B1100 Cytology -39-52 PDF

39 CH 3. EUKARYOTES, PROKARYOTES AND VIRUSES I. DEFINITION OF EUKARYOTES AND PROKARYOTES Free living cells and cells of multicellular organism are divided into two major types which are eukaryotes and prokaryotes. The prefix "eu" means true and "karyon" means nucle...

39 CH 3. EUKARYOTES, PROKARYOTES AND VIRUSES I. DEFINITION OF EUKARYOTES AND PROKARYOTES Free living cells and cells of multicellular organism are divided into two major types which are eukaryotes and prokaryotes. The prefix "eu" means true and "karyon" means nucleus, thus eukaryotes are cells Q having "true" nuclei. A Lysosome nucleus is said to be "true" when it is delimited by two membranes. The prefix "pro" means before and refers to cells without nuclei. This nomenclature is related to the theory of t 0 evolution. Prokaryotes 0 " (J existed long before "(') ti Golgi complex eukaryotes according to the discovered fossils. Viruses do not belong to these groups because they are not living things as long as they can not reproduce in an autonomous manner. The main difference between eukaryotes and prokaryotes is the presence of membranes between DNA and ribosomes. In fact, eukaryotic cells have membranes that delimit many compartments Figure 3.1: Organelles and structures common to most animal cells. (organelles) within the cell, including the nucleus. By contrast, in prokaryotes cytoplasm and "nucleus" are not separated and all cellular structures are found in the same compartment. Eukaryotes and prokaryotes differ in many other respects. Procaryotes have no organelles. Eukaryotes may be unicellular or multicellular, photoautotrophs (perform photosynthesis) or heterotrophs. All animal and plant cells are eukaryotes. Protists (e.g. algae), fungi and protozoa are also eukaryotes. Prokaryotic cells include eubacteria, archaebacteria and pleuropneumonia-like organisms (PPLO) or mycoplasma. Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  40 II. BRIEF PRESENTATION OF EUKARYOTIC CELLS 1. Animal Cells There is no typical cell that can serve as an example of all animal cells because cells vary considerably in their size, shape, physiological functions, abundance of organelles and structures. The range of size of animal cells is between 10 and 30 µm while plant cells are usually larger reaching 100µm. Many cellular structures are common to the majority of animal cells (e.g. mitochondria, Golgi body, endoplasmic reticulum) but others are specific to certain cell types (e.g. cilia, flagella and microvilli). Most of the structures are depicted in a composite animal cell shown in Figure 3.1. The diverse organelles and constituents listed below will be treated with more details in the coming chapters. The main structures and organelles of eukaryotic celles are: the plasma membrane or plasmalemma that interacts with the extracellular matrix as well as with neighbouring cell membrane the nucleus (genetic information) the cytoskeleton is a set of filaments and microtubules (shape, adhesion and movement) the endoplasmic reticulum (ER, synthesis and maturation of diverse cell components) the cytosol (also named hyaloplasm) which is the Cell walls aqueous solution that fills the space between the Interior { plasma membrane and the of cell nuclear envelope. It is bathing the organelles of the cell. It is the site diverse metabolic reactions. 0.5 µm Plasmodesmata Plasma membranes - the ribosomes (translation) Figure 3.2: A typical plant cell seen by TEM (top) and - the mitochondria (oxidation longitudinal (bottom). section of the cell wall showing plasmodesma and cell respiration) - the Golgi body (a set of dictyosomes for maturation and sorting of diverse cell components) - the lysosomes (intracellular digestion) - the peroxisomes (oxidation) - the centrosome (movement) - cilia and flagella (locomotion in certain cell types). Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  41 2. Plant Cell Properties All of the organelles and structures which are regular constituents of animal cells (Figure 3.1) are also found in similar forms in many types of plant cells; the centrosome and lysosomes which are restricted to animal cells are exceptions. Several other organelles are exclusively found in plant cells and include the carbohydrate-rich cell wall, plasmodesmata and plastids such as chloroplasts (Figure 3.2). Plant cell wall is a thick structure, located outside the plasma membrane. It is made up of three classes of polysaccharides which are cellulose (polymer of 13-D-glucose), hemicellulose (glucose+pentoses) and pectin (uronic acids-rich polysaccharides). Furthermore, small amounts of glycoproteins are found in plant cell wall. Matrix of the cell wall is hemicellulose, pectin and proteins. The cell wall plays a supportive and protective role in plant tissues since it surrounds the plasma