1. Introduction to genomes Content  the human genome CNVs SNPs Alternative splicing  genome projects Celia van Gelder CMBI UMC Radboud June 2009 c.vangelder@cmbi.ru.nl

2. The human genome Genome: the entire sequence of DNA in a cell 3 billion basepairs (3Gb) 22 chromosome pairs + X en Y chromosomes Chromosome length varies from ~50Mb to ~250Mb About 22000 protein-coding genes Human genome is 99.9% identical among individuals

3. Eukaryotic Genomes: more than collections of genes Protein coding genes RNA genes (rRNA, snRNA, snoRNA, miRNA, tRNA) Structural DNA (centromeres, telomeres) Regulation-related sequences (promoters, enhancers, silencers, insulators) Parasite sequences (transposons) Pseudogenes (non-functional gene-like sequences) Simple sequence repeats

4. Annotating the genome Genome annotation is the process of attaching biological information to sequences. It consists of two main steps: 1.identifying elements on the genome, a process called Gene Finding, and 2.attaching biological information to these elements. Automatic annotation tools try to perform all this by computer analysis, as opposed to manual annotation which involves human expertise. Ideally, these approaches co-exist and complement each other in the same annotation pipeline.

5. The human genome cntnd From: Molecular Biology of the Cell (4 th edition) (Alberts et al., 2002) Only 1.2% codes for proteins, 3.5-5% is under selection Long introns, short exons Large spaces between genes More than half consists of repetitive DNA

6. Eukaryotic Genomes: High fraction non-coding DNA Blue: Prokaryotes Black: Unicellular eukaryotes Other colors: Multicellular eukaryotes (red = vertebrates) From: Mattick, NRG, 2004

7. Variation along genome sequence Nucleotide usage varies along chromosomes – Protein coding regions tend to have high GC levels Genes are not equally distributed across the chromosomes – Housekeeping generally in gene- dense areas – Gene-poor areas tend to have many tissue specific genes From: Ensembl

8. Chromosome organisation (1) From: Lodish (4 th edition)

9. Chromosome organisation (2) From: Lodish (4 th edition) DNA packed in chromatin Non-active genes often in densely packed chromatin (30-nm fiber) Active genes in less dense chromatin (beads-on-a-string) Gene regulation by changing chromatin density, methylation/acetylation of the histones Genes that are OFF Genes that are ON

10. Today’s focus 1.Copy number variations (CNV) 2.Single Nucleotide Polymorphisms (SNPs) 3.Alternative transcripts

11. Copy Number Variation People do not only vary at the nucleotide level (SNPs) Copy Number Variations (CNVs): duplications and deletions of pieces of chromosome When there are genes in the CNV areas, this can lead to variations in the number of gene copies between individuals CNVs may either be inherited or caused by de novo mutation

12. Why study CNVs? CNVs are common in cancer and other diseases. CNVs are also common in normal individuals and contribute to our uniqueness. These changes can also influence the susceptibility to disease. Since CNVs often encompass genes, they can have important roles both in characterizing human disease and discovering drug response targets. Understanding the mechanisms of CNV formation may also help us better understand human genome evolution.

13. CNV & disease, examples CNVs have been implicated in Cancer EGFR higher copy number in non-small cell lung cancer Low copy number of FCGR3B can increase susceptibility to SLE & other autoimmune disorders Autism Schizophrenia (dept. human genetics) Mental retardation (dept. human genetics)

14. Single Nucleotide Polymorphisms (SNPs) SNPs are DNA sequence variations that occur when a single nucleotide (A,T,C,or G) in the genome sequence is altered. Similar to mutations, but are simultaneously present in the population, and generally have little effect Are being used as genetic markers (a genetic disease is e.g. associated with a SNP) T T T T T T A A A A A C C CG G G G A T T T T T T A A A A A C C CG G G G A CG GCTA Single Nucleotide- Polymorphism (SNP)

15. SNP fact sheet For a variation to be considered a SNP, it must occur in at least 1% of the population. SNPs, which make up about 90% of all human genetic variation, occur every 100 to 300 bases along the 3-billion-base human genome. Two of every three SNPs involve the replacement of cytosine (C) with thymine (T). SNPs can occur in coding (gene) and non coding regions of the genome.

16. SNPs & medicine Although more than 99% of human DNA sequences are the same, variations in DNA sequence can have a major impact on how humans respond to: – disease; – environmental factors such as bacteria, viruses, toxins, and chemicals; – and drugs and other therapies. This makes SNPs valuable for biomedical research and for developing pharmaceutical products or medical diagnostics. SNPs are also evolutionarily stable—not changing much from generation to generation—making them easier to follow in population studies.

17. SNP & disease, example Alzheimer's disease & apolipoprotein E ApoE contains two SNPs that result in three possible alleles for this gene: E2, E3, and E4. Each allele differs by one DNA base, and the protein product of each gene differs by one amino acid. Each individual inherits one maternal copy of ApoE and one paternal copy of ApoE. Research has shown that a person who inherits at least one E4 allele will have a greater chance of developing Alzheimer's disease.

18. The HapMap Project is a multi-country effort to identify and catalog genetic similarities and differences in human beings. Using HapMap, researchers will be able to find genes that affect health, disease, and individual responses to medications and environmental factors. HapMap is a collaboration among scientists and funding agencies from Japan, the United Kingdom, Canada, China, Nigeria, and the United States All of the information generated will be released into the public domain. www.hapmap.org HapMap

19. Alternative splicing

20. Alternative splicing (2) ~ 15 % of the mutations that cause genetic diseases affect pre-mRNA splicing

21. Genome projects, a bit of history http://www.genomesonline.org/

23. Sequenced genomes 1995Haemophilus influenzae 1.8 Mb 1996Yeast 12 Mb 1998C. elegans100 Mb 1999Fruit fly125 Mb 2000Arabidopsis115 Mb 2001Human (draft) 2002Mouse 2.6 Gb 2002 Rice 2004Human (“finished”) 3 Gb 2006 Sea urchin 2007 Grapevine 2008 Platypus (draft) 2009 Cow

24. Some genome sizes OrganismGenome size (base pairs) Virus, Phage Φ-X174;5387 First sequenced genome Virus, Phage λ5×10 4 Bacterium, Escherichia coli4×10 6 Plant, Fritillary assyrica13×10 10 Largest known genome Fungus,Saccharomyces cerevisiae2×10 7 Nematode, Caenorhabditis elegans8×10 7 Insect, Drosophila melanogaster2×10 8 Mammal, Homo sapiens3×10 9

25. Genome browsers can be used to examine …. –Genomic sequence conservation –Duplications en deletions of pieces chromosome (Copy Number Variations, CNVs) –Single Nucleotide Polymorphisms (SNPs) –Alternative splicing –And much more…. LET’S GO BROWSE GENOMES!

26. Alternative Transcripts Source: Wikipedia (http://www.wikipedia.org/)