Eukaryotes organisms that contain a membrane bound nucleus and organelles. Prokaryotes organisms that lack a nucleus or other membrane-bound organelles. Viruses small, non-cellular (lacking a cell), infectious agents that can replicate only within the cells of living organisms. LESSON 1.4 WORKBOOK Viral sizes and structures In this lesson we will learn about viruses what their relative sizes are compared to bacterial and host cells; what structures they use to infect host cells and what their lifecycle is. As with bacterial structures, viral structures are precisely adapted for propagation in the host. You may even be able to predict the function of the viral structures based on what you have learned from bacteri Just how small are viruses? As we mentioned before, there are many different classes of pathogens. The larger pathogens, such as fungi and multicellular parasites, are eukaryotic, which means they have a membrane-bound nucleus. Bacteria are prokaryotic, which, as we saw before, means they lack a membrane-bound nucleus. Viruses are neither prokaryotic nor eukaryotic in fact they are not even alive! They are simply a collection of genetic material and proteins that are able to replicate by exploiting the cell they infect. Viruses are much, much smaller than bacteria and this impacts our ability to detect them they are so small we can't even see them under a simple microscope! How big are viruses compared with other pathogens? The microbial world generally exists below the limit of what the unaided human eye can see. Sizes can vary considerably within each category. Eukaryotes are the biggest, followed by prokaryotes, and then by viruses. However, within each group, sizes can vary in volume by a thousand fol In fact, the largest viruses fall within the lower size range of prokaryotes similar to the size of small bacteria such as Chlamydiae that cause chlamydia infections. For example, the largest virus found till date, Pithovirus sibericum, was revived from 30,000 year-old Siberian permafrost and can easily be seen under a light microscope. 41
Micron (micrometer, µm) one millionth of a meter. Because the differences in microbe sizes are so large, we need more than one unit of measurement Most bacteria are on the order of 1 micron in diameter. A micron, or a micrometer, is one millionth of a meter. To give you an idea of how small this is, bacteria are so small that when they are in suspension in a fluid like urine, it only appears cloudy when there are between one million to ten million bacteria per milliliter. This means that even when water looks completely clear, it can contain millions of bacteri Viruses are even smaller, on average being about 8,000 times smaller in volume than the average bacterium. To give you a sense of scale, if a virus were the size of an orange, a bacterium would be about the size of a sof Practice Calculation: Conversion Conversion between units is important for every aspect of science. Fortunately, in this course we are only concerned with a few conversions that relate to microbe size. So, let s practice: How big is a micron? 1 millimeter = 1 X 10-3 meters or 0.001 meters 1 micron = 1 X 10-6 meters or 0.000001 meters 1 nanometer = 1 X 10-9 meters or 0.000000001 meters 1. Which shows the correct increasing order of the relative average sizes of microbes? virus < proteins < prokaryotes < eukaryotes eukaryotes < bacteria < virus < proteins proteins < virus < bacteria < eukaryotes prokaryotes < eukaryotes < virus < proteins 2. How big is 2 microns? 2 x 10-3 m 2 x 10-4 m 2 x 10-5 m 2 x 10-6 m 42
Genome the genetic material of an organism. Envelope membrane layer surrounding the capsid that is derived from the host cell membrane. Capsid a protein shell of a virus surrounding and protecting its genome. Viral structure: It's all about delivering the genome to the host cell Now back to viral structures. When you are reading about viral structures, it is important to keep in mind that viruses require their host cells to supply everything they need to make more copies of themselves, with the exception of a few key viral proteins and the viral genome. For this reason, viruses can be thought of simply as vessels that carry genetic information. Because the virus uses so many of the host cell proteins to replicate, it does not need to code for these genes. Hence, viral genomes are much, much, smaller than either bacterial or eukaryotic genomes. Viral genomes can be either DNA or RNA, as we will discuss later. Viral genomes are enclosed within two structures: 1. The protein coat (capsid) all viruses have it 2. The envelope some viruses have it Figure 1: There are two types of viruses: Naked and Envelope Both have a capsid, but only the enveloped viruses have an envelope. All viruses also have a genome (RNA or DNA), and a few viral proteins that they need for replication these are proteins that the host cells lack. 3. Viral structures are composed of: viral genome, capsid, envelope capsule, murein, flagella RNA, envelope, capsule none of the above 43
The capsid protects the genome All viruses have a capsid that surrounds their genome. The capsid is a shell that can be single or double layered, and is arranged in one of three symmetrical patterns: icosahedral, helical or complex. Icosahedral capsids have the shape of a regular polygon with 20 identical equilateral triangular faces. Helical capsids are arranged as a helix or spiral. Complex capsids are neither purely helical nor icosahedral in shape, and may have additional structures. The capsid is composed of viral proteins. Along with the genome, sometimes viral proteins required for replication in the host cell are also enclosed within the capsi For non-enveloped or naked viruses, the capsid is the only protection the genome has from the environment! The envelope is made of host cell lipids/proteins and viral proteins For enveloped viruses, the envelope is vital to their life cycle. It surrounds the capsid and is made of viral proteins that help viruses enter host cells, plus lipids and proteins stolen from the cell membrane of the host cell. The inner surface of the envelope has virus-specific proteins that form attachments to the capsid to anchor it. These proteins stabilize the interactions between the envelope and the capsi As we will see later, they also determine the location of viral assembly in the host cell. The four stages of the viral life cycle The rest of this reading describes viral structures in the context of how they function. To simplify we can divide the viral life cycle into four steps: 1. Attachment 2. Entry 3. Replication 4. Exit Figure 2: Helical capsid (upper left); Complex capsid (upper right); Icosahedral capsid (bottom). We will focus on viral replication in later units. While you are reading keep in mind that, like bacterial structures, each viral structure performs its own unique key function. 