Question

what is the sctrucute of the coronavirus?

what is the role of the cytoskeleton in the three main parts of the SARS-CoV-2 virus (covid19) life cycle: viral entry (internalization), transport via early endosomes to the ER-Golgi for replication, synthesis and packaging, and lastly, transport via late endosomes back to the plasma membrane for exocytosis of new viruses. For each of these phases, explain which cytoskeletal components are most likely involved, that is, choose microtubules or microfilaments as the major polymer (or “railroad tracks”) the virus uses, and which motor protein (dynein, kinesin and myosin) would be involved. You need to describe specific experiments you will use to confirm your hypothesis. In the experiments you design, be sure to indicate which drug(s) you will use, how you will fluorescently label the molecule you want to follow, and the kind of microscopy (confocal epifluorescence or electron microscopy) you will use. You should also describe how you will measure replication of the virus, for example “Real Time PCR.”

Answer 1

Structure of corona virus Covid-19

Coronaviruses are large, mostly spherical, sometimes pleomorphic (changeable in shape), particles with bulbous surface projections. The average diameter of the virus particles is around 125 nm (.125 μm). The diameter of the envelope is 85 nm and the spikes are 20 nm long. The envelope of the virus in electron micrographs appears as a distinct pair of electron-dense shells (shells that are relatively opaque to the electron beam used to scan the virus particle). The viral envelope consists of a lipid bilayer, in which the membrane (M), envelope (E) and spike (S) structural proteins are anchored. The ratio of E:S:M in the lipid bilayer is approximately 1:20:300. On average a coronavirus particle has 74 surface spikes. A subset of coronaviruses (specifically the members of betacoronavirus subgroup A) also have a shorter spike-like surface protein called hemagglutinin esterase (HE).

Cross-sectional model of a coronavirus

The coronavirus surface spikes are homotrimers of the S protein, which is composed of an S1 and S2 subunit. The homotrimeric S protein is a class I fusion protein which mediates the receptor binding and membrane fusion between the virus and host cell. The S1 subunit forms the head of the spike and has the receptor binding domain (RBD). The S2 subunit forms the stem which anchors the spike in the viral envelope and on protease activation enables fusion. The E and M protein are important in forming the viral envelope and maintaining its structural shape. Inside the envelope, there is the nucleocapsid, which is formed from multiple copies of the nucleocapsid (N) protein, which are bound to the positive-sense single-stranded RNA genome in a continuous beads-on-a-string type conformation. The lipid bilayer envelope, membrane proteins, and nucleocapsid protect the virus when it is outside the host cell.

The role of the cytoskeleton in the three main parts of the SARS-CoV-2 virus (covid19) life cycle: viral entry (internalization), transport via early endosomes to the ER-Golgi for replication, synthesis and packaging, and lastly, transport via late endosomes back to the plasma membrane for exocytosis of new viruses.....The Coronavirus superfamily (Coronaviridae) includes several human pathogens with large RNA-encoded genomes, e.g., influenza, common cold, and viral encephalitis, which are classified into alpha-, beta- and gamma-coronavirus families with further division into Lineages A, B, and C. COVID-19 is classed as a Lineage B beta-coronavirus with high similarity to SARS-CoV, thus it'-s recentl renaming to SARS-CoV-2. Although the first members of the beta-coronavirus family were recorded in the 1960’s, the family’s rate of new virulent human pathogens has increased rapidly over the past 20 years, now numbering six in total with the latest ones having the familiar pseudonyms SARS , HKU , MERS , and now COVID-19 / SARS-CoV-2. Their emergence is linked to increased density of human and animal populations which has enhanced zoonotic transmission rates.Here, we describe the four stages of virus “life” cycle with respect to its interaction with the cytoskeleton.Coronaviruses are enveloped positive-stranded RNA viruses that replicate in the cytoplasm. To deliver their nucleocapsid into the host cell, they rely on the fusion of their envelope with the host cell membrane. The spike glycoprotein (S) mediates virus entry and is a primary determinant of cell tropism and pathogenesis. It is classified as a class I fusion protein, and is responsible for binding to the receptor on the host cell as well as mediating the fusion of host and viral membranes—A process driven by major conformational changes of the S protein. This review discusses coronavirus entry mechanisms focusing on the different triggers used by coronaviruses to initiate the conformational change of the S protein: receptor binding, low pH exposure and proteolytic activation. We also highlight commonalities between coronavirus S proteins and other class I viral fusion proteins, as well as distinctive features that confer distinct tropism, pathogenicity and host interspecies transmission characteristics to coronaviruses.

Viral entry relies on a fine interplay between the virion and the host cell. Infection is initiated by interaction of the viral particle with specific proteins on the cell surface. After initial binding of the receptor, enveloped viruses need to fuse their envelope with the host cell membrane to deliver their nucleocapsid to the target cell. The spike protein plays a dual role in entry by mediating receptor binding and membrane fusion. The fusion process involves large conformational changes of the spike protein. Coronaviruses use a variety of receptors and triggers to activate fusion, however fundamental aspects that enable this initial step of the viral life cycle are conserved. In this review, we will address entry strategies of coronaviruses and how these mechanisms are related to host tropism and pathogenicity.

The anterograde pathway, from the endoplasmic reticulum through the trans-Golgi network to the cell surface, is utilized by trans-membrane and secretory proteins. The retrograde pathway, which directs traffic in the opposite direction, is used following endocytosis of exogenous molecules and recycling of membrane proteins. Microbes exploit both routes: viruses typically use the anterograde pathway for envelope formation prior to exiting the cell, whereas ricin and Shiga-like toxins and some nonenveloped viruses use the retrograde pathway for cell entry. Mining a human genome-wide RNA interference (RNAi) screen revealed a need for multiple retrograde pathway components for cell-to-cell spread of vaccinia virus. We confirmed and extended these results while discovering that retrograde trafficking was required for virus egress rather than entry. Retro-2, a specific retrograde trafficking inhibitor of protein toxins, potently prevented spread of vaccinia virus as well as monkeypox virus, a human pathogen. Electron and confocal microscopy studies revealed that Retro-2 prevented wrapping of virions with an additional double-membrane envelope that enables microtubular transport, exocytosis, and actin polymerization. The viral B5 and F13 protein components of this membrane, which are required for wrapping, normally colocalize in the trans-Golgi network. However, only B5 traffics through the secretory pathway, suggesting that F13 uses another route to the trans-Golgi network. The retrograde route was demonstrated by finding that F13 was largely confined to early endosomes and failed to colocalize with B5 in the presence of Retro-2. Thus, vaccinia virus makes novel use of the retrograde transport system for formation of the viral wrapping membrane.

A real-time reverse transcription–polymerase chain reaction (RT-PCR) assay was developed to rapidly detect the severe acute respiratory syndrome–associated coronavirus (SARS-CoV). The assay, based on multiple primer and probe sets located in different regions of the SARS-CoV genome, could discriminate SARS-CoV from other human and animal coronaviruses with a potential detection limit of <10 genomic copies per reaction. The real-time RT-PCR assay was more sensitive than a conventional RT-PCR assay or culture isolation and proved suitable to detect SARS-CoV in clinical specimens. Application of this assay will aid in diagnosing SARS-CoV infection.