Viral Replication - Lecture notes 5 PDF

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Summary

Define and describe different steps of virus growth curve, i.e. Attachment, Penetration, Un-coating, Synthesis of viral components, Assembly and Release
Describe Strategies of Replication for DNA and RNA viruses
Define and describe different the major events in the eclipse period of vi...

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Viral Replication Learning Objectives Define and describe different steps of virus growth curve, i.e. Attachment, Penetration, Un-coating, Synthesis of viral components, Assembly and Release Describe Strategies of Replication for DNA and RNA viruses Define and describe different the major events in the eclipse period of virus replication • Recognition of viral growth • Infection and death in animals Lab animals or –Hosts • Embryonated eggs –Infection and/or –Death • Cell culture –Cell lysis –Infection Intracellular replication of viruses Because of limited genome composition which obligates viruses to utilize host cell machinery system, they can multiply only in host cells. A single cycle of virus replication range from 6 to 40 hours. One-Step Growth Curve Virus replication cycle Viruses Replication Cycle Attachment –> Penetration –> Uncoating –> Synthesis of –> Uncoating –> Synthesis of viral components –> Assembly –> Release Attachment In order to initiate infection, virions must be bind to cells. Binding occurs between ligands on the surface of the virions (viral attachment proteins) and receptors on the plasma membrane of the target cells, this step is ATP independent.

The lack of correlation between ligands and receptors leading to lack of infectivity of the viruses in different hosts. Cellular receptor Sialic acid –Oligosaccharide side chain of a glycoprotein (or glycolipid) exposed at the cell surface Immunoglobulin super-family –A large group of cell surface and soluble proteins that are involved in the recognition, binding, or adhesion processes of cells. Penetration (Uptake) • Receptor mediated Endocytosis • Fusion with Plasma Membrane In complex viruses like herpesviruses, several viral proteins may serve as attachment proteins and several cellular receptors may be involved, in sequential order, first achieving a loose attachment via one receptor, then irreversible binding via another. Although there is a degree of specificity about the recognition of particular cellular receptors by particular viral ligands, in some cases quite different viruses (e.g., orthomyxoviruses and paramyxoviruses) may utilize the same receptor and, conversely, viruses in the same family or genus may use different receptors. • Receptor mediated Endocytosis • Fusion with Plasma Membrane Receptor mediated endocytosis Fusion Polykarion or Giant cells Events during Eclipse Period Uncoating: – Release of the viral genome from its protective capsid to enable the viral nucleic acid to replicate. • Transcription: Synthesis of viral m-RNA • Translation: The viral m-RNA is translated on cell ribosomes into structural and nonstructural proteins.

Replication of the viral genome: –Providing enough copy of mature genome for assembly Un-coating For viral genes to become available for transcription, it is necessary that virions be at least partially uncoated. Uncoating can occur close by to the cytoplasmic membrane or nucleus membrane. The viral envelope fuses with the lysosomal membrane and the viral nucleocapsid is expelled into the cytoplasm. Release of the viral genome from its protective capsid to enable the viral nucleic acid to replicate. the viral nucleic acid to replicate. Strategies of Replication DNA Viruses Replication of most DNA viruses involves mechanisms that are familiar in cell biology. (transcription of mRNA from double- stranded DNA and replication of DNA using cellular enzymes) RNA Viruses • The situation is quite different for RNA viruses, which are unique in having their genetic information encoded in RNA. (All RNA viruses must have their own RNA polymerases. All negative sense RNA viruses carry their RNA polymerase inside of the virions but positive sense RNA viruses synthesized then use their RNA polymerase from their own genes.) RNA viruses with different types of genomes (single or double stranded, positive or negative sense, non segmented or segmented) have necessarily evolved different routes to the production of mRNA. In the case of positive-sense, single-stranded RNA viruses, the genomic RNA itself functions as messenger RNA, whereas all other types of viral RNA must first be transcribed into mRNA. Because eukaryotic cells are not containing RNA dependent RNA polymerase, all negative-sense, single stranded RNA viruses and double-stranded RNA viruses must carry an RNA-dependent RNA polymerase in their virions. All RNA viruses must carry the gene(s) for RNA polymerases.