4. All viruses have a/an but only some have a/an. envelope, DNA capsid, envelope envelope, capsid DNA, capsid 44
Uncoating the process of release of the viral genome from the capsi Virion a complete virus particle. Receptor a molecule, usually protein, on a cell or virus surface that binds to a corresponding specific molecule that it encounters. Tropism the property of a specific virus to attach to only a specific type of cells. The virus has a major goal to make more virus particles. To reproduce, a virus needs to enter its specific host cell where it will make new viral particles and finally exit the cell. Figure 3: An overview of the virus lifecycle in a host cell. First, the virus attaches to the cell it is going to infect. Second, the virus enters the cell and uncoats its genome so that replication can begin. Third, viruses replicate their genomes and make the structural proteins they need to assemble a new virus particle. Fourth, once the virus has made all its parts it assembles new virions that exit the host cell to infect other cells. 1) Attachment usually requires a specific receptor Both naked and enveloped viruses need to attach to a host cell before they can enter it. To do this all viruses have receptors on their surface that act like keys, which interact with select receptors on the host cell surface that act like locks. For example, HIV is an enveloped virus that uses its receptor protein gp120 to attach to a receptor on the host cell surface called CD4 (see Fig. 4 below). HIV can therefore only infect cells that have CD4 on them, which happens to be the helper T cells and dendritic cells of the immune system (more about immune cells in Lesson 1.5 and Unit 5). This lock-and-key interaction, determines which host cells can be targets for a given virus a phenomenon called tropism. 5. Which shows the accurate viral life cycle? entry > attachment > exit > replication attachment > entry > replication > exit entry > attachment > replication > exit replication > entry > attachment > exit 45
H1N1 a strain (type) of the flu virus. Hemagglutinin a protein on the surface of flu viruses which is used as a receptor to bind to host cells. Endocytosis the process which cells use to absorb molecules from the outside by engulfing and bringing them inside the cytoplasm. Extracellular outside of the cell. Endosome a membrane bound vesicle in eukaryotic cells used to transport molecules within the cytoplasm. A"achment* Genome& Viral&protein& Capsid& Figure 4: HIV, like all viruses attach to their host cells using specific surface receptors. Consider the influenza virus. Influenza virus has a receptor on its envelope called hemagglutinin (the H in H1N1). Hemagglutinin binds to sialic acid sugars that are on the surface of epithelial cells in the respiratory tract, making the virus tropic for those cells. Different variants of influenza have different H receptors that attach to different sialic acid sugars. These different sialic acid sugars are located on cells that are in different locations in the respiratory tract. This means that different variants of H1N1 infect different sites in the respiratory tract, which can impact the severity of disease. 2) Entry depends on whether the virus is enveloped or nake In order to replicate, all viruses must cross the plasma membrane and get inside the host cell. As we will see, enveloped and naked viruses enter cells differently. Naked viruses cross the host membrane Naked viruses aren t coated in a lipid envelope, so they can t fuse with membranes. Instead, naked viruses either inject their genomes into the host cell cytoplasm through the plasma membrane. An alternative route naked viruses can take is via endocytosis, a process that host cells usually use to take up molecules from the extracellular space. A virus particle may hijack the endocytosis process to enter the host cell in a vesicle (sac), formed by the host membrane, called an endosome. Next the capsid and the genome are released into the cytoplasm where the genome can start to replicate. Figure 5: Both naked (shown here) and enveloped viruses can be taken up through endocytosis. 6. Attachment of both enveloped and naked viruses depends on: the tropism of the virus and host cell s receptors how fast uncoating of the genome occurs the presence and secretion of sialic acid all of the above 46
