Till mamma, pappa, bror och Lasse List of Papers This thesis is based on the following papers, which are referred to in the text by their Roman numerals. I Díaz de Ståhl TD, Sandgren J, Piotrowski A, Nord H, Anders- son R, Menzel U, Bogdan A, Thuresson AC, Poplawski A, von Tell D, Hansson CM, Elshafie AI, Elghazali G, Imreh S, Nor- denskjöld M, Upadhyaya M, Komorowski J, Bruder CE, Du- manski JP. Profiling of copy number variations (CNVs) in healthy individuals from three ethnic groups using a human ge- nome 32 K BAC-clone-based array. Hum Mutat. 2008 Mar;29(3):398-408. II Piotrowski A, Bruder CE, Andersson R, Díaz de Ståhl TD, Menzel U, Sandgren J, Poplawski A, von Tell D, Crasto C, Bogdan A Bartoszewski R, Bebok Z, Krzyzanowski M, Jan- kowski Z, Partridge EC, Komorowski J, Dumanski JP. Somatic mosaicism for copy number variation in differentiated human tissues. Hum Mutat. 2008 Sep;29(9):1118-24. III Sandgren J, Díaz de Ståhl T, Andersson R, Menzel U, Pio- trowski U, Nord H, Bäckdahl M, Kiss N, Braukhoff M, Komo- rowski J, Dralle H, Hessman O, Larsson C, Åkerström G, Brud- er C, Dumanski J.P and Westin G. Recurrent genomic altera- tions in benign and malignant pheochromocytomas and para- gangliomas revealed by whole-genome array comparative genomic hybridization analysis. Endocr Relat Cancer. 2010 June;17(3):561-79. IV Sandgren J*, Andersson R*, Rada-Iglesias A, Enroth S, Du- manski J.P, Åkerström G, Komorowski J, Westin G and Wade- lius C. Integrative Epigenomic and Genomic Analysis of Ma- lignant Pheochromocytoma. *Both authors contributed equally. Exp Mol Med. 2010 July;42(7):484-502. Reprints were made with permission from the respective publishers. Related publications by the author i Meyer-Rochow GY, Jackson NE, Conaglen JV, Whittle DE, Kunnimalaiyaan M, Chen H, Westin G, Sandgren J, Stalberg P, Khanafshar E, Shibru D, Duh QY, Clark OH, Kebebew E, Gill A, Clifton-Bligh R, Robinson BG, Benn DE, Sidhu SB. MicroRNA profiling of benign and malignant pheochromocy- toma identifies novel diagnostic and therapeutic targets. En- docr Relat Cancer. 2010 Aug 16;17(3):835-846. ii Popławski AB, Jankowski M, Erickson SW, Díaz de Ståhl T, Partridge EC, Crasto C, Guo J, Gibson J, Menzel U, Bruder CE, Kaczmarczyk A, Benetkiewicz M, Andersson R, Sandgren J, Zegarska B, Bała D, Srutek E, Allison DB, Pio- trowski A, Zegarski W, Dumanski JP. Frequent genetic dif- ferences between matched primary and metastatic breast can- cer provide an approach to identification of biomarkers for disease progression. Eur J Hum Genet. 2010 May;18(5):560- 8. iii Nord H, Segersten U, Sandgren J, Wester K, Busch C, Men- zel U, Komorowski J, Dumanski JP, Malmstrom PU, Díaz de Ståhl T. Focal amplifications are associated with high-grade and recurrences in stage Ta bladder carcinoma. Int J Cancer. 2010 Mar 15;126(6):1390-402. iv Nord H, Hartmann C, Andersson R, Menzel U, Pfeifer S, Piotrowski A, Bogdan A, Kloc W, Sandgren J, Olofsson T, Hesselager G, Blomquist E, Komorowski J, von Deimling A, Bruder CE, Dumanski JP, de Ståhl TD. Characterization of novel and complex genomic aberrations in glioblastoma using a 32K BAC array. Neuro Oncol. 2009 Dec;11(6):803-18. v Bruder CE, Piotrowski A, Gijsbers AA, Andersson R, Erick- son S, Díaz de Ståhl T, Menzel U, Sandgren J, von Tell D, Poplawski A Crowley M, Crasto C, Partridge EC, Tiwari H, Allison DB, Komorowski J, van Ommen GJ, Boomsma DI, Pedersen NL, den Dunnen JT, Wirdefeldt K, Dumanski JP. Phenotypically concordant and discordant monozygotic twins display different DNA copy-number-variation profiles. Am J Hum Genet. 2008 Mar;82(3):763-71. vi Andersson R, Bruder CE, Piotrowski A, Menzel U, Nord H, Sandgren J, Hvidsten TR, Díaz de Ståhl T, Dumanski JP, Komorowski J. A segmental maximum a posteriori approach to genome-wide copy number profiling. Bioinformatics. 2008 Mar 15;24(6):751-8. Contents INTRODUCTION ........................................................................................ 13 Human Genetic Variation ......................................................................... 15 Copy number variations ....................................................................... 16 Somatic mosaicism ................................................................................... 22 DNA copy number changes in the context of somatic mosaicism ...... 22 Epigenetics ............................................................................................... 25 DNA Methylation ................................................................................ 25 Chromatin and genomic regulation ..................................................... 26 Histone modifications .......................................................................... 