Nucleic acid polymerases DNA dependent DNA polymerase – (DNA polymerase) DNA dependent RNA polymerase –(Transcriptase) RNA dependent RNA polymerase –(RNA Transcriptase) RNA dependent DNA polymerase –(Reverse transcriptase) RNA dependent DNA polymerase –(Reverse transcriptase) Transcription (Synthesis of mRNA) The viral RNA of most positive-sense, single-stranded RNA viruses (with the complete structure of mRNA) binds directly to ribosomes and is translated in full or in part without the need for any prior transcriptional step. From all other classes of viral genomes, mRNA must be transcribed in order to begin the process of expression of the infecting viral genome. In the case of DNA viruses that replicate in the nucleus, cellular DNA-dependent RNA polymerase II transcribes the genome to make mRNA. All other viruses require a unique and specific transcriptase that is virus coded and is an integral component of the virion. The double-stranded DNA viruses that replicate in the cytoplasm carry a DNAdependent RNA polymerase. Double-stranded RNA viruses have a specific double-stranded RNA-dependent RNA polymerase. Negative-sense single-stranded RNA viruses carry a specific single-stranded RNAdependent RNA polymerase. Translation (Production of proteins by decoding mRNA produced in transcription) Each monoisotopic mRNA molecule binds via its capped 5' terminus to the 40S ribosomal subunit, which then moves along the mRNA molecule until stopped at the initiation codon. The 60S ribosomal subunit then binds, together with methionyl- transfer RNA and various initiation factors, after which translation proceeds. Classes of Viral Proteins

The non-structural proteins translated from the early transcripts of viral genome include enzymes and other proteins required for the replication of viral nucleic acid, as well as proteins that suppress viral genome include enzymes and other proteins required for the replication of viral nucleic acid, as well as proteins that suppress host cell RNA and protein synthesis. Most of the late proteins are viral structural proteins and are often made in considerable excess. Assembly and Release In non-enveloped DNA viruses a particular protein binds to a nucleotide sequence at one end of the viral DNA known as the packaging sequence; this enables the DNA to enter the capsid. Most non-enveloped viruses accumulate within the cytoplasm or nucleus and are released only when the cell eventually lyses. An infected cell may indeed produce hundreds or thousands of virus particles, but not all of these will be infectious, usually less than 10% are infectious. Maturation and Release of Enveloped Viruses All mammalian viruses with helical nucleocapsids, as well as some with icosahedral nucleocapsids mature by acquiring an envelope by budding through cellular membranes. Budding from Cellular Membranes –Rough Endoplasmic Reticulum Membrane (Arenaviruses) –Cytoplasmic Membrane (Paramyxoviruses) –Nuclear Membrane (Herpesviruses) –Golgi Membrane (Flaviviruses) Cell membrane proteins replace with the Viral proteins during budding Maturation and Release of Enveloped Viruses (cont.) Cell lysis Exocytosis The viruses that budded through inter-cytoplasmic membranes make vesicles containing the virus then migrate to the plasma membrane with which they fuse, thereby releasing the virions by exocytosis membrane with which they fuse, thereby releasing the virions by exocytosis

Sample questions: 1- Copying the Retrovirus' nucleic acid depends on DNA polymerase B) Reverse transcriptase C) Transcriptase D) Endonuclease 2- The structural portions of viruses made: A) Envelope proteins. B) DNA polymerase. C) Viral Capsid . D) A and C 3- Which of the following proteins plays a key role in receptor mediated endocytosis. A)Fusion protein B)Haemagglutinin protein C)Clathrin protein D)Matrix protein...