CLASS: 9:00-10:00 Scribe: Eric Larson La
DATE: 11-1-10 Proof: Lauren Morris
PROFESSOR: Dr. Lefkowitz Negative Sense RNA Viruses Page 7 of 7
I. Title [S1]
II. Contact Info [S2]
III. Objectives [S3]
a. The objectives of the lecture is to first go through some common and distinguishing characteristics of RNA viruses in general
b. Discuss basic replication mechanisms of RNA viruses – how they produce some of their proteins and from there cause disease
c. And finally discuss each family of viruses that make up this group, and talk a little bit about their pathogenesis and disease in humans
IV. Reading [S4]
V. References [S5]
VI. Virus Classification [S6]
a. Ok so: virus classification
VII. The Virosphere[S7]
a. This is a depiction of the world of viruses
b. The virus we’re interested in are the negative sense single strand viruses
VIII. The Virosphere [S8]
a. Here we see the group of single stranded RNA viruses
b. It’s worth pointing out, that in this group of viruses there are organisms that infect plants, invertebrates, and vertebrates
c. Most of the organisms that infect vertebrates can also infect humans, and these are the viruses we’re interested in
IX. Features Used to Classify RNA Viruses [S9]
a. Here are the features we’re interested in for RNA viruses
b. We’re interested in viron symmetry: the virus can be icosahedral, and helical
c. The viruses may or may not have a lipid envelope derived from the host cell, and if they don’t, you’ll see them as naked icosahedral capsids
d. The size of the particles are generally characteristic
e. The genome structure is especially characteristic for viruses
f. While the previous characteristics don’t necessarily indicate specific origins, genome structure is telling in discerning the origins and properties of viruses
X. RNA Virus Genome Structure[S10]
a. Most RNA viruses are single stranded, some are double stranded
b. Strand polarity can be positive or negative
c. The segments are often single, but others have one, two, or up to nine segments
XI. Single Strand Virus Genome Polarity [S11]
a. So when we talk about polarity, that is the characteristic which distinguishes the RNA viruses you studied last week from those we are covering now
b. Plus sense RNA can be directly can be directly translated into a protein
c. Negative sense mRNA must be first transcribed into positive sense before it can be translated into a protein
d. All the viruses we discuss today are negative sense RNA viruses, though ambisense RNA genomes are a little different
XII. Negative/Ambisense RNA Viruses [S12]
a. Let’s now talk about particular groups of this class of viruses
XIII. Properties of Negative-sense RNA viruses [S13]
a. So let’s talk about properties
b. They all have an eveloped virion derived from the host cell
c. They all have helical nucleocapsids inside that envelope
d. Some are negative sense single segment RNA
e. Some are negative sense and ambisense multi segment RNA viruses
f. They generally all replicate in the cytoplasm, but there are some exceptions such as Bornaviruses, and orthomyxoviruses which both have a component of the replication cycle that directly requires the nucleus
g. The genomes themselves are not infections – that is they must be first transcribed into positive sense RNA before they can begin to produce new viruses
h. That’s different from the positive sense RNA viruses where you can introduce the RNA genome directly into the cell and make viruses
XIV. Classification [S14]
a. So how do we classify some of these viruses?
b. Negative sense single strand viruses are part of the Mononegavirales order
c. That order is made up of several different families: bornaviruses, filoviruses, paramyxoviruses and rhabdoviruses
d. All of these viruses here, if you compare their proteins, you can see that they all have a common ancestor
e. However, they happen to cause different types of diseases, despite their relation
XV. Multi-Segment Negative and Ambisense ssRNA viruses [S15]
a. Ambisense viruses are a combination of both positive sense and negative sense RNA
b. They copy information off of both positive sense and negative sense segments
c. Arenaviruses and Bunyaviruses are two examples of ambisense viruses
d. They not natural pathogens of humans - you get them from bites of ticks or mosquitos
XVI. Multi-Segment Negative Sense ssRNA Viruses[S16]
a. Finally, we’ll talk about orthomyxoviruses
b. They are all negative sense viruses, but these all have multiple segments that encode their proteins
XVII. Viral Replication Cycle [S17]
a. What are the features of a replication cycle that allows these viruses to produce progeny?