membrane. It protects the cell against fluctuation of osmotic pressure as well as against intruders (viruses, bacteria, fungi,...). It determines the cell shape, growth direction and mediates cell-cell adhesion through the middle lamella. In fact, the walls of neighboring cells are fused together by a pectin-rich layer called middle lamella. Plasmodesmata (Figure 3.2) are interruptions of the cell wall by cytoplasmic bridges between a cell and its neighbor. These channels serve for intercellular exchange of nutrients and diverse materials. Their structure and further details about the cell wall are given in Ch 4. Ill. 7.. Plastids form a family of organelles involved in many functions such as storing of organic compounds and photosynthesis. An example of plastid is chloroplast found in green plant tissues. Chloroplast is specialized in using light energy in order to reduce CO2 and H20 into glucose (the process of photosynthesis). Although the term vacuole was utilized in animal cells to indicate certain vesicles, real vacuoles are restricted to plant cells. In mature plant cells, vacuoles are particularly large compartments, they often occupy a major proportion of the cell volume. The large vacuole in a mature plant cell results from fusion of smaller vacuoles in the young dividing cells. The vacuoles are bounded by a single membrane named Anaerobic bacteria: Acidophiles (low pH, e.g. tonoplast. Vacuoles serve to sulfur bacteria1 ), Thermophiles (high temperature) Archaebacteria Halophiles bacteria (growing in extreme saline expand the plant cell volume conditions, Dead Sea) without diluting its cytoplasm; Anaerobic bacteria that reduce carbon dioxide they also function as sites for into methane (methanogens) Gram positive bacteria water storing as well as for Gram negative bacteria storage of some cell products Eubacteria Green photosynthetic bacteria (anaerobic) or metabolic intermediates. Cyanobacteria (blue-green algae) Further details will be explained in the coming Purple photosynthetic bacteria chapters. Table 3.1: Classification of bacteria. Ill. PROKARYOTES From a structural point of view, prokaryotes are characterized by being simple living things. However, they are diverse and complex from the biochemical standpoint. In other words, we encounter inside prokaryotes a huge diversity in terms of biochemical reactions. Some of these reactions are beneficial to humans (example of lactic bacteria that ferment milk) while others are harmful and produce toxins (example of tetanus toxins, diphteria toxin). Bacteria are widespread on earth but most of them are not pathogens. Bacteria differ in terms of oxygen requirement. Some can grow only if oxygen is present (aerobic), others are anaerobic (strict or facultative). Strict anaerobic cannot grow Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  42 in the presence of oxygen. Bacteria also differ in terms of trophic mode: heterotrophs, photoautotrophs (perform photosynthesis), chemoautotrophs (perform chemosynthesis). Prokaryotes differ from eukaryotes by many properties such the absence of nuclear envelope around DNA which is the most important difference, but also they lack all membrane-bound organelles. Although known to be free living cells (unicellular), certain prokaryotic species may grow into clusters or colonies in a semi-solid medium. However, such colonies cannot be considered as multicellular organisms since the individual cells do not differentiate; all of the cells in a colony remain structurally and functionally 1 t identical. Prokaryotes grow rapidly asexually by fissiparity (or..Figure 3.3:. :.!!!! a::::SIII!......... Colony of cocci (left, SEM), bacilli (center, SEM) and binary fission, Figure 3.6, not by spirochetes (left, TEM). mitosis) in rich culture media. The frequency of such rapid division may be as fast as one generation every 20 minutes. Bacteria grow freely whenever nutrients are available in the medium. However, some bacterial species are intracellular parasites (e.g. Chlamydia), that is they live and proliferate inside a eukaryotic cell. Prokaryotes are numerous and may be classified in different ways. They are divided into eubacteria (true bacteria) and archaebacteria (ancient bacteria, Table 3.1 ). These classes differ in many respects. For instance, most eubacteria CAPSul live in mild conditions that are quite similar to those required for eukaryotes. By contrast, archaebacteria live in extreme conditions of pH, temperature, and salinity. Thermophile and F---_ NUCLEAR AREA (nucleoid) containing halophile bacteria live at high ONA temperature and high salt concentration, respectively. Methanogen bacteria are anaerobic and produce methane in their metabolism. Archaebacteria are extremophile bacteria and are chemoautorophs. Figure 3.4: Scheme of a typical bacterium where most structures are assembled. See text for details. In addition to diversity of their physiology, prokaryotes are diverse in terms of size and shape (Figure 3.3). Nommenclature of bacteria uses prefixes related to their shape. Many prokaryotes are spherical (genus: coccus), or elongate (genus: Bacillus, e.g. Bacillus anthracis which is a gram-positive bacterium that causes anthrax). Another common shape is the spiral one (genus: spirochetes). Certain bacteria change their shape depending on the medium where they are found. Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  43 Cocci bacteria are about 1 µm in diameter. Escherichia coli are elongate (2µm in length). Some bacteria are about 5µm to 10µm in length (e.g. cyanobacteria, spirochetes). However, the average size of bacteria is about 1 to 2µm. In addition, there are very small bacteria, about 0.2µm in diameter, even smaller than some viruses. These are named mycoplasma and may be spherical or filamentous (Figure 3.7). They belong to the class of Mollicutes (e.g. Mycoplasma pneumonia). Mycoplasma or PPLO cause a number of diseases in human, animals (especially in the respiratory and urogenital tracts) as well as in plants. Mycoplasma behave as intracellular parasites although they can grow in cell-free media. They are highly polymorphic since they lack rigid envelope. Mycoplasma are bounded by a special membrane made up of lipids (including cholesterol) and proteins, but they have no cell wall and capsule. Some mycoplasma are classified as eubacteria, others (e.g. thermoplasma) are classified with archaebacteria. Prokaryotes are simpler in organization than eukaryotes since the cytoplasm is not compartmentalized. However prokaryotic envelope (cell wall) is more complex than eukaryotic Figure 3.5: TEM view of cyanobacterium. section showing envelope. Although the cell photosynthetic lamella and carboxysomes (dark circles). structures we describe in this paragraph are mainly those of Gram positive9, Gram negative and cyanobacteria, most of the cellular structures described next are common to all prokaryotes. a. Nuc/eoid The nucleoid is a dense region in the bacterial cytoplasm. It contains the bacterial genetic material combined with proteins and is not separated from the remaining cell components by any membrane (Figure 3.4 ). The genetic material is DNA in form of a single circular chromosome (size about several 1000 kb). This chromosome carries all necessary genes for bacterial life. The chromosomal DNA which is much longer the the bacterium, is condensed in the nucleoid region by specific proteins. The nucleoid segregates the two chromosome copies (contained within two nucleoids and resulting from DNA replication) during bacterial division by fissiparity. Nucleoli do not exist in bacteria. In certain bacteria and certain free- living eukaryotes, there is an extrachromosomal genetic material called plasmid. Plasmids are circular Figure 3.6: Bacterial division by binary fission. DNA molecules that are about several 9 Gram is a staining technique. Gram+ refer to the species that can be stained by the technique, Gram- are those that can not. Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  44 kb in size, they carry a few genes that confer certain advantages to the host cell (e.g. genes responsible for resistance to antibiotics). Plasmids can be transferred between bacteria, naturally by conjugation or experimentally. They are used as vectors indispensable for molecular biology and genetic engineering experiements. b. Cytoplasmic structures The cytoplasm is a single compartment that contains organic and inorganic compounds required for diverse cell physiology processes. There are no mitochondria, no chloroplasts, no ER no Golgi apparatus, no cytoskeleton, no transport by vesicles (no endocytosis, no exocytosis). All organelles of eukaryotes are absent in prokaryotes. Despite that, bacteria are able to perform many processes in a way similar to eukaryotes. For instance, although mitochondria are absent, the enzymes that catalyze oxidation-reduction reactions (e.g. cell respiration) and synthesize ATP are usually present in the bacterial plasma membrane and in the cytoplasm. L__ Although no organelles are found in prokaryotes, (B) 0.2 µm infoldings of the plasma membrane may occur especially Figure 3.7: Mycoplasma genitalium seen by SEM (whole, a) and TEM in photosynthetic bacteria 10 forming photosynthetic (section, b). lamellae. These lamellae may separate from the membrane and form independent structures named chromatophores (Figure 3.5). However, such lamellae are not comparable to ER of eukaryotic cells. Similarly, cyanobacteria have no organelles, but they are provided with membrane-bounded gas vacuoles that control buoyancy and carboxysomes which form a complex of enzymes involved in CO2 fixation for photosynthesis. Other bacteria, such as Chlorobium, have chlorophyll-containing vesicles. Within the cytoplasm of all bacteria there are ribosomes that translate mRNA into Pllus Flagellum Gram-positive bacteria Gram-negative bacteria Pllus teichoic surface lipoteichoic acid protein add outer membrane cell membrane phospholipid space Figure 3.8: Cell wall structure of Gram positive and Gram negative bacteria. proteins. Prokaryotic ribosomes are slightly different from eukaryotic ribosomes in terms of 10 Note that not all autotrophs rely on normal photosynthesis to produce energy (photoautotrophs). Chemoautotrophs use light but rely on chemiosynthesis (using energy stored in inorganic molecules such as H2S, nitrites and ammonia). Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  45 size and fine composition (see Ch 5. II. ). Ribosomes are free or attached to the plasma membrane, but no ribosomes are attached to cytoplasmic lamella when they occur. Moreover, in the cytoplasm, there are granules or inclusions that represent stored inorganic or organic compounds such as sulfur, phosphates, and carbohydrates. c. Bacterial envelope A bacterium is always enclosed in a plasma membrane which is surrounded by a cell wall in all species (except mycoplasma). So, the bacterial envelope consists of a plasma membrane which is enveloped by a cell wall. The plasma membrane is a lipid bilayer that has the same basic structure as that of eukaryotic cell membranes (but without steroids 11 ). The plasma membrane contains many protein types that are involved in diverse metabolic reactions (e.g. transporters, receptors, respiratory chain Hook (universal joint) proteins, ATP synthase, cell wall-synthesizing enzymes,...). The plasma membrane is characterized by a selective and controlled permeability that is indispensable for cell life. The plasma membrane gives rise to a specialized Peptidoglycan layer structure named cytoplasmic Periplasmic space lamella (see previous Inner (plasma) paragraph). For more than two membrane decades it was thought that the plasma membrane gives rise to another infolding named mesosome that is involved in the formation of septum between two daughter cells and in the DNA segrargation during cell division. The septum is the inward growth of Figure 3.9: Detailed scheme of the bacterial flagellum elements (top). A bacterium forming an endospore (bottom). cell envelope between the two daughter cells during division by binary fission (Figure 3.6). Recently, many scientists showed that mesosomes do not exist in reality and they are artifacts of sample preparation for microscopy analysis. Bacterial cell wall determines its shape and provides protection against changes of osmotic pressure (solute concentration). The cell wall is rich in peptidoglycans which consist of complex 3D network of heteropolysaccharides (rich in glucose and N­ acetylglucosamine) that are joined by peptide bridges (several "aa" in length). However, the fine structure and composition of the cell wall differ depending on the species. For instance, the cell wall is thick and enveloped by a capsule in gram positive bacteria (comprising peptidoglycans + teichoic acid) while it is thin and enveloped by a gelatinous layer in cyanobacteria. However, the cell wall of Gram-negative bacteria is thin peptidoglycan layer (without teichoic acid) and enveloped by a second lipid bilayer named outer membrane. The outer leaflet of the outer membrane in these bacteria contains lipopolysaccharides (LPS = endotoxin) rather than phospholipids whereas the inner leaflet contains special lipoproteins that anchor the outer membrane to the peptidoglycans (Figure 3.8). The outer membrane presents porins (for transport) and may be surrounded by a 11 Except mycoplasma which have steroids. Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  46 capsule. Bacillus subtillis is an example of gram positive and Escherichia coli is an example of gram-negative bacteria. The capsule is sometimes named glycocalyx and corresponds to the outer structure of cell envelope and is present in many species. It consists of extracellular polymers of polysaccharides. The capsule plays an important role in cell-cell and cell-support adhesion. For example, some bacteria adhere to tooth enamel by means of their capsules and cause plaque formation. The capsule is also involved in the invasiveness of pathogenic bacteria by preventing the process of phagocytosis. The pathogenicity of some bacterial strains depends on the presence of a capsule. d. Flagella Many bacteria have one or more flagella that serve for cellular locomotion (Figure 3.4,Figure 3.8). Flagella may be located at one pole of the cell, or they are distributed over the entire cell surface. Flagella are thread-like appendages composed of three parts: a filament, a hook, and a basal body. The filament