osteoporosis............................................. 39 1.5.2.3. Glucose and diabetes......................................................................... 40 1.5.2.4. Uric acid, gout and urinary tract stones ............................................ 41 1.6. A brief overview of the population genetics of the human population ........ 42 1.7. A local study population example: Croatian island isolates ......................... 51 2. AIMS AND OBJECTIVES ................................................................................. 55 2.1. Introduction .................................................................................................... 55 2.2. Aims ................................................................................................................ 55 2.3. Objectives ........................................................................................................ 55 2.3.1. Study population ...................................................................................... 55 2.3.2. Individual genome-wide heterozygosity estimation ................................. 56 2.3.3. Heterozygosity-fitness correlations .......................................................... 56 ii 3. MATERIALS AND METHODS ........................................................................ 57 3.1. Setting ............................................................................................................. 57 3.1.1. Historical accounts and demographics of the Vis Island ......................... 57 3.1.2. Historical accounts and demographics of the Korcula Island ................. 61 3.2. Measurements ................................................................................................ 63 3.3. Derived variables ............................................................................................ 66 3.4. Genealogy reconstruction and pedigree information ................................... 67 3.5. Genotyping ...................................................................................................... 68 3.6. Statistical analysis .......................................................................................... 70 4. RESULTS ............................................................................................................. 73 4.1. Characteristics of the study population ......................................................... 73 4.2. Descriptors of the heterozygosity estimates ................................................... 79 4.3. Comparison of different heterozygosity measures ........................................ 84 4.4. Heterozygosity-fitness correlations ................................................................ 93 5. DISCUSSION ..................................................................................................... 104 5.1. Measuring individual genome-wide heterozygosity .................................... 105 5.2. Heterozygosity-fitness correlations .............................................................. 117 5.3. Study limitations ........................................................................................... 124 6. CONCLUSION................................................................................................... 130 7. REFERENCES................................................................................................... 131 8. APPENDICES .................................................................................................... 150 8.1. Systematic review (Appendix I) ................................................................... 150 8.2. Standard operating procedures (Appendix II) ............................................ 155 8.3. Descriptive statistics of the investigated traits (Appendix III) .................... 191 8.4. An overview of lipidomics and glycomics (Appendix IV) ........................... 193 8.5. Publications (Appendix V) ........................................................................... 195 iii ABSTRACT Aim The aim of this study was to investigate the most commonly used approaches to measure individual genome-wide heterozygosity (IGWH) and to investigate whether IGWH can be considered as a health risk factor or a protective factor in humans. Methods This study was based on two samples from isolated communities of Croatian Adriatic islands, with a total of 1,930 adult examinees from Islands of Vis (N=986) and Korcula (N=944). Examinees were genotyped with a total of 302,662 single nucleotide polymorphisms. Heterozygosity was estimated using five commonly calculated methods. Results Correlation coefficients between different heterozygosity methods were generally in the range of 0.7-0.8. A worsening in some phenotypic traits, including cholesterol and triglycerides as well as increased odds for osteoporosis and metabolic syndrome was recorded in cases of IGWH reduction. Nevertheless, in these cases heterozygosity explained a relatively low amount of variance, generally in range of 0.4-0.6% of total trait variance. Conclusion However, these results were significant in Vis Island sample, while in the replication sample, Korcula Island, most of the associations were not significant, possibly due to the overall lower amount of inbreeding and higher heterozygosity in Korcula Island sample. The results warrant further research in order to provide more information on the extent and importance of individual genome-wide heterozygosity, which might have an important role in communities which experience consanguinity on a greater scale. Two main shortcomings of the study include possible lack of power to detect inbreeding depression and the need to replicate the results in other populations. iv FOREWORD This Thesis is the result of my involvement in a large genetic epidemiology programme that was initiated in 1999. I joined the research group in 2003 and have been substantially involved in this work ever since. Initially, I joined as a medical doctor and was immediately involved in field work in the Croatian islands. Firstly I worked for brief periods in the islands of Lastovo and Mljet, then in 2003 and 2004 I worked in the island of Vis, sampling and organising the field work that constitutes a major part of this Thesis. This stage of the field work was largely organised and performed by the staff of the Institute for Anthropological Research (notably Professor Pavao Rudan, Branka Janicijevic and Nina-Smolej Narancic and many more who worked in the field). After the field work was completed, I created a database, entered the data, and organised and carried out most of the genealogy reconstruction work with the help of Dr. Ivana Kolcic (Zagreb University School of Medicine). In 2005 I was awarded a PhD Scholarship by the University of Edinburgh. The same year I was also awarded two additional scholarships which enabled my further involvement in this field – a scholarship from the Croatian Ministry of Science, Education and Sports and another from the Association of the Schools of Public Health in the European Region (ASPHER). During my second year I was awarded a scholarship by the Overseas Research Scheme,