b. I’ll give a general overview of the while replication cycle
XVIII. Viral Replication Cycle [S18]
a. The virus must first attach, enter the cell, and uncoat their nucleic acids and any associated proteins into the cell
b. At that point, if the virus is a positive sense RNA virus, translation can begin immediately
c. If the virus is a negative sense RNA virus you must first transcribe positive sense mRNA before translation can begin
d. After translation, you now have enough proteins that you can start assembling into new virions
e. Assembly is inside the cell cytoplasm itself or at the membrane of the cell
f. In essence you have association with virion proteins with virion DNA
g. They then leave by budding off and picking up part of the cell membrane on the way out
h. Lot’s of detail, but this general scheme is about all you’ll need to know
XIX. Diagram of the Viral Replication Cycle [S19]
XX. ViralProteins [S20]
a. There are a number of different viral proteins
b. The different viruses mix and match combinations of these proteins depending on their functions
c. Important thing is: they all need proteins for attachment, proteins for the structure of the virus, and proteins that mediate replication
d. For attachment, you might have the membrane glycoprotein, you might have a fusion protein and a hemagglutinin, and you also may have a neuraminidase
e. All these viruses have a matrix protein which underlies the lipid bilayer and gives structure to the virion itself
f. Finally, there are a number of proteins required for replication, which we’ll discuss in more detail soon
XXI. Rhabdovirus Virion [S21]
a. Rabies virus is a characteristic Rhabdovirus
b. For this virus, we can see how those proteins form up to give its characteristic structure
c. You have a lipid bilayer, you have G glycoprotein spikes coming out of it responsible for attachment
d. Underneath you have matrix responsible for the virion structure
e. Within that you have viral genome that has attached to it some of the replication proteins which I’ll talk about in a second or two
XXII. Virus Coding Strategies [S22]
a. The simplest mechanism for coding is to have individual open reading frames (ORFs)
b. But these viruses have a lot of tricks to maximize the information that can be derived from their single, small genome
c. So in addition to individual ORFs they might translate one large protein, which is then cleaved into two or more individual proteins – that is polyprotein processing
d. Also, when they transcribe some of their DNA, they play with the sequence of the RNA itself by adding or deleting bases to shift the reading frame one direction or the other which will produce a different protein
e. They also have multiple ribosomal initiation sites which means that by having the ribosome bind to different sites, they can produce different proteins
f. Some have a stop codon read-through, which means that in some cases the codon causes a stop of the translation, while in other cases it might be skipped over to give a different protein product
XXIII. Genome Organization Mononegavirales [S23]
a. This gives you an idea of what the genome organization looks like
b. This is for the order Mononegavirales
c. You can see how all these related viruses have similar proteins in their genomes
i. They all have the same proteins such as M and P proteins for replication, a matrix protein, a glycoprotein, and they have a polymerase protein
You can also see here that all these proteins are in the same general order in the genome
ii. This is one piece of evidence that leads us to believe they have a common evolutionary origi
XXIV. Virus Replication Machinery [S24]
a. Here’s the replication machinery
b. You have RNA dependent RNA polymerase (RdRp) that reads the RNA and can either produce mRNAs for proteins or virion RNAs
c. There’s a nucelocapsid protein which is what makes the nucelocapsid
d. That polymerase protein is dependent on the P protein for proper translation – it acts as a cofactor
e. The P protein binds to both the polymerase (L) and the nucleocapsidprotein (N)
XXV. RNA-dependent RNA Polymerase [S25]
a. The RdRp is the catalytic subunit of the polymerase complex that brings in nucleotides and allows for elongation of the transcribed or replicated RNA genome
b. Transcription and genome replication are functions which are also associated with RNA polymerase, and it can also have functions like capping, methylation and polyadenylation
XXVI. Source of the RNA-dependent RNA polymerase [S26]
a. Some viruses might need proteins contained in the host cell in their replication process
