1 Genome Composition Dan Graur

2 Genome Composition in Bacteria

Carsonella ruddii has a very low GC content.

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5 The selectionist explanation views GC content as an adaptation. G:C pairs are more stable than A:T pairs. Preferential usage of amino acids encoded by GC-rich codons (e.g., ala and arg) and avoidance of amino acids encoded by GC- poor codons (e.g., ser and lys). T-T dimers are sensitive to UV radiation. Noempiricalevidence

6 The mutationist explanation Rate of substitution G/C  T/A is  Rate of substitution T/A  G/C is Noboru Sueoka University of Colorado

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9 Mycoplasma capricolum Escherichia coli Micrococcus luteus

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11 Differences in the way the leading and lagging strands of DNA are replicated can result in strand-dependent mutation patterns. The expectation under no-strand-bias conditions is f A = f T and f C = f G

12 Deviations from equal mutation rates between the two strands are quantified by the skew.

13 The skew is a measure of inequality between the frequencies of nucleotides X and Y on a strand.

14 If there are no violations of the no-strand-bias conditions:

15 Skew values are calculated for sliding windows of predetermined lengths, and are plotted on a skew diagram.

16 Bacillus subtilischirochorechirochore

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18 Chlamidia trachomatis

19 Compositional Properties of Eukaryotic Genomes

20 GC content of bacterial genomes ranges from ~24% to ~74% Intergenomic variability GC content of vertebrate genomes ranges from ~40% to ~45%

21 TTGACCGATGACCCCGGTTCAGGCTTCACCACAGTGTGGAACGCGGTCGTCTCCGAACTT AACGGCGACCCTAAGGTTGACGACGGACCCAGCAGTGATGCTAATCTCAGCGCTCCGCTG ACCCCTCAGCAAAGGGCTTGGCTCAATCTCGTCCAGCCATTGACCATCGTCGAGGGGTTT GCTCTGTTATCCGTGCCGAGCAGCTTTGTCCAAAACGAAATCGAGCGCCATCTGCGGGCC CCGATTACCGACGCTCTCAGCCGCCGACTCGGACATCAGATCCAACTCGGGGTCCGCATC GCTCCGCCGGCGACCGACGAAGCCGACGACACTACCGTGCCGCCTTCCGAGAGATTGATG ACAGCGCTGCGGCACGGGGCGATAACCAGCACAGTTGGCCAAGTTACTTCACCGAGCGCC CGCACAATACCGATTCCGCTACCGCTGGCGTAACCAGCCTTAACCGTCGCTACACCTTTG ATACGTTCGTTATCGGCGCCTCCAACCGGTTCGCGCACGCCGCCGCCTTGGCGATCGCAG AAGCACCCGCCCGCGCTTACAACCCCCTGTTCATCTGGGGCGAGTCCGGTCTCGGCAAGA CACACCTGCTACACGCGGCAGGCAACTATGCCCAACGGTTGTTCCCGGGAATGCGGGTCA AATATGTCTCCACCGAGGAATTCACCAACGACTTCATTAACTCGCTCCGCGATGACCGCA AGGTCGCATTCAAACGCAGCTACCGCGACGTAGACGTGCTGTTGGTCGACGACATCCAAT TCATTGAAGGCAAAGAGGGTATTCAAGAGGAGTTCTTCCACACCTTCAACACCTTGCACA ATGCCAACAAGCAAATCGTCATCTCATCTGACCGCCCACCCAAGCAGCTCGCCACCCTCG AGGACCGGCTGAGAACCCGCTTTGAGTGGGGGCTGATCACTGACGTACAACCACCCGAGC TGGAGACCCGCATCGCCATCTTGCGCAAGAAAGCACAGATGGAACGGCTCGCGGTCCCCG ACGATGTCCTCGAACTCATCGCCAGCAGTATCGAACGCAATATCCGTGAACTCGAGGCCG AGGAATTCACCAACGACTTCATTAACTCGCTCCGCGATGACCGCAAGGTCGCATTCAAAC GCAGCTACCGCGACGTAGACGTGCTGTTGGTCGACGACATCCAATTCATTGAAGGCAAAG Interspecific variation among vertebrate genomes is low. However, vertebrates seem to have a much more complex intragenomic compositional organization (internal structure) than prokaryotic genomes.

