Non-conserved intronic motifs in human and mouse are associated with a conserved set of functions

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1 Non-conserved intronic motifs in human and mouse are associated with a conserved set of functions Aristotelis Tsirigos Bioinformatics & Pattern Discovery Group IBM Research

2 Outline. Discovery of DNA motifs in human (PNAS 26) 2. Explore potential regulatory role in introns 3. Link to function 4. Relevance to cancer 2

3 Motif discovery

4 Preparation of the input pattern discovery Color legend INTERGENIC patterns INTRONIC 5 UTR pyknons CODING 3 UTR 4

5 Summary of pyknon properties Discovery constraints: 6 nucleotides long 3 intact copies in intergenic/intronic regions additional copies in exonic regions Discovered pyknon set properties: comprise the most frequent motifs in the genome ~2, distinct, non-overlapping overlapping pyknons in human cover ~2% of human genome appear in ~9% of human genes 5

6 Summary of pyknon properties Discovery constraints: 6 nucleotides long 3 intact copies in intergenic/intronic regions additional copies in exonic regions Discovered pyknon set properties: comprise the most frequent motifs in the genome ~2, distinct, non-overlapping overlapping pyknons in human cover ~2% of human genome appear in ~9% of human genes 6

7 Pyknons are not conserved

8 Genomic regions region human mouse entire genome 5.7 Gbp 5.3 Gbp conserved regions repeat elements 33% 24% 36% 2% introns 8% 7% pyknons 2% 9% Recent findings (ENCODE project, Nature 27): conserved non-exonic regions are not necessarily functional large fraction of non-conserved regions are functional 8

9 Questions Are pyknons conserved across human and mouse? Are pyknons related to repeat elements? Can we link pyknons to function? 9

10 Human intron decomposition

11 Mouse intron decomposition

12 Human genome decomposition more conservation 2

13 Mouse genome decomposition more conservation 3

14 Pyknons vs. conserved regions intra-genomic signature human inter-genomic conservation inter-genomic conservation mouse intra-genomic signature DNA sequence conserved sequences human pyknons mouse pyknons 4

15 Key observations Pyknons are not conserved across human and mouse Pyknons do not represent a rediscovery of repeats Pyknons lie inside unexplored genomic territory But, can we link pyknons to function? 5

16 Linking pyknons to function

17 Input: Computational method intronic sequences per gene region type (conserved, repeats, or pyknons) to be tested Functional annotations: a set of GO terms per gene Statistical test: over-concentration in genes associated with some GO term Output: a set of enriched GO terms for each region type 7

18 Step-by by-step: introns sequences gene # gene #2 gene #3 gene #n intronic sequence 8

19 Step-by by-step: pyknons sequences gene # gene #2 gene #3 gene #n intronic sequence pyknon instances 9

20 Step-by by-step: concentrations sequences concentrations gene # gene #2 gene #3 x x 2 x 3 gene #n x n intronic sequence pyknon instances 2

21 Step-by by-step: GO terms sequences concentrations GO terms gene # gene #2 gene #3 x x 2 x 3 A, C, F B, C, D, E C, E gene #n x n A, F, H intronic sequence pyknon instances 2

22 Step-by by-step: GO terms sequences concentrations GO terms gene # gene #2 gene #3 x x 2 x 3 A, C, F B, C, D, E C, E gene #n x n A, F, H intronic sequence pyknon instances 22

23 Step-by by-step: GO term sets concentrations x x 2 x 3 GO terms A, C, F B, C, D, E C, E x n A, F, H 23

24 Step-by by-step: GO term table concentrations GO terms A B C D E F G H x A, C, F x 2 B, C, D, E x 3 C, E x n A, F, H 24

25 Step-by by-step: test each GO term concentrations GO terms A B C D E F G H x A, C, F x 2 B, C, D, E x 3 C, E x n A, F, H 25

26 Statistical test concentrations for each GO term T, compute t-test t test statistic x x 2 x 3 probability x n pyknon concentrations in rest of genes pyknon concentrations in genes whose annotation contains GO term T Output = a p-value p for each GO term 26

27 Step-by by-step: next GO term concentrations GO terms A B C D E F G H x A, C, F x 2 B, C, D, E x 3 C, E x n A, F, H 27

28 Algorithm output GO term A B C p-value p A p B p C H p H 28

29 Random permutation test permuted concentrations GO terms A B C D E F G H x 7 A, C, F x B, C, D, E x C, E x 2 A, F, H Permute concentrations and repeat previous steps The result is a random background distribution of p-valuesp 29