b. One protein no host cell has the code for is the RNA-dependent RNA polymerase
i. Host cells don’t code for this, so the virus is dependent on its own RNA dependent RNA polymerase to surve
c. So the virus must provide that polymerase in one of two ways:
i. For positive strand RNA viruses, the virus only has to carry in the RNA genome which can be translated by the host ribosome
ii. Since the negative strand can’t be directly translated, the virus must bring in the polymerase with them so they can initiate mRNA transcription once it reaches the host cell
XXVII. VSV Transcription and Replication [S27]
a. Overview of VSV transcription and replication cycle of a prototypic virus
b. So at top you have host genome that codes for N P M G and L proteins
c. Transcription is initiated to produce mRNA
d. This begins with transcribing the code for N, and once it finishes transcribing there the polymerase terminates and reinitiates at the start of the P code
e. So you don’t have the drop off of the polymerase, but just the termination and reinitiation of the same polymerase at the junction of each subsequent reading frame
f. This will lead to the production of N proteins which encapsulate the virion RNA
g. Once you get enough N proteins produced you can begin replication
i. So initially we have transcription of mRNA – the switch to replication occurs when enough of the N protein has been produced
h. In replication, you don’t have the termination-reintitiation process of transcription but rather you just make the whole transcript continuously
i. So this is the process for VSV, but it works similarly for all the viruses we discuss today
XXVIII. Important Human Pathogens from the Group of Negative Strand RNA Viruses [S28]
a. Now we’ll talk about each individual group of viruses and their pathogenesis
XXIX. Major Target Tissues [S29]
a. In general, these are the major targets of viruses
b. Just want to point out some of the negative-sense RNA viruses
c. You have different types of viruses that infect the various target tissues
d. Moth of the viruses we’ll look at here are paromyxo and orthomyxo viruses
i. These are human pathogens in that they spread human to human without animal reservoirs
XXX. Oral and Respiratory Viral Diseases [S30]
a. These are the diseases some of these viruses cause
b. Notice a lot of respiratory diseases caused by paramyxo and orthomyxo viruses
XXXI. Arboviruses and Zoonoses [S31]
a. Arboviruses are viruses that have an insect vector that transmit them from their normal animal host to humans
b. Zoonoses are viruses that humans are infected of as result of a contact of an animal – you pick it up directly from animal rather than through the insect vector
XXXII. Arenaviruses/Bunyaviruses [S32]
a. Let’s talk about some individual viruses now
XXXIII. Arenavirus and Bunyavirus Disease [S33]
a. These cause disease by accidental contact with animals, or some are transmitted by contact with insect vectors
b. The diseases are often seen in Africa – rare in U.S. and Europe, but people traveling can be infected by these
c. Among the arenaviruses is the junin virus which causes hemorrhagic fever
d. Again, with the bunyaviruses the main disease is hemorrhagic fever
i. One type is the Hantaviruse (comes from contact with rodent droppings) which does occur in the U.S.
e. You can get serious hemorrhagic fevers from all these viruses
XXXIV. Arenavirus Genome Organiation [S34]
a. So for ambisense viruses, some parts of virion mRNA must be transcribed before it can be translated, and even though other parts of the RNA appear as if they can be directly translated (like + sense), they must be first transcribed before they can be translated as well
b. Arenaviruses have two segments: large and small segments
c. On their genome they have various proteins which we’ve seen before
d. They have polymerase protein (L), and N protein which encapsulated the virion RNA
e. Glycoprotein used to attach to the cell
XXXV. Bunyavirus Genome Organization [S35]
a. Bunayavirus again has multiple segments – a small segment, a medium segment, and a large segment
b. Various segments acquired by cleaving of polyprotein
c. All of these segments are negative sense, although some of them look a little similar to the ambisense arenavirus RNA
XXXVI. Filoviruses [S36]
a. So there are very few of Bunyaviruses and Arenoviruses that are purely human viruses (few that don’t have an animal host or insect vector)
b. Filoviruses is another group of viruses that aren’t normal human pathogens
c. They have a characteristic, long, extended virion structure