Revenge of the phages


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Revenge of the phages

  1. 1. Revenge of the phages: defeating bacterial defences Raffia Siddique National University Of Sciences And Technology Julie E. Samson, Alfonso H. Magadán, Mourad Sabri and Sylvain Moineau NATURE REVIEWS | MICROBIOLOGY
  2. 2. Abstract • Constant fight b/w bacteria and bacteriophage • What bacteria has? strong defense • What bacteriophage has? 1. adsorption inhibition, 2. restriction–modification 3. CRISPR–Cas 4. abortive infection.
  3. 3. Bacterial anti-phage system 1. Inhibition of the phage attachment to the host cell surface receptors 2. Cleavage of the invading phage genome 3. Abort phage infection by making cell suicide
  4. 4. Access to host receptors • Adsorption • Interaction b/w RBP present on the tail of bacteriophage and the bacterial cell surface receptors. • bacterial receptors are the surface proteins, polysaccharides and LPSs
  5. 5. Adapting to new receptors Tailed phages are able to modify their RBPs to acquire novel receptor tropism 1- Evolving a new RBP 2- Gaining access to masked receptors 3- Stochastic expression of RBPs
  6. 6. 1-Evolving a new RBP Receptor of bacteria modified through mutation or replaced by a different molecule, then the phage is unable to adsorb to the host. To infect this receptor- modified host, the phage can evolve to target a new receptor, by acquiring mutations in the genes encoding the RBP or tail fibres For example, mutations in the gene encoding protein J of coliphage λ enables this RBP to recognize a new receptor, OmpF, in addition to the cognate receptor, LamB.
  7. 7. 2- Gaining access to masked receptors surface molecules (for example, capsule or exopolysaccharide (EPS)) at the receptor site can limit or prevent phage access if the phage possesses a depolymerase, it can degrade these substances to unmask the receptor.
  8. 8. 3- stochastic expression of RBPs Phages can modify their RBPs in a manner that allows them to interact with a surface component that is expressed by the host at that time This can be achieved by mutations in the gene encoding major tropism determinant (Mtd) for the Bordetella spp. phages, by proteolytic cleavage of tail fibres for Lactococcus lactis phages TP901-1 and Tuc2009 or by duplication of a His box element in the coliphage T4 tail proteins.
  9. 9. Adapting to new receptors
  10. 10. Battling restriction–modification systems • When a phage genome manages to enter the cell, it can still face a myriad of intracellular antiviral barriers. • Restriction– modification (R–M) systems are widespread in bacteria. These systems typically use a restriction endonuclease (REase) to cut invading foreign DNA at specific recognition sites.
  11. 11. These systems are classified into four types based upon; 1-structure, 2- recognition site 3- mode of action
  12. 12. phage anti-restriction strategies Passive mechanisms of phage evasion. The phage genome can be modified by the host methyltransferase (Mtase) Some phages have few restriction sites in their genome, or these sites are too far apart to be recognized by the host restriction endonuclease (REase), thus preventing targeting
  13. 13. Active mechanisms of phage evasion A phage can co-inject proteins such as DarA and DarB (of phage P1) with its genome to bind directly to the phage DNA and mask restriction sites. A phage protein (such as Ocr of phage T7) mimics the target DNA and sequesters the restriction enzyme. Ocr binds to both the MTase and the REase of the type I restriction–modification system EcoKI and inhibits its activity. A phage protein such as Ral of phage λ can activate the activity of the MTase and thereby accelerate protection of the phage DNA
  14. 14. Evading CRISPR–Cas systems These are the part of bacterial immune system which detects and recognize the foreign DNA and cleaves it. 1. The CRISPR (clustered regularly interspaced short palindromic repeats) loci and 2. Cas (CRISPR-associated) proteins can target and cleave invading DNA in a sequence-specific manner. Spacer=The direct repeats in a CRISPR locus are separated by short stretches of non-repetitive DNA called spacers that are typically derived from invading plasmid or phage DNA. Protospacers = The nucleotide sequence of the spacer must be similar to a region in the phage genome called a protospacer in order to recognize and subsequently block phage replication.
  15. 15. 1. Adaptation The direct repeats of the CRISPR locus are separated by short stretches of non-repetitive DNA called spacers, which are acquired from the invading DNA of plasmids or viruses in a process known as adaptation. 2. biogenesis of CRISPR RNA The CRISPR locus is transcribed as a long primary pre-crRNA transcript, which is processed to produce a collection of short crRNAs (a process referred to as biogenesis of crRNA. Each crRNA contains segments of a repeat and a full spacer and, in conjunction with a set of Cas proteins, forms the core of CRISPR–Cas complexes. These complexes act as a surveillance system and provide immunity against ensuing infections by phages or plasmids encoding DNA complementary to the crRNA. 3. Interference On recognition of a matching target sequence, the plasmid or viral DNA is cleaved in a sequence-specific manner (known as interference). The nucleotide sequence of the spacer must be identical to a region of the viral genome or plasmid (known as the protospacer) for the CRISPR–Cas complex to block replication of the foreign element.
  16. 16. Evading CRISPR-Cas System Mutations in the phage protospacers or in the protospacer- adjacent motif (PAM) render the phage insensitive to the interference step of the CRISPR–Cas system,
  17. 17. The phage-encoded anti-CRISPR protein blocks the interference step by preventing the formation or blocking the action of the CRISPR–Cas complexes.
  18. 18. The CRISPR–Cas system of Vibrio cholerae phages. target an uncharacterized antiphage system of the V. cholerae host; this system is contained within a locus resembling a phage-inducible chromosomal island (PICI), referred to as a PICI-like element (PLE). The spacers in the phage CRISPR locus are complementary to PLE sequences, and the CRISPR machinery is then able to specifically target this genetic element and inactivate it.
  19. 19. Escaping abortive-infection mechanisms Abi systems inhibit various steps of the phage replication cycle Abi systems typically consist of a single protein or protein complex and are often found on mobile genetic elements, such as prophages and plasmids, Toxin–antitoxin (TA) systems represent a subgroup of Abi systems that lead to bacterial death following activation by phage infection. TA systems are composed of a toxin and a neutralizing antitoxin that renders the toxin ineffective during normal bacterial growth The antitoxin is more labile than the toxin, so when a stress is encountered, the antitoxin is degraded and the toxin is free to induce either dormancy or cell death
  20. 20. During normal bacterial growth, the antitoxin neutralizes the toxin, thereby preventing bacterial cell death. During phage infection, an imbalance in the toxin–antitoxin ratio or inactivation of the antitoxin results in liberation of the toxin, which is free to act on its target and inhibit bacterial growth. This growth inhibition also leads to the abortion of phage infection.
  21. 21. Thanks Questions?