30 False discovery rate GO term p-value permutation () permutation (k) A p A p () A p (k) A B p B p () B p (k) B C p C p () C p (k) C H p H p () H p (k) H FDR(p) = { p (j) f p f=a,b,...,h and j k k } /k { p f p f=a,b,...,h } 3

31 Analyzed regions We analyzed the following intronic regions for functional links in human and mouse (% FDR): pyknons conserved regions repeat elements (no significant GO terms found) 3

32 Enriched GO terms per region conserved human conserved mouse human pyknons mouse pyknons conserved human 385 conserved mouse 46 human pyknons 87 mouse pyknons 24 32

33 Pair-wise GO term overlaps conserved human conserved mouse human pyknons mouse pyknons conserved human 385 conserved mouse 334 (87%) 46 human pyknons 87 mouse pyknons 24 33

34 Pair-wise GO term overlaps conserved human conserved mouse human pyknons mouse pyknons conserved human 385 conserved mouse 334 (87%) 46 human pyknons 87 mouse pyknons 94 (76%) 24 34

35 Pair-wise GO term overlaps conserved human conserved mouse human pyknons mouse pyknons conserved human 385 conserved mouse 334 (87%) 46 human pyknons (%) 4 (2%) 87 mouse pyknons (%) (%) 94 (76%) 24 35

36 Enriched GO terms conserved intronic regions cellular process cell communication regulation of cellular process cell adhesion cell differentiation regulation of biological process negative regulation of biological process regulation of development regulation of physiological process positive regulation of biological process regulation of growth interaction between organisms growth development sex differentiation developmental maturation anatomical structure development embryonic development pattern specification segmentation response to stimulus defense response response to biotic stimulus response to chemical stimulus response to stress response to external stimulus behavior pyknons in introns cellular physiological process chromosome segregation cellular metabolism transport cell division cell cycle metabolism catabolism biosynthesis macromolecule metabolism primary metabolism protein localization response to endogenous stimulus regulation of hydrolase activity organelle localization 36

Enriched GO terms pyknons in introns cellular physiological process chromosome segregation cellular metabolism (chromatin modification, DNA repair, ubiquitin cycle, ) transport cell division

37 Enriched GO terms pyknons in introns cellular physiological process chromosome segregation cellular metabolism (chromatin modification, DNA repair, ubiquitin cycle, ) transport cell division cell cycle (meiosis, mitosis, ) The set of direct targets of BCL6 is enriched in modulators of transcription, chromatin structure, protein ubiquitylation, cell cycle, and DNA damage responses. 37

38 Pyknons and cancer-associated associated regions pyknon densities (%) loss of heterozygocity sites amplified sites fragile sites introns conserved regions genome % 2% 4% 6% 8% % 2% 4% 6% 8% 2% 38

39 Summary Feature Conserved regions Pyknons Length long short Cross-species conservation YES NO Organism-specific conservation NO YES Functional conservation YES YES 39

40 Connection to pirnas

41 Introduction to pirnas Basic facts about pirnas: novel class of small RNAs,, form RNA-protein complexes longer than mirnas (~29nt long) ~3, distinct molecules present during meiosis in human and mouse stem-cell maintenance in Drosophila Main result: pyknons computationally predicted most pirnas 4

42 pirna overlaps 42

43 Conclusions

44 Conclusions Pyknons are not conserved and distinct from repeats Pyknons are linked to a conserved set of functions A subset of pyknons corresponds to pirnas 44

45 Publication 45

46 Future work Is there a connection? pyknons = organism-specific specific signature motifs pyknon function = DNA maintenance (signature preservation) pirna function = stem-cell maintenance disruption of signature-preserving mechanism = cancer? 46

47 Next steps Experimentally link pyknons to function/disease collaboration with George Calin (MD Anderson Cancer Center) design human pyknon array analysis of differential expression (normal vs. cancer) 47

48 Acknowledgments Bioinformatics group at IBM Research: Isidore Rigoutsos Tien Huynh Dan Platt Alice McHardy Kevin Miranda 48

Evidence for convergent evolution of ALU repeats in human and mouse

Evidence for convergent evolution of ALU repeats in human and mouse Aristotelis Tsirigos - IBM Computational Genomics Group May 2 I. ALU repeats: a class of mobile elements II. Evolution of ALUs III. Genomic