virology lec.3 molecular bio/host cell contraints

Capsid

Protects nucelic acid genome of virus from nucleases

Nucelocapsid

Consists of capsid and genome

Two types of viruses

Naked and enveloped

Genes to code for

Capsid protein

Viral receptor-binding used for attachment to host cell

Genome length varies greatly depending on composition

SsRNA

DsRNA

Giruses

Viruses dependent on host for replication

Where is replication, transcription, and translation localized processes?

In the eukaryotic host cell

DNA replication and RNA transcription happen where?

Within nucleus

Translation of processed mRNAs happen where?

Cytoplasm

Central dogma of molecular biology

DNA →RNA → Proteins

The human genome

Protein-coding sequences: genes

Non-protein-coding sequences

• introns (no coding intervening sequences)

• Promoters and enhancers

• Telomeres

• Non-coding RNAs

Eukaryotic DNA replication

• DNA polymerases synthesize dsDNA

• Copied/read in 3’ → 5’ direction

• Synthesis in 5’ → 3’ direction

• Additional cellular proteins involved: Primases and ligases

Eukaryotic RNA transcription and RNA splicing

• RNA polymerases

• Transcription factors

• Enhancer and promoters

• RNA splicing

Eukaryotic translation

3 steps

• initiation

• Elongation

• Termination

Initiation is Key for versus

Translation and ORFs

• created when introns removed from RNA

• Series of triplets representing amino acids

• End of termination codons (UAA, UGA, UAG)

• Ribosomal frameshfiting and translational readthrough

Cap-dependent initiation of translation

all eucaryotes initiation of translation requires

the presence of the 5’m7Gppp cap of the

cellular mRNA

Cap-independent initiation of translation

viruses synthesize viral mRNA that contain

internal ribosomal entry sites (IRES) that

allow ribosomes to enter 5’ end of viral

mRNA independently of cap.

Ribosomal scanning model

Ribosome scans for the best context of the Kozak’s consensus sequence

Leaky scanning

• Occurs when 40S ribosomal subunit bypasses first AUG to initiate ranslation farther downstream

• Allows multiple viral proteins to be synthesized from a single mRNA

Posttranslational processing of proteins

Modifications serve many functions

• glycosylation (enveloped viruses use carbohydrates on cell surface as receptors for entry to the host cell)

• phosphorylation ( nucleic acid binding)

• proteolytic cleavage (maturation and assembly of viruses)

Why do all viruses use the host’s protein synthesis machinery?

Viral genomes do not have capacity to contain all genes necessary to synthesize proteins

Membranes and endocytosis

Plasma membrane

Phospholipid bilayer

Integral and peripheral proteins

Fluid mosaic model

Why do enveloped viruses steal part of host cell membrane?

For protective envelope

Endocytic pathways

• phagocytosis (“cell eating”)

• Pinocytosis(“cell drinking”)

• Macropinocytosis (growth-factor induced, actin-dependent pinocytosis)

• Receptor-mediated endocytosis

Intracellular membranes and organelles

cytoskeleton

• network of protein filaments or polymers

-microtubules

-actin filaments

-intermediate filaments

The nuclear envelope

Eukaryotic cells contain …

Barriers for viral infections

Molecular hurdles of the host cells

1. Receptors and polymerases

2. Actin remodeling

3. Ribosomes and viral mRNA compatibility

4. The virus-host cell mRNA competition

Hurdle 1: receptors and polymerases

RNA-dependent RNA polymerase are incapable of replicating viral genomes

• Cellular RNA-dependent RNA polymerase are

required for gene regulation or in host defense

mechanisms

• RNA viruses contain a gene that encodes virus-

specific RNA-dependent RNA polymerase

Hurdle 2: actin remodeling

• affects every stage of a viral replication cycle

• Stress fibers

Hurdle 3: ribosomes and viral mRNA compatibility

Eucaryotic host translational machinery can translate only monocistronic RNAs

• Classic or alternative (differential) splicing used by DNA viruses to overcome this hurdle

• Segmented genomes generate separate mRNAs through classic or alternative splicing

• Viral genome transcribed as a single mRNA encompassing several genes, translated into a large precursor polyprotein

Hurdle 4: the virus-host cell mRNA competition

Viruses evolved ways to produce abundant amounts of their own protein to combat expression of numerous cellular genes from host cell

• HSV-1 and poxviruses degrade host mRNA to inhibit cellular translation

Describe the basic structure of a virus

Which DNA genome, upon entry to the cell, can be immediately copied into mRNA?

dsDNA

Why is mRNA synthesis a critical step in the life cycle of the virus?

Because mRNA must be made from all viral genomes