PE class proteins of M. tuberculosis

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Studies on the implications of PE class proteins of Mycobacterium tuberculosis as determinants of pathogenicity and Immuno-prophylaxis

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PE class proteins of M. tuberculosis

  1. 1. Studies on the implications of PE class proteins of Mycobacterium tuberculosis as determinants of pathogenicity and Immuno-prophylaxis Sameer Tiwari Division of Microbiology L/O/G/O
  2. 2. M. tuberculosis 1882 Robert Koch 2011 2011 ~9 million new cases ~2 million deaths worldwide ~2 million new cases in India
  3. 3. M. tuberculosis genome 4,000 genes 40% orphans 100 highly homologous PE/PPE genes Cole et al. (1998) Nature 393: 537-544
  4. 4. General Classification of M. tuberculosis genes
  5. 5. PE/PPE proteins 100 highly homologous genes with signature sequence of Pro-Glu (PE) amino acid near amino terminus.
  6. 6. PE-PGRS (polymorphic GC rich repetitive sequences) 1901 aa 550 aa 175 aa Rv3508 Average Rv0742 Larger proteins domain extremely rich in Gly (~ 40%) & Ala (~ 25%) residues PGRS domain occur as repetitive sequences (repeated >30times within domain)
  7. 7. Properties of PE proteins Extensive functional redundancy Close genomic & evolutionary association with ESX regions Highly immunogenic
  8. 8. Classification of PE/PPE protein families Sequence variation between M. tuberculosis H37Rv & M. bovis BCG Cole et al. (1998) Nature 393: 537-544
  9. 9. Role of PE_PGRS Homeostasis of mycobacterial cell Intracellular survival Multiplication within its chosen environment
  10. 10. Why to study? Unique gene family No counterpart in bacteria Potential source of antigenic variation No strict correlation with pathogenic strains Little is known
  11. 11. Known facts….. Role in virulence (Ramakrishnan et al.) pathogenic Mycobacterium marinum expresses two PE_PGRS genes in granulomas of infected frogs and that M. marinum mutants containing deletions in these genes replicate poorly in macrophages Several PE_PGRS proteins have been localized to the cell surface, and allelic diversity among M. tuberculosis isolates indicates that they could serve as a source of antigenic variation for this pathogen. Microarray analysis has also suggested that variable expression of certain PE_PGRS genes occurs under conditions that mimic in vivo pathogenesis such as nutrient depletion, low pH or oxidative stress
  12. 12. Objectives Evaluation of complete cellular immune responses incurred by these proteins Delineation of protective immune response of the proteins, upon challenge with MTB Regulation of PE genes in mycobacteria in Culture grown, persistent, Hypoxic and from infected macrophages/ from infected lungs. .
  13. 13. What we Achieve? Characterization of PE_PGRS gene expression mechanisms of pathogenesis of mycobacteria in macrophages Vaccine candidates as well as virulence factors. Quantification of the amounts of CFP-10/ESAT-6 secreted into the macrophages during early stages of infection will give an insight of their role in virulence. Identification of domains involved in binding to cell surface or other intracellular components will help in designing targets against these proteins.
  14. 14. Impact on antigen-presentation pathways Ensuing host immune responses, and Also provide a mechanism for generating antigenic diversity in mycobacteria
  15. 15. Research Plan 1 2 3 4
  16. 16. 5 6 7 8
  17. 17. Conclusions The PE proteins/PPE functions to be the rich source of protective antigens involved in antigenic variation and immune-pathogenic importance. PE/PPE genes involve in host-pathogen interactions, such as antigenic variability, virulence, and persistence of the bacillus. Their surface-exposed domains are also involved in the shaping of the bacterial cell structure. These proteins are capable of forming heterodimers which are secreted and thought to play a role in signal transduction. The study will investigate the characteristics of PE3 (Rv0159c) & PE4 (Rv0160c) and their associations with ESAT-6 & CFP-10 and finally their interactions with macrophage.
  18. 18. Laboratory: Microbiology Division, CDRI, Lucknow Equipments Bio-hood, CO2 Incubator, Refrigerated Centrifuge, Microfuge, Fluorescent Microscope, Incubator Shaker, Water Bath, Incubators, -860C Deep freezer, Power supplies, Gel apparatus, Gradient Thermal Cycler, Gel Documentation System, Sonicator, Protein Purification System, 2-D Gel Electrophoresis System, ABI Real Time PCR. Other resources Animal House and BSL-III facility
  19. 19. Thank you for your patience….

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