27 Cancer ...................................................................................................... 30 Hallmarks of cancer ............................................................................. 30 Cancer genetics .................................................................................... 32 Genomic profiling of cancer ................................................................ 33 Epigenetics in cancer ........................................................................... 34 Remarks on large scale cancer genetic/epigenetic research ................ 38 Pheochromocytoma .................................................................................. 40 Diagnosis and management of pheochromocytoma ............................ 42 Malignant pheochromocytomas ........................................................... 42 Familial pheochromocytomas .............................................................. 45 Genetic testing in pheochromocytoma patients ................................... 47 Signaling pathways and genes involved in pheochromocytoma development ......................................................................................... 48 Somatic genetic and epigenetic alterations in pheochromocytomas .... 50 PRESENT INVESTIGATIONS ................................................................... 53 Aims ......................................................................................................... 53 METHODS ................................................................................................... 54 Array- CGH .............................................................................................. 54 ChIP-chip ................................................................................................. 56 SUMMARY OF INCLUDED PAPERS ....................................................... 57 Paper I ...................................................................................................... 57 Profiling of Copy Number Variations (CNVs) in Healthy Individuals from Three Ethnic Groups Using a Human Genome 32 K BAC-Clone- Based Array ......................................................................................... 57 Paper II ..................................................................................................... 58 Somatic Mosaicism for Copy Number Variation in Differentiated Human Tissue ...................................................................................... 58 Paper III .................................................................................................... 59 Recurrent Genomic Alterations in Benign and Malignant Pheochromocytomas and Paragangliomas Revealed by Whole-Genome Array-CGH Analysis ........................................................................... 59 Paper IV ................................................................................................... 61 Integrative Epigenomic and Genomic Analysis of Malignant Pheochromocytoma ............................................................................. 61 CONCLUDING REMARKS ........................................................................ 63 Ongoing research and future perspectives ................................................ 63 General and future genomic and epigenomic views on pheochromocytoma and cancer ................................................................ 66 ACKNOWLEDGMENTS ............................................................................ 69 REFERENCES ............................................................................................. 73 Abbreviations Array-CGH Array-based comparative genomic hybridiza- tion BAC Bacterial artificial chromosome ChIP Chromatin immunoprecipitation CNV Copy number variation DNA Deoxyribonucleic acid FoSteS Fork stalling and template switching GWAS Genome wide association studies H3K4me3 Histone 3 lysine 4 trimethylation H3K27me3 Histone 3 lysine 27 trimethylation kb kilo base pair LCR Low copy repeats LOH Loss of heterozygosity Mb Mega base pair MEN2 Multiple endocrine neoplasia type 2 miRNA microRNA MOR Minimal overlapping region MZ Monozygotic NAHR Nonallelic homologous recombination ncRNA non coding RNA NHEJ Nonhomologous end joining NF1 Neurofibromatosis type 1 PcG Polycomb group protein PCR Polymerase chain reaction PRC Polycomb repressive complex PGL Paraganglioma syndrome RNA Ribonucleic acid SD Segmental duplication SNP Single nucleotide polymorphism SNV Single nucleotide variant TSS Transcription start site VHL von Hippel-Lindau syndrome INTRODUCTION The genome can be described as the entirety of an organism’s hereditary information that lies in the DNA sequence. The human nuclear genome con- sists of more than 3 billion bases constituting the DNA molecules present