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30511 prions.ppt
- 1. PRIONS THE INFECTIOUS PROTEINS Paras Yadav*,Jaspreet Singh Arora*, Sachinandan De*, Tirtha Kumar Datta*, Surender Lal Goswami*, Aarti Bhardwaj$, Shalini Jain# and Hariom Yadav# *Animal Biotechnology, #Animal Biochemistry Division, National Dairy Research Institute, Karnal-132001, Haryana, India $Meerut Institute of Engeenering and Technology, Meerut, U.P., India
- 2. • Stanley B.Prusiner coined the term proin from Proteinaceous infective particle and changed to prion to sound it rhythmic. • Prion diseases were caused by misfolded proteins. • Elucidated the gene and mechanism by which wild type protein bring about the clinical disease. INTRODUCTION
- 3. • Kuru • FatalFamilial Insomnia (FFI) • Creutzfeldt-Jakob disease (CJD) • Scrapie • Bovine Spongiform Encephalopathy (BSE) • Chronic Wasting Disease (CWD) Prion Diseases Human Animal
- 4. Classification of priondiseases • Infectious/Exogenous – e.g., Kuru, BSE (mad cow disease), Scrapie – Spread by • Consumption of infected material. • Transfusion. • Sporadic • Familial/Hereditary – Due to autosomal dominant mutation of PrP.
- 5. Differences between cellularand scrapie proteins PrPC PrPSC Solubility Soluble Non soluble Structure Alpha-helical Beta-sheeted Multimerisation state Monomeric Multimeric Infectivity Non infectious Infectious Susceptibility to Proteinase K Susceptible Resistant
- 6. Steps in thebiosynthesis of PrPc
- 7. Post-translational processing ofPrP S S A C B
- 8. ER Golgi Endosome Cellular traffickingand cleavage of PrP
- 9. Mechanism of Internalizationof PrPC
- 10. Hypothetical model fora PrPc receptor
- 11. Model for thefunction of LRP- LR as the receptor for PrP HSPG Dependent binding domain Direct binding domain (aa 161-179) LRP/LR PrP BDI (aa 144-179) Heparan sulfate chain (HS) Sulfated domains HSPG Proteogl ycan moiety GPI BDII (aa 53-93)
- 12. Cell Culture Systemsfor Prion Propagation
- 13. Sequence of prionprotein
- 14. PrPC + Cu(Copper) Antioxidant activity Resistance to oxidative stress Prevent neuronal dysfunction (Brown et al., 2002) • Antioxidative Functions of Prion protein • Other functions
- 15. Models for theconversion of PrPc to PrPsc
- 16. Time taken forTransformation of mutant PrP c to a PrP Sc state Detergent insoluble PIPLC- resistant Synthesis of Mutant PrP c Protease- resistant <10 min BFA 180 C 0.5-1 hr 1-6 hr Endoplasmic Reticulum Plasma membrane/ Endocytic Pathway
- 17. Effect of conformationof PrP on Pro K
- 18. Model of thecellular pathways involved in generation of PrPSc
- 19. Proposed model ofPrPc aggregation and induction of CtmPrP
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- 21. Mechanism of PrPsc inducedapoptosis
- 22. They generatea C-terminal fragment(C2) which has molecular weight of 27-30 kDs. Increase in intracellular levels of Calcium increase production of terminal fragments . Calpastatin prevents production of C2. Inhibitors of lysosomal proteases has no effect on C2 production. Telling et al.,2004 What are Calpains?
- 23. It wasproposed that prion-associated toxicity involves altered trafficking of PrP c . Inhibition of ubiquitin-proteasome system(UPS). Deposition of aggresomes of PrP Sc in nerve cells. Induction of Apoptosis with activation of Caspase 3 and Caspase 8. Complete molecular basis for neuronal death is not known. Aggregates of over expressed PrP c does not cause cell death. Tabrizi et al., 2005 Role of Caspases
- 24. The AGAAAAGA Palindromein Prion Generation Factors Responsible for Prion Propagation Norstrom & Mastrianni, 2005
- 25. Factors Responsible forPrion Propagation cont… • PrP c association with lipid rafts in the early secretory pathway. • Creutzfeldt–Jakob disease (CJD) in Libyan Jews, linked to the E200K mutation in PRNP (E200KCJD).
- 26. E.g. Hsp70, GroEL Modelfor chaperone-supervised PrP conversion
- 27. Phospholipase A2 Inhibitorsprevent prion replication. Platelet-activating Factor Antagonists also inhibits prion replication. Bate et al.,2004 Drugs which share a N-benzylidene-benzohydrazide core structure. Trimethylamine N-oxide (TMAO), can prevent formation of PrP Sc . Factors that prevent Prion Replication Bertsch et al.,2005 Bennion, 2004
- 28. Gross and MicroscopicChanges Grossly there is Cortical atrophy and ventricular dilatation may also be present. Gross changes
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- 30. • Gliosis withinplaques. • Loss of oligodendrocytes within plaques. • Axons usually remain intact in plaques. • Both CD4+ and CD8+ lymphocytes are present in active lesions. (Kretzschmar et al.,1996, Wilesmith et al., 1997). Other microscopic changes
- 31. Diagnosis can bemade by: 1. Clinical signs and Symptoms. 2. Detection of Scrapie Associated fibrils. 3. Detection of Abnormal Prion protein (PrPsc) by Western blotting. 4. Two dimensional Gel Electrophoresis. 5. Imunodiagnosis of Prion disease. 6. Bioassay in Mice. Diagnosis Scrapie Associated fibrils.
- 32. PrPC PrPSC PrPC Mechanism of plaqueformation
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- 35. Molecular hallmark ofthe disorder is the accumulation of abnormal prion protein(PrP Sc ). Physiological functions of cellular prion protein (PrP c ) is not clear. Identity of intracellular compartment where PrPc to PrPSc occurs is not established. Prion peptide of 106-126 residues is found to be neurotoxic. Studies on prion protein will open the avenues for treatment of other neurodegenerative disorders. Conclusion