22 TTGACCGATGACCCCGGTTCAGGCTTCACCACAGTGTGGAACGCGGTCGTCTCCGAACTT AACGGCGACCCTAAGGTTGACGACGGACCCAGCAGTGATGCTAATCTCAGCGCTCCGCTG ACCCCTCAGCAAAGGGCTTGGCTCAATCTCGTCCAGCCATTGACCATCGTCGAGGGGTTT GCTCTGTTATCCGTGCCGAGCAGCTTTGTCCAAAACGAAATCGAGCGCCATCTGCGGGCC CCGATTACCGACGCTCTCAGCCGCCGACTCGGACATCAGATCCAACTCGGGGTCCGCATC GCTCCGCCGGCGACCGACGAAGCCGACGACACTACCGTGCCGCCTTCCGAGAGATTGATG ACAGCGCTGCGGCACGGGGCGATAACCAGCACAGTTGGCCAAGTTACTTCACCGAGCGCC CGCACAATACCGATTCCGCTACCGCTGGCGTAACCAGCCTTAACCGTCGCTACACCTTTG ATACGTTCGTTATCGGCGCCTCCAACCGGTTCGCGCACGCCGCCGCCTTGGCGATCGCAG AAGCACCCGCCCGCGCTTACAACCCCCTGTTCATCTGGGGCGAGTCCGGTCTCGGCAAGA CACACCTGCTACACGCGGCAGGCAACTATGCCCAACGGTTGTTCCCGGGAATGCGGGTCA AATATGTCTCCACCGAGGAATTCACCAACGACTTCATTAACTCGCTCCGCGATGACCGCA AGGTCGCATTCAAACGCAGCTACCGCGACGTAGACGTGCTGTTGGTCGACGACATCCAAT TCATTGAAGGCAAAGAGGGTATTCAAGAGGAGTTCTTCCACACCTTCAACACCTTGCACA ATGCCAACAAGCAAATCGTCATCTCATCTGACCGCCCACCCAAGCAGCTCGCCACCCTCG AGGACCGGCTGAGAACCCGCTTTGAGTGGGGGCTGATCACTGACGTACAACCACCCGAGC TGGAGACCCGCATCGCCATCTTGCGCAAGAAAGCACAGATGGAACGGCTCGCGGTCCCCG ACGATGTCCTCGAACTCATCGCCAGCAGTATCGAACGCAATATCCGTGAACTCGAGGCCG AGGAATTCACCAACGACTTCATTAACTCGCTCCGCGATGACCGCAAGGTCGCATTCAAAC GCAGCTACCGCGACGTAGACGTGCTGTTGGTCGACGACATCCAATTCATTGAAGGCAAAG How are nucleotides distributed along the genome? Uniform? Patchy? Clines?

23 “When vertebrate genomic DNA is randomly sheared into fragments kb in size and the fragments are separated by base composition, the fragments cluster into a small number of classes distinguished from each other by their GC content. Each class is characterized by bands of similar, but not identical, base compositions.” (Macaya et al. 1976; Thiery et al. 1976; Bernardi et al. 1985) Equilibrium centrifugation in Cs 2 SO 4 density gradient

24 carp

25 The Isochore Theory - Giorgio Bernardi carp

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27 Isochores do not merit the prefix “iso.” Lander et al. (2001)

28 Post genomic era (2001) Objections against the isochore theory: “We can rule out a strict notion of isochores as compositionally homogeneous.” Lander et al. (2001) “There are no isochores in chromosomes 21 and 22.” Häring and Kyper (2001) Defense of the isochore theory: “The conclusion of the authors that ‘isochores’ are not ‘strict isochores’ is correct, however isochore are fairly homogeneous regions.” Bernardi (2001)

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30 In search of isochores… Questions:  Do isochores exist?  Is the isochore theory a useful (or practical) concept?

31 Segmentation Models Assumption: Sequences can be partitioned into a number of segments each with a characteristic GC content. Each segment has a certain degree of internal homogeneity (or similarity).

32 In search of isochores… Methodology:  Define rigorously 6 attributes of isochores and of the isochore theory as applied to humans  Test attributes against the human genomedata

33 Attributes of isochores A1. Distinguishability: An isochore is a DNA segment that has a characteristic GC content that differs significantly from the GC content of adjacent isochores. A2. Homogeneity: An isochore is more homogeneous in its composition than the chromosome on which it resides. A3. Minimum length: The length of an isochore exceeds a certain cutoff value. In the literature, the most commonly mentioned value is 300 Kb.

34 Attributes of the isochore theory in humans A4. Genome coverage: The overwhelming majority of the human genome consists of segments abiding by A1- A3. Non-isochoric DNA takes up only a small fraction of the genome.

35 A5. Isochore families: The human genome comprises of five isochore families, each described by a particular Gaussian distribution of GC content. Attributes of the isochore theory in humans

36 A6. Isochore assignment into families: It is possible to classify each isochore into its isochore family based solely on its compositional properties. Practicality of the isochore theory

37 Segment length distribution The fitted regression line (solid line) indicates that the tail of the distribution exhibits power-law decay with an exponent of –2.38. P  L –2.38

38 Power laws everywhere!

39 Isochore families Most parsimonious Gaussian fit to putative isochores

40 Homogeneous “isochores” in vertebrates

41 Assignment into families Classification errors reach values of 70%. Only a minute fraction of segments can be classified with an expected error under 5%.

42 Summary (A1) Distinguishability  (A2) Homogeneity  (A3) Minimum length X (A1) Genome coverage  (A2) Isochore families  families (A3) Isochore assignment into families X

43 Conclusion: The isochore theory may have reached the limits of its usefulness as a description of genomic compositional structures.

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46 As of December genetic codes 11 mitochondrial 5 nuclear 1 nuclear + mitochondrial

47 Lock & Key Hypothesis

48 Frozen accidents Evolutionary Dead Ends

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50 The codon- capture hypothesis Thomas Jukes

51 AAA = lysine Universal genetic code

52 AAA = asparagine Echinodermata

53 Hemichordata AAA = unassigned

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