Virus, Viroids, and Prions PDF

10/22/24 Origin Story -- Cells Cellular life arose one time Evolved linearly, with branches Can trace a common heritage and map incremental evolution through all cellular life forms! 5 Origin Story -- Viruses Viruses arose many times, from many diffe...

10/22/24 Origin Story -- Cells Cellular life arose one time Evolved linearly, with branches Can trace a common heritage and map incremental evolution through all cellular life forms! 5 Origin Story -- Viruses Viruses arose many times, from many different cells Using many different strategies and components Usually restricted to a limited range of host cells All use nucleic acid chains (some form of RNA or DNA) to store information à so they can evolve Use same genetic code, processes But small size of genome limits amount of information that can be stored! All use proteins, but which proteins and how used vary widely There is an elaborate naming convention, but less valuable than the similar system for cellular life. à We will focus on characteristics, common names, and/or small groups of related viruses. 6 3  10/22/24 Informal Taxonomy Can Be Important Groups of unrelated viruses sharing roues of infection Enteric Viruses – Oral/Fecal route Respiratory viruses – infecting through inhalation Arboviruses (“Arthropod Borne”) – vector transmission STI/STD – Sexual contact Zoonotic – Animal reservoir, transmitted to humans either directly or through vector 7 Two general stages Virion -- Consist of Protein and Nucleic acid Infective stage (e.g., “poison”) Structure mostly protein (some have membrane envelop) All carry some form of RNA or No measurable metabolism! Behaves more like a complex chemical or protein Can crystalize, precipitate in Ethanol, store for years – still infective Infected cell Mostly hijack cell systems. In many cases, only the nucleic acid gets inside cell! There is no replication, transcription, or translation without the host cell 8 4  10/22/24 Structure – Protein Capsid Small size à limited DNA à limited number of proteins But need to build structures that can hold all the viral DNA How balance competing goals? To build capsid, viruses generally code for one or a few “lego-like” proteins that fit together to form larger structures Means that there are limited number of optimal structures Very different viruses might have “similar looking” structures. 9 Three basic shapes of Viruses Icosahedral 20 flat triangles Helical Subunits in a helix of various lengths Complex viruses (mainly some bacteriophage) Both shapes, plus ancillary proteins. 10 5  10/22/24 Viron Structure Example 1 Virion (viral particle) is nucleic acid surrounded by a capsid (protein coat) Capsid composed of simple identical subunits called capsomeres Capsid plus nucleic acids called nucleocapsid 11 Viron Structure Example 1 Some viruses are surrounded by a lipid bilayer obtained from the host cell. (can have protein structures between virion and membrane envelope.) Called “Enveloped Viruses” Exposed membrane is essential for infection à makes them more susceptible to disinfectants like alcohol or soap and water. 12 6  10/22/24 First Focus: Bacteriophages – well understood laboratory systems Three general types of host/phage relationship Lytic (aka, virulent) phages Always a destructive infection, rupturing the host cell Temperate phages Any given infection could go either lytic or “lysogenic” (latent stage) Filamentous (or helical) phage Continuous production of phage from a “living” host cell Two other terms? Productive infection à new virus particles are produced Latent state à viral genome along for the ride, for now… 13 Diagram of three types of Bacteriophage infection 14 7  10/22/24 Lytic Phage Infection – Example T4 T4 is a complex phage with double stranded DNA. Attachment by tail fibers Attach to susceptible host outer membrane. Only attaches to living host (not membrane fragments) Genome Entry Phage lysozyme makes small hole in cell wall Tail contracts, injecting only the DNA into the bacterium (Note that if we were to break open cell at this point, no infective virus particles are found!) 15 Lytic Phage Infection, cont. Synthesis stage – As genome is inserted, set of early genes (using host promoters!) are transcribed and translated. Includes a viral nuclease that destroys the host cell genome! Now a zombie cell! Also makes new DNA polymerase to start Phage replication. Replicates DNA and starts making capsid parts for assembly. 16 8  10/22/24 Lytic Phage Infection, cont. Assembly stage – Phage parts self assemble, including packaging DNA inside icosahedral head While still inside host, assemble hundreds of mature, infective virons! If we were to lyse cell at this point, there would be numerous infective particles. 