PABIO 551 -- Welcome

1. PABIO 551 -- Welcome

2. PABIO 551

3. PABIO 551 grades

4. PABIO 551 readings

5. Problem sets

6. Midterm exam

7. Final paper

8. Final paper guidelines

9. Course goals

10. The Central Dogma

11. Why does biology have so many exceptions?

12. Why does biology have so many exceptions?

13. Lineage tree of life on Earth

14. Copyright (c) by W. H. Freeman and Company DNA Structure DNA composition DNA forms Chromosomes

15. All nucleotides have a common structure

16. The five principal bases in nucleic acids

17. The phosphodiester backbone of DNA

18. The double helix

19. DNA can adopt different conformations

20. DNA forms

21. Non-B DNA structures

22. Non-B DNA conformations Non-B DNA complexes are mutagenic Due to the non-B conformation not DNA sequence Have contorted bond angles or unpaired nucleotides Associated with human genomic disorders: ~20 neurological diseases (simple sequence amplifications) ~50 hereditary diseases (e.g. genomic rearrangements, and deletions) Some psychiatric disease (polymorphisms of simple repeat sequences)

23. Disease associations with Z-DNA Rheumatoid arthritis Multiple sclerosis Type 1 Diabetes Infectious disease susceptibility Chronic hepatitis C Leukemia Lymphoma

24. Z- DNA and disease Alzheimer�s disease Severely infected � Z DNA conformation Moderate - probable B-Z intermediate form Normal - B-DNA conformation Amyloid Beta and aluminum (an etiologic factor) can modulate helical alterations in vivo Alternate purine pyrimidine sequences found in promoter regions of AD specific genes (amyloid precursor proteins, presenilin and ApoE

25. A role of Z-DNA binding in poxvirus pathogenesis E3L gene product is essential for virulence N-terminal domain has sequence similarity to Za family (binds Z-DNA) Tested virulence of E3L with decreased Z-DNA binding and chimeric viruses with different Z-DNA binding abilities Kim et al. 2003. PNAS. 100:6974

28. Role of E3L Z-DNA binding Modulates expression of host cellular genes Transactivation of IL-6, nuclear factor of activated T cells (NF-AT), and p53 transcription Inhibits hygromycin induced apoptosis

29. DNA can undergo reversible strand separation

30. UV absorption and DNA denaturation

31. GC content and DNA denaturation

32. Genome complexity and Cot�

33. Genome complexity and Cot�

34. Simple-sequence DNAs are concentrated in specific chromosomal locations

35. Simple sequence DNA and neurodegenerative disease

36. DNA supercoiling

37. Types of cellular DNA

38. DNA forms separate on agarose gels

39. Organizing cellular DNA into chromosomes Most bacterial chromosomes are circular with one replication origin. But Borrelia, Rhodococcus have multiple, linear chromosomes. Eukaryotic chromosomes each contain one linear DNA molecule and multiple origins of replication. But the S. pombe genome may be circular. Bacterial DNA is usually associated with polyamines. Eukaryotic DNA usually associates with histones to form chromatin.

40. Chromatin exists in extended and condensed forms

41. The solenoid model of condensed chromatin

42. Core histones are extensively modified

43. The histone code hypothesis

44. Nonhistone proteins provide a structural scaffold for long chromatin loops

45. Chromatin packing in metaphase chromosomes

46. Three functional elements for stable inheritance of eukaryotic chromosomes Origin for initiation of DNA replication. The centromere (point of spindle fiber attachment). The two ends (telomeres).

47. Experimental evidence for the importance of the origin of replication

48. Experimental evidence for the importance of the centromere

49. Experimental evidence for the importance of telomeres

50. Lecture 1 (part 2) Genes and genomes Pathogenicity islands Organellar genomes

51. Gene:

52. DNA content and phylogeny

53. The human genome

54. Orthologs and paralogs

55. Orthologs and paralogs II

56. Gene families

57. Gene classes

59. Concept of essential genes vs. minimal genome Both concepts are situationally defined Essential genes - knock out one gene at a time and ask if organism survives Minimal genome � number of genes considered essential for organisms to survive

60. What really are essential genes? Clouded by functional redundancy, which has been proposed to under predict �essential genes� E. coli � 134 essential genes � now 245 M. genitalium � only 480 genes 256 essential genes � now 382

61. How much genetic diversity within a species?

62. Definitions Pan-genome � global gene repertoire of a bacterial species Core genome + Dispensable Genome Core genome � genes shared by all strains of the same species Dispensable genome � consisting of partially shared and strain-specific genes, i.e. genes present in some but not all of the same species

