Emerging

Sue Delos CWA Jan 23, 30, and Feb 6 2018 Part I Introducon to Virology

What is a ? Viral Lifecycle Common Needs Unique soluons Transmission Routes Classificaon Methods of Visualizing Virion Components Structures Viral Envelopes Funcons of Structural Funcons of Nonstructural Proteins Part II Virus-Host Interacons, Vaccines and Ihibitors Part III Emerging Viruses

The Immune System Zoonoses and Why They Occur Innate Influenza, A Respiratory Virus Adapve Ebola, A Blood-Borne Virus Viral Countermeasures Zika, A Mosquito Transmied Virus Vaccines Vaccines Improve Life Expectancy Dependence upon Herd Immunity Passive Vaccines Acve Vaccines Inacvated Virus Subunit Vaccines Subunit Vaccines Anvirals Requirements for success Potenal Targets The Path of Drug Discovery The HIV experience The HCV experience

Emerging Viruses

Part I Introducon to Virology What is a Virus?

A virus is an organism with two phases. What would you do if you were a parasite that could only reproduce inside a cell?

Cell A Virus is an Organism with Two Phases Intracellular Extracellular

Disassembly Stable parcle Reproducon New, stable parcle assembly and release Common Needs 2 phases of viral lifecycle: extracellular and intracellular

• Find permissive cell • Synthesize viral mRNA • Bind cell (transcripon) • Penetrate membrane • Synthesize viral proteins • Uncoat (translaon) • • Reprogram Cell for own Replicate genome replicaon • Assemble and mature • Establish replicaon virion center • Release stable virion (lysis or budding)

Structural Funcons Nonstructural Protein Funcons Unique Soluons

ENTRY: • Uncoang • Mulple receptors • Replicaon site • Mulple modes of entry • Assembly • Surface proteins • Egress • Site of membrane • (Cell-to-cell) penetraon transmission Sites of Viral Entry into the Host

V. Raconelli, MID 31 In humans

Viruses cause a wide range of misery

Image credit: "Prevenon and treatment of viral infecons: Figure 1, by OpenStax College, Biology, CC BY 4.0. Modificaon of original work by Mikael Häggström.

Viruses are Nature’s Nanoparcles

(too small to be observed by tradional microscope)

1 μM = 1000 nM The Tools of Viral Structural Biology

• X-ray Crystallography (must have crystals) • But most viruses and their proteins are flexible and do not crystallize

• Electron microscopy (can see parcles)

• CryoEM and Tomography (structures of flexible virions and proteins)

• NMR (structures of regions within protein) The Structural Proteins Capsid Funcons

• Encases and protects c material • Contains the budding funcon • Maintains stable structure during transit to new host/cell • Undergoes orderly disintegraon during entry and travel to replicaon site • For non-enveloped viruses, the capsid is the outer shell, contains the receptor binding site, and is the target for immune responses (cores have icosahedral symmetry) Icosahedral Capsid Structures HPV 16 VLPs

Viral Envelopes

• Membranes derived from Host Cell Membranes • Contain Viral Proteins termed Surface Proteins • Surface Proteins – bind target cells – define internalizaon pathways – Fuse viral and cellular membranes – Are targets of the immune response • Because usually contain budding informaon, enveloped viruses are promiscuous for surface protein incorporaon

Receptors Come in Many Forms What we study: How enveloped viruses bind to, (enter), and fuse with cell membranes

Any thoughts on why we study this?

INFLUENZA HA: Conformaonal Changes During Fusion

Native: First change: Activated: Post-fusion: metastable Head-group separation prehairpin 6-helix bundle structure During fusion, fusion proteins convert from unique conformaons to 6-helix bundles

Common Viral Fusion Mechanism

target membrane

virus membrane

Model is for a Class I (flu HA, HIV , Ebola GP) Viruses Exploit All Modes of Internalizaon

Adenovirus Picornaviruses Influenza, VSV, SV40, (induces) ?, Influenza, SFV polyoma, Sendai? MLV?

Journal of General Virology (2002), 83, 1535–1545. Replicaon

• DNA viruses replicate in the nucleus • Most RNA viruses replicate in the cytoplasm • reverse transcribe their RNA into DNA which then integrates into the host DNA for replicaon • Viral polymerases make mRNAs • Many viral proteins are synthesized as single polymer which must be cleaved into separate proteins for virus funcon • Unique viral proteins include reverse transcriptases (RT), (IN), polymerases (Pol), and

All Virions Complete a Common Set of Assembly Reacons

Formaon of individual structural Acquision of an envelope units of the protein shell from one or several viral proteins Release from host cell Assembly of the protein shell by appropriate, and somemes Maturaon of the virus parcles variable, interacons among structural units

Selecve packaging of the nucleic Common to all viruses acid genome and other essenal virion components Unique to enveloped viruses

All Steps in the Viral Life Cycle are Targets for Intervenon

The HIV Life Cycle Quesons? Emerging Viruses

Part IIa Virus-Host Interacons The Two Arms of the Immune Response: Innate vs Adapve Immunity Innate Adapve • Primive: In all mulcellular • Only in vertebrates organisms • Directed toward classes of • Directed toward specific molecules epitopes on molecules • Effectors broadly reacve • Effectors highly specific • No anamnesc response • Memory persists • Effectors = epithelial cells, • Effectors = lymphocytes, phagocytes, endothelial Angen Presenng Cells cells, fibroblasts (APCs) Interferons (IFN) are major components of innate immune response B-Cells

Make Anbodies Make Cytokines to Smulate Innate Immunity Steady-state Immune Readiness Regulate Immune Response Smulate T-cells

NIH Publicaon No. 03-5423 Three broad categories of T cells

Helper T cells augment the immune response by recognizing the presence of a foreign angen and then smulang anbody producon and producing cytokines that “turn on” or acvate other T cells.

Regulatory T cells funcon in an opposite manner: they dampen or turn off the immune response . Cytotoxic or “killer” T cells directly aack and destroy cells bearing angenic material.

NIH Publicaon No. 03-5423

Viral Interference with Early B-cell Acvaon : 10.1038/nr, 2017.133i doi , Iannacone and M. Kuka M. Viral Interference with Anbody Producon and High Affinity Maturaon

M. Kuka and M. Iannacone, doi: 10.1038/nr, 2017.133i M. Kuka and M. Iannacone, doi: 10.1038/nr, 2017.133i M. Kuka and M. Iannacone, doi: 10.1038/nr, 2017.133i Quesons?