17 Lytic Phage Infection, cont. Release stage – large amount of T4 lysozyme is synthesize and exported to outside of cytoplasmic membrane. Breaks down peptidoglycan, causing cell to burst and freeing the mature phate Mature phage drift through the fluid, looking for a new, susceptible host. 18 9  10/22/24 Temperate phage add a new wrinkle. Example: Lambda Phage Double stranded DNA phage that infects E. coli. Attachment and Genome Entry occur similarly to T4 But, after entry, the linear chromosome circularizes and makes a biochemical coin toss! Sometimes it decides to continue down the “Lytic” pathway, similarly to T4 19 Lysogenic Infection Other times, the injected genome makes an “integrase” that inserts the lambda DNA into the bacterial genome. Integrated phage DNA à prophage Bacterial cell carrying phage DNA à lysogen Prophage replicates along with host chromosome and is passed on to all progeny. Most phage genes are blocked from synthesis by a phage encoded repressor -- for the time being… 20 10  10/22/24 Lysogenic Infection cont. A prophage can remain in latent state indefinitely But, about every 10,000 division, prophage makes an enzyme that excises the phage genome and initiates a lytic infection. Happens about once per 10,000 divisions of lysogen Also triggered by damage to the host. If phage detects that bacteria is under stress (e.g., UV light), quickly replicates more phage and kills host. Called phage induction; allows phage escape damaged host 21 Beneficial (to the Bacterium) Consequences of Lysogeny Immunity to Superinfection Lysogen is immune to superinfection (infection by same phage) Repressor maintaining integrated prophage also binds to operator on incoming phage DNA, prevents gene expression Lysogenic Conversion— lysogen may show change in phenotype due to prophage DNA Toxins encoded by phage genes; only strains carrying prophage produce the toxins 22 11  10/22/24 Filamentous (Helical) Phage – Example M13 Single Stranded (+ strand) DNA genome, filamentous phage, that infects E. coli F-pilus When genome inserted, host polymerases immediately complete the second strand (- strand) of DNA, creating a double stranded Replicative Form (RF) of DNA RF is template to make new phage proteins that makes copies of the (+) strand for virus genomes and makes mRNA from (-) strand to make phage enzymes and capsomeres. 23 Assembly M13 Capsomeres insert into cytoplasmic membrane Phage “protein pores” assemble that span cell wall and outer membrane. DNA copy extruded through pore while being coated with capsomere Continuously produces phage while keeping cell alive! 24 12  10/22/24 SEA-PHAGES (HHMI and Dr. Graham Hatfull) UTEP part of the Alliance! Freshman students Isolating and sequencing bacteriophage Started as science study Grew to a model of the potential of phage therapy. Isolate phage that kill specific bacteria that are resistant to antibiotics. 25 Animal Viruses Informal classification given to viruses that infect cells in animals, including humans. Details of infection and identification of virus enzymes is critical for understanding transmission and developing antiviral agents that can slow or block virus multiplication. And being animal cells, organization is more complex than in Bacteria (endocytosis, membrane bound vesicles, nucleus, etc!) 26 13  10/22/24 Animal Viruses – Attachment (Adsorption) Virus attachment proteins (spikes) bind to receptors on host cell surface, usually glycoproteins on cytoplasmic membrane Often more than one receptor attachment is required (e.g., HIV binds to two) Normal function of receptor molecule unrelated to viral infection Particular viruses must attach to specific receptor; limits cell types and tissues a virus can infect Most viruses infect a single species (dogs do not contract measles from humans), but that is not always true (rabies virus infects dogs and humans) 27 Enveloped Viruses – Option 1 – Membr. Fusion… We have seen before – eukaryotic cells, tracking inside and outside! Virus is inside an exterior membrane vesicle But when virus membrane with cytoplasmic (or plasma) membrane, find it was already “inside”! 28 14  10/22/24 Option 2 – enveloped or non-enveloped Virus particle (enveloped or not) is phagocytosed – surrounded by a new membrane. On track to fuse with lysosome to be degraded. But as pH drops, virus proteins lyse membrane and release into cytoplasm!! (why does pH drop?) 29 Next steps Once the virion (or contents) is in cytoplasm, next steps depend on the nature of the nucleic acid in virus! Almost all DNA viruses push on into the nucleus (through the nuclear pore complex!) and begin replication. Most RNA viruses uncoat (degrade the capsid) and release contents into the cytoplasm 30 15  10/22/24 A moment of explanation on (+) and (-) -- Single Strand RNA (ssRNA) designated (+) or (-) ssRNA+ is equivalent to mRNA – can be read by Ribosomes to make protein. ssRNA- is the complement to ssRNA+. CANNOT MAKE PROTEIN DIRECTLY. First must make complement! ssDNA viruses also labeled (+) or (-) Neither strand makes protein directly. ssDNA(+) “reads” the same as ssRNA(+) HOWEVER, ssDNA(-) is what is TRANSCRIBED to MAKE ssRNA(+) 31 Animal Virus Synthesis Synthesis requires two events Expression of viral genes to produce viral structural and catalytic genes (e.g., capsid proteins, enzymes required for replication) Often synthesized as polyprotein that is cleaved by viral proteases à WHY???? Is a target site of some antiviral medications Synthesis of multiple copies of viral genome. Three general replication strategies depending on type of genome of virus DNA viruses (single or double stranded) RNA viruses (single stranded (+), single stranded (-), or double stranded) Reverse transcribing viruses 32 16  10/22/24 dsDNA DNA (+/ −) is replicated to form viral genome (−) strand transcribed to produce mRNA; translated to make viral proteins 33 ssDNA, (+) and (-) Complement to DNA synthesized first; then acts as template to produce more copies of viral genome (−) strand transcribed to produce mRNA 34 17  10/22/24 Replication of RNA Viruses Cells normally do not have ability to replicate RNA!! So no need to go to nucleus… RNA Viruses encode a NEW (to the cell) enzyme: RNA-dependent RNA polymerase (aka, Replicase) Evolved from DNA Dependent RNA polymerase (so no error checking) Allows virus to replicate RNA genome. Remember: ssRNA(+) also works as mRNA ssRNA(-) is complementary. Can also create double stranded RNA! 35 Replication of ssRNA(+) Viruses Cell Ribosome reads genome directly, making viral replicase Viral replicase makes multiple copies of (-) strand which are used to make many more (+) strands (more mRNA, plus copies of genome for assembly. 