Lysis the process of rupturing or breaking down of a cell. Enveloped viruses fuse with the host membrane The most direct way for enveloped viruses to deliver their capsid to the host cell cytoplasm is to fuse their own envelope membrane with the membrane of the host. Since both viral envelopes and host cell membranes are composed of similar host cell membrane lipids, fusion happens easily, like two oil droplets combining in water. After the membranes have fused the virus can then release its capsid, which contains its genome, into the cytoplasm. HIV is an example of a virus that fuses its envelope with the T cell membrane. The genome is then released from the capsi Some enveloped viruses such as influenza are taken up by the host cell through endocytosis in a similar manner to some naked viruses as described above. The virus enters the cell via the endosome. Next, the virus fuses its envelope with the endosomal membrane, and the capsid and nucleic acid are released into the cytoplasm. 3) Viral replication will be discussed in detail in Unit 4. 4) Exit also depends on whether the virus is enveloped or nake Once the step of replication has occurred and the virus has made many copies of itself, these virions must leave the infected cell to infect other cells so that further cycles of propagation can take place. Again, viruses have two different exit strategies depending on whether they are enveloped or nake Naked viruses lyse the host cell Figure 6: Enveloped viruses fuse with the host membrane. Naked viruses employ a more straightforward exit strategy than enveloped viruses. Quite simply, the virus replicates rapidly, and the huge number of virus particles that accumulate in the cytoplasm burst the cell open. This process of rupturing a cell is termed lysis. It is an important phenomenon and we will use the term routinely to describe when a cell is broken open. An example of a naked virus that exits by lysis is human papillomavirus or HPV, the virus that causes genital warts and is linked to forms of cervical cancer. 47
Budding the process in which viruses leave the host cells by exiting its surface by wrapping their capsids in the host cell membrane. Enveloped viruses bud off of host cells Enveloped viruses leave the cell in distinct stages. First, the envelope proteins collect in the host cell membrane close to where the genome is making the capsid proteins. Then the genome and capsid proteins travel together to the site on the host cell membrane where the envelope proteins are located, and attach to them on the inside of the membrane. Next, the fully formed capsid containing the genome buds out of the host cell. As it does so, the capsid surrounds itself with the stolen host membrane, studded with viral envelope proteins, and forms a new complete virus particle or virion. This is true of both HIV and influenz Whether or not the budding off of virus particles will cause the host cell to die depends on the rate with which a virus is replicating; if a virus is replicating slowly the cell may survive, but rapidly replicating viruses will probably cause the host cell to die. Figure 7: Enveloped viruses exit the host cell by budding. Naked viruses lyse the cell. 7. The exit of naked and enveloped viruses differ because naked viruses result in cell lysis while enveloped viruses bud off enveloped viruses result in cell death while naked viruses bud off there is no difference in the exit of naked and enveloped viruses naked viruses exit through endocytosis while enveloped viruses use enzymes 48
How do viruses cause illness? As you have just seen, in replicating through a host, viruses can cause widespread cell death and damage. Sometimes infected cells are killed by the host s own immune system in order to stop the virus from spreading through the body. In either case, cell death can cause serious illness if the cells that are being killed are important to the body s functioning, such as those in the lungs or in the central nervous system, or if the host cannot replace the cells that are being destroyed quickly enough. Some viral infections seem to transform cells into a cancerous state that makes them grow out of control. One of the most common cancers, is of the liver, and is caused by persistent infection with Hepatitis B or C viruses. Another example is cervical cancer caused by human papillomavirus (HPV). 49
STUDENT RESPONSES Predict what happens to a virus once it exits its human host (e.g. through sneezing or out the gut into the fecal matter). How does it survive? Remember to identify your sources If a lab dish is 10cm across, how many average bacterial cells in a row would it take to form a side-by-side chain across the middle? How many average-sized viruses would it take to make a chain of the same length? How many average-sized eukaryotic cells would it take? 50
TERMS TERM DEFINITION Budding Capsid Complex Cytoplasm Endocytosis Endosome Envelope Eukaryotes Extracellular Genome H1N1 Helical Hemagglutinin Icosahedral shape Lysis The process in which viruses leave the host cells by budding off its surface by wrapping their capsids in the host cell membrane. Protein shell of a virus surrounding and protecting its genome. Viruses that have a capsid that is not icosahedral nor helical, and may have additional structures. The internal content of a cell, excluding the nucleus when present. The process which cells use to absorb molecules from the outside by engulfing and bringing them inside the cytoplasm. A membrane bound vesicle in eukaryotic cells used to transport molecules within the cytoplasm. These molecules can be engulfed from outside or produced inside the cell. Membrane layer surrounding the capsid that is derived from the host cell membrane. Organisms that contain a membrane bound nucleus and organelles. Outside of the cell. The genetic material of an organism. A strain (type) of the flu virus. Having the shape of a spiral or a helix. A protein on the surface of flu viruses which is used as a receptor to bind to host cells. Having the shape of an icosahedron that has 20 identical triangular faces. The process of rupturing or breaking down of a cell. Micron (micrometer, µm) One millionth of a meter. 51
TERMS TERM DEFINITION Prokaryotes Receptor Sialic acid Tropism Uncoating Virion Viruses Organisms that lack a nucleus or other membrane-bound organelles. A molecule in or on a cell that receives chemical signals. A modified sugar molecule present on the surface of many cells and used as a receptor by the flu virus. The property of a specific virus to attach to only a specific type of cells. The process of release of the viral genome. One viral particle that contains all the components of a mature virus. Small, non-cellular (lacking a cell), infectious agents that can replicate only within the cells of living organisms. 52