Objectives
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Objectives. To understand the general principles involved in RNA replication discussed in Chapter 6 pages 175-192 To use the following + stranded RNA viruses as examples and to understand the different processes that during replication by RNA viruses:- Picornavirus Togaviruses Flaviviruses.

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Objectives

Objectives

  • To understand the general principles involved in RNA replication discussed in Chapter 6 pages 175-192

  • To use the following + stranded RNA viruses as examples and to understand the different processes that during replication by RNA viruses:-

    • Picornavirus

    • Togaviruses

    • Flaviviruses


General principles

General Principles

  • De novo initiation:

    • RNA polymerase

    • RNA template

    • The initiating NTP and a second NTP


General principles1

General Principles

  • Primer Dependent Initiation

    • Protein priming

    • Capped RNA fragments

  • All synthesis begins with the formation of a complex of polymerase, template-primer, and initiating NTPs


General principles2

General Principles

  • Template specificity

    • Appears to be very strongly related to :

      • Affinity of RNA polymerase for the initiating NTP

      • The sequence

      • The structure of the viral RNA molecules

      • Encoding proteins that bind to genomic RNA segments which then activates an endonuclease that cleaves host cell RNAs.


More facts

More facts

  • Unwinding the RNA template

    • Encoded in genome of ds RNA viruses

    • Prevent base pairing between template and new strand.

  • Role of host cell proteins

  • Different RNA polymerases for mRNA synthesis and genome replication

  • Switching from mRNA to genome replication


Positive strand rna viruses

Positive Strand RNA Viruses


Picornaviruses picornaviridae

PICORNAVIRUSES   (PICORNAVIRIDAE)

  • Properties

    • These are small (28nm),

    • naked

    • icosahedralviruses

    • RNA is single-stranded, plus sense, polyadenylated.

    • functions as mRNA immediately upon infection

    • E.g poliovirus


Adsorption and penetration

Adsorption and penetration

  • A viral protein recognizes a receptor on the host cell membrane (this is important in the tropism of virus).It seems that binding to the receptor alters capsid structure in some way, a channel forms across the cell membrane and the RNA is released into cytoplasm. The mRNA is now available for translation.


Synthesis of viral proteins

Synthesis of viral proteins

  • Poliovirus virion RNA functions as an mRNA but does not have the methylated cap structure typical of eucaryotic mRNAs

  • It has a "ribosome landing pad" (known as the internal ribosome entry site or IRES) which enables ribosomes to bind without having to recognize a 5' methylated cap structure

  • Most host cell translation is cap-dependent, so this inhibits a lot of host protein synthesis but not viral protein synthesis. 


Synthesis of viral proteins1

Synthesis of Viral Proteins

  • The mRNA is translated into a single polypeptide (polyprotein), which is cleaved.

  • The cleavages occur before translation is complete ( i.e. on the nascent (=growing) chain) and are carried out by virally coded proteases.

  • Products of cleavage include:

    • An RNA polymerase (replicase)

    • Structural components of the virion

    • Proteases


Rna replication

RNA replication

  • We now have newly made viral proteins to support replication.

  • Viral RNA polymerase copies plus-sense genomic RNA into complementary minus-sense RNA requiring:

    • VPg (Viral Protein genome-linked)

    • Viral RNA polymerase (replicase)

    • Certain Host proteins

  • VPg may act as a primer for RNA synthesis, this would explain why it is at the 5' end of all newly synthesized RNA molecules

  • New minus sense strands serve as template for new plus sense strands

  • Again, poliovirus RNA polymerase and VPg are needed. VPg is linked to the 5' ends of the new plus sense strands (again, it probably functions as a primer).

  • The new plus strand has three alternative fates:i. It may serve as a template for more minus strandsii. It may be packaged into progeny virionsiii. It may be translated into polyprotein (In this case VPg is usually removed prior to translation)


Assembly

Assembly

  • When sufficient plus-sense progeny RNA and virion proteins have accumulated, assembly begins.

  • Particles assemble with VPg-RNA inside and 3 proteins in the capsid [VP0,1 and 3].

  • VP0 is then cleaved to VP2 and VP4 as the virions mature and these mature virions are infectious.

  • Virionsare released following cell lysis.

  • Excess capsids are formed and inclusion bodies may be seen in the cytoplasm.


Objectives

  • NOTE: THE ENTIRE LIFE CYCLE OCCURS IN THE CYTOPLASM

  • THERE IS NO DIVISION INTO EARLY AND LATE GENE EXPRESSION


Non segmented negative strand viruses

NON-SEGMENTED NEGATIVE STRAND VIRUSES


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