36 18  10/22/24 Replication of ssRNA(-) Viruses Virus must provide BOTH genome AND REPLICASE!!!! When released from capsid, viral replicase transcribes a complementary (+) RNA strand. Cell machinery makes viral proteins from (+) strand. Replicase also transcribes (+) strand to (-) for viral genome! 37 Replication of dsRNA Viruses Also requires a Viral Replicase enzyme in the capsid because cell cannot translate dsRNA! Replicase produces (+) strand copies to serve as mRNA for viral proteins. Late in infection, makes dsRNA for genome packaging! 38 19  10/22/24 RNA Viruses Encourage Mutations Replicases generally evolve from RNA polymerase – no Proofreading! But in Virus replication – errors are incorporated into viral genomes! This means that significant percent of virus particles cannot actually cause a productive infection. But this also makes for rapid mutation of RNA viruses. Can result in frequent changes in surface antigens, meaning that immunity to previous infections is not as effective to the new mutation. 39 Some RNA viruses even funkier…. Example: Influenza Has 8 linear segments of RNA(-), totaling ca. 13,500 bases Single virus infection, cell has usual “Antigenic Drift” However, if cell is infected by two or more viruses, infected cell packages a copy of each of the 8 segments into each new virion à but mix and match segments from among the different parent viruses. Can result in dramatic Antigenic “Shifts” that combine very different antigens that are completely different from anything that anyone has immunity to! Can result in a Pandemic! 40 20  10/22/24 Retroviruses…Special ssRNA(+) Viruses Genome functions as mRNA, translating a new enzyme – RNA dependent DNA polymerase – “Reverse Transcriptase” (RT) RT makes a DNA complementary copy of the ssRNA(+), then makes a complementary DNA strand of the new DNA. This dsDNA is then integrated into the host cell chromosome! Irreversible. Can hide or direct formation of viruses from there. 41 Assembly of Virons Capsomeres matrix proteins are self assembling. Some viruses use their genome as a scaffold, attaching capsomeres that assemble around it. Others assemble a “procapsid” into which all components are added. 42 21  10/22/24 Release of virions – Non-enveloped Viruses As for Lytic Bacteriophage, mature virions are assembled in the cell Viruses then trigger cell destruction (frequently by initiating apoptosis), releasing virions to new infections 43 Release of virions – Enveloped Viruses Similar to budding bacteriophage (e.g., M13), Enveloped viruses insert proteins into specific areas of cell membranes (can be cytoplasmic or other membranes) These proteins cluster, attach matrix proteins, bind mature virons, and extrude through membrane! 44 22  10/22/24 Kinds of Viral infections in Humans Acute infections Rapid onset; short duration Burst of virions released from infected host cell Immune system gradually eliminates virus Influenza, mumps, and poliomyelitis Persistent Infections Continue for years (or lifetime) May or may not have symptoms – or may have intermittent symptoms. 45 Persistent Infections (expansion) Chronic infections Continuous production of low levels of virus particles ”carriers” may lack symptoms but still transmit virus Latent infections – Virus genome remains, usually latent, in host cell Can reactivate to cause productive infection (e.g., when immunity wanes) Virus genome MAY be integrated into host cell chromosome as a provirus. 46 23  10/22/24 47 Latent Infections Provirus integrates into host chromosome or replicates separately, much like a plasmid Cannot be eliminated; can later be reactivated Examples are varicella-zoster virus (VSV) and herpes simplex type 1 virus (HSV-1) 48 24  10/22/24 Viroids Small single-stranded RNA molecules that form a closed ring Mainly in plants; cause serious disease Examples include potato spindle tuber, chrysanthemum stunt, citrus exocortis, cucumber pale fruit, and cadang-cadang Enter through wound sites Lots that we don’t know -- Where did they come from How do they replicate? How do they cause disease? 49 Prions Prions are proteinaceous infectious agents Composed solely of protein; no nucleic acids Linked to slow, fatal diseases in humans and animals Human diseases: Creutzfeldt-Jakob disease, fatal familial insomnia, and kuru Animal diseases: scrapie (sheep and goats), mad cow disease or bovine spongiform encephalopathy (cattle), and chronic wasting disease (deer and elk) 50 25  10/22/24 In disease, Prion proteins accumulate in neural tissue Neurons die Tissues develop holes Brain function deteriorates Characteristic appearance gives rise to general term for all prion diseases transmissible spongiform encephalopathies 51 Mode of action – Folding Rearrangement Cells produce normal form of neuronal protein PrPc Infectious prion proteins or PrPs Infectious form appears to cause normal protein to refold into new infectious forms Resistant to proteases, insoluble forms aggregate Resistant also to heat and chemical treatments Cause damage (structural or inflammatory?) to surrounding tissue 52 26