T4 Life Cycle

T4 phage Lytic cycle

The T4 bacteriophage is among the largest phages, encoding roughly 200 genes. It is a dsDNA virus which infects E. coli. T4 has been extensively studied and has a rich history in the advancement of genetics. Some of the first essential ideas of genetics came from studies using T4 including: the basis of genetic code, even ribosomes, mRNA, and the codon. It is surprising that science has learned so much using a virus which is extremely complex. Nonetheless, we owe much of our current knowledge to T4. The following stages of the life cycle are a brief outline of what is known about T4. [16]

T4 body plan

Complex life cycle of a complex capsid

  1. Adsorption – T4 adheres with tail fibers to lipopolysaccharide [4] and tryptophan [16] receptors on the bacterial host.
  2. Penetration – the bacteriophage penetrates the bacterial cell by bringing the base plate into contact with the cell surface, this allows for a conformational change which causes the sheath to contract piercing the cell membrane allowing the viral core to enter the cell, and release the dsDNA viral genome. [4] Thus begins the complex behavior of this viral genome. Some of the first genes which are transcribed, called immediate-early, encode enzymes which break down host DNA. Host DNA is broken down in order to use host nucleotides to produce more viral DNA. In fact, the breakdown of host-genome DNA can be visualized under a microscope in the few several minutes following infection. Many T4 genes are involved in nucleotide metabolism because T4 uses 5-hydroxymethycytosine instead of plain old cytosine. [16]
  3. Replication – Delayed early and middle genes encode the 20 proteins which are involved in viral replication. In fact, T4 encodes nearly all of its own replicative machinery. There are two distinct replicative processes of T4 which are outlined below.Stage 1 – Replication occurs in a bidirectional manner with multiple origins of replication within the genome. The first several rounds of replication are initiated by RNA primers synthesized by the host RNA polymerase. These RNA primers can travel to their complimentary region in dsDNA and displace the other strand to produce a structure called an R-loop. The attached RNA can now act as a primer for the leading strand of DNA replication. The lagging strand is then synthesized using the replicative helicase, gp41. DNA replication is finished by gp30 which is the T4 encoded ligase as well as the host DNA ligase. Several minutes after infection, host RNA primers cannot be used because the promoting recognition specificity is altered on the host RNA polymerase and recombinant-dependent replication (RDR) is favored. The mechanism of RDR is outlined in stage 2 [16]
    Stage 2 – At this point the virus uses its own replicative machinery to transcribe its late genes with a process known as the gp45 sliding clamp model. The primers for leading strand synthesis are recombination intermediates instead of RNAs made by RNA polymerase.[16]

In this particular model, the clamp loading proteins gp44 and gp62 help load the gp45 clamp onto the viral DNA. At the same time the viral DNA polymerase (gp43) synthesizes okazaki fragments and the viral helicase (gp41) helps keep the strands separate.

Once the okazaki fragment is synthesized, gp43 unbinds from the gp45. The gp45 clamp can now interact with gp33 on RNA polymerase and initiate transcription of late genes.

Another gp45 clamp is loaded onto the DNA strand to interact with gp33 on RNA polymerase and initiate another round of replication. This process can continue until long strands of concatemers are made and ready to be packaged into empty capsid head complexes.

4. Assembly – Once late genes are expressed, the (1) viral base plate is first assembled, this then attaches to the (2) tail and (3) tail fiber proteins. These three different protein pathways combine to form a mature T4 phage capsid. DNA is packaged into the mature capsid protein by packaging and cutting the concatemers using a terminase complex found at the end of the concatemerized DNA strand. The terminase complex binds to the capsid head and moves DNA into the empty capsid head. The capsid also encases necessary enzymes for future infections such as virally encoded DNA polymerase.[16]

5. Release – The viral encoded enzyme holin (gpt) creates the holes in the inner membrane of the bacterial host-cell to allow lysozymes to exit and degrade the peptidoglycan cell wall. Cell lysis subsequently follows releasing a shower of bacteriophage progeny into the extracellular space.[16]

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