63. The �Open� Pan Genome Group B strep Core genome -1806 genes Dispensable genome � 906 genes with each genome sequenced � 33 new genes S. pyogenes - 5 strains sequenced With each genome sequenced � 27 new genes E. coli Strains � 7 sequenced Core genome � 2,865 genes Each new genome sequenced � 441 new genes

64. Update on the E. coli pan genome Comparative genomic analysis of commensal and pathogenic isolates (n=17) E. coli pan genome � 13,000 genes Core genome � ~2200 genes Few conserved �pathovar specific genes� e.g. among EPEC genomes or EAEC genomes Predict 300 new genes per genome sequenced Significant genetic mosaicism between pathogen and commensal Features thought to be pathogen associated � found in commensal Commensals serve as reservoir potential virulence factors Commensals interact with �precursor pathogens� to allow development of �pathogens� Rasko et al. J. Bacteriol. 2008. Published online ahead of print, Aug. 1, 2008

65. The �Closed� Pan-Genome Bacillus anthracis Number of specific genes added to Pan-genome converged to 0 after the addition of a fourth genome Four genome sequences are sufficient to characterize species Mycobacterium tuberculosis Chlamydia trachomatis

66. The Bacterial Pan-Genome Analyzed 573 sequenced genomes Chose 15,000 random ORF Distinguished 3 groups of ORF Conclusion: the bacterial pan genome is of infinite size

67. The Bacterial Pan Genome

68. Genomes, Pathogenicity Islands, and Virulence

69. Pathogenicity Islands

70. A model pathogenicity island

71. Some virulence factors on pathogenicity islands

72. Adhesins mediate bacterial attachment

73. PAI Toxins

74. Iron uptake systems

75. Invasins, modulins, effectors

76. Secretion systems

77. Type III Secretion Systems Injectisome permits bacteria �docked� at the cell membrane to deliver effector proteins across the membrane Various families of injectisomes (e.g. SP2-2, SPI-2, YSC) Found in a wide variety of pathogens Chlamydia trachomatis, Yersinia pestis, Pseudomonas aeruginosa, Bordatella pertussis, Salmonella enterica, Vibrio parahemolyticus, Yersinia enterocolitica,

78. Type III Secretion Over 100 different effector proteins Roles of effector proteins Invasion of cells Inhibition of phagocytosis Down regulation of pro inflammatory responses Induction of apoptosis Prevention of autophagy Modulation of intracellular trafficking

79. Targets of effector proteins Small GTP binding proteins Mitogen-activated protein kinases I?B phosphoinositides

80. Type IV � ex. H. pylori, B. pertussis, N. gonorrhoeae, Coxiella burnetii. Delivery of bacterial effector proteins Across the bacterial membrane and the eucaryotic plasma membrane � contributes to pathogenesis Mediate horizontal gene transfer Contribute to genome plasticity Evolution of infectious pathogens Dissemination of antibiotic resistance and other virulent factors Discovered a T4SS responsible for formation of conjugative pilus and conjugative transfer of a genomic island (Juhas et al. 2006. J. Bacteriol)

82. Organelle Genomes

83. Mitochondrial DNAs Mitochondria contain multiple mtDNA molecules. Genes in mtDNA encode rRNAs, tRNAs, and proteins of the mt respiratory chain. apcytochrome B (CYb) and cytochrome c oxidase I (COI) The size and coding capacity of mtDNA varies considerably in different organisms. The products of mitochondrial genes are not exported. Mutations in mtDNA cause several genetic diseases in humans. Mitochondrial inheritance is distinct from nuclear.

84. Human mtDNA

85. Mitochondrial diseases in humans

86. Examples of diseases Leber�s hereditary optic neuropathy missense mutation in gene encoding subunit 4 of NADH-CoQ reductase Degeneration of optic nerve Myclonic epilepsy and red fiber disease single mutation in the TCG loop of mitochondrial lysine tRNA (thus, pool of defective tRNA) Decreased synthesis of proteins required for electron transport and ATP production inhibition of several mitochondria 1 proteins results in �ragged muscle fibers� Muscle weakness, heart problems, epilepsy, or dementia

87. Mitochondrial diseases in humans

88. Parasite mtDNAs can be very varied

89. Trypanosome mtDNAs - the kinetoplast

90. kDNA

91. Many trypanosme mtRNAs are edited

92. MtDNA of the Apicomplexans

93. The MtDNA of P. falciparum

94. The Plastid of the Apicomplexa