is a polymer entirely made up of one protein type (flagellin) that forms hollow cylinder. The hook connects the filament to the basal body which is ankored in the envelope. The basal body generates flagellum rotation. As shown in Figure 3.9, the flagellum structures arise from the basal body in the cytoplasm and cross the plasma membrane, the cell wall and capsule when they are present. Unlike eukaryotic flagella, the bacterial flagellum is naked, not enveloped by extension of the plasma membrane. Cyanobacteria lack flagella. They usually occur as individual cells, as small clusters or colonies or as long filamentous chains. These bacteria are able to move by gliding due to a gelatinous layer that externally covers the cell wall. e. Pili (Fimbriae) Pili means hair and fimbriae means fringes. These are rigid appendages on cell surface of many bacteria (Figure 3.4, Figure 3.8). They are made up of proteins and are shorter and finer than flagella. They serve for bacteria-bacteria adhesion and conjugation (through sexual pilus, a very narrow channel that mediates passage of genetic material such as plasmid from one bacterium to another). Pili mediate also bacteria-host cells adhesion. In addition, pili may serve as receptors for viruses in the early stages of infection. f. Endospores Members of several bacterial genera can form endospores under condition of stress or nutritional depletion. An endospore is a resting cell that is highly resistant to desiccation (drought), heat and chemical stress. The endospore consists of the bacterial DNA plus a few cellular structures which are all enveloped by a membrane that is surrounded by special cell wall and coat (Figure 3.9). An endospore may resist harsh conditions for tens of years and when the medium is favorable again it germinates and reforms the original bacterium. IV. VIRUSES Viruses are not cells. They may be referred to as multimolecular complexes. They cannot divide by their own even if they are supplied with all the nutrients. This inability of division is since viruses lack metabolic enzymes and do not have the metabolic (anabolic and catabolic) pathways that are required for growth and division. Therefore, viruses are not living things. Viruses are associated with cells and live by exploiting them. They are responsible for some diseases in humans such as AIDS (acquired immunodeficiency syndrome), Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  47 smallpox, chickenpox, rabies, poliomyelitis, mumps, measles, hepatitis, mononucleosis, influenza and the common cold. In addition, some viruses may cause certain cancers. Viruses are extremely small but they are diverse (Figure 3.10) in terms of size (range between 20 to 200 nm), shape and structural organization. a) Tob sa,c rus (T"1V) (b) Bactenophage MS2 (c) Tomato bushy stunt virus (TBSV) Figure 3.10: Examples of viruses and phages of diverse shapes. Despite their diversity, nearly all viruses consist of one or more nucleic acid chain (DNA or RNA, single stranded or double stranded, linear or circular) assembled with very few proteins such as polymerases and integrase, and enveloped by a capsid which is made up of proteins. The capsid proteins are named capsomers (usually one or few types). These three types of components are common to most viruses. The term nucleocapsid is often used to describe the nucleic acid of the virus with the combined capsomer proteins. Certain viruses have their capsids enveloped by a lipid bilayer; others have complex tail anchored to the head (Figure 3.11). Furthermore, there are very simple viruses, named viroids, which represent an exception consisting of a naked nucleic acid (usually RNA) without any associated proteins. The typical viroid is an RNA molecule of about 50 nm in length. Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  48 Viruses do not contain the enzymatic equipment that is necessary for reproduction (no ribosomes, no oxidation of organic compounds, no energy production,...). Thus, viruses are not capable of autonomous growth or division, this is why they are not considered as living things and Reverse require specific host cells to multiply transcriptase inside them. Even the smallest cells......,. ,-Nucleic such as bacteria and mycoplasmas acid may be infected by viruses Viruses are, therefore, parasitic entities that infect plants and animals as well as bacteria. Host cells are infected by viruses which deviate the cell metabolism to their own benefit. In other words, a virus has proteins that alter cell metabolism (transcription, translation, replication,...) and deviate cell metabolism to promote viral a:--- ---Upid bilayer reproduction. The cell is therefore exploited by the virus. It should be known that the relationship between a virus and its host cell is relatively specific. As a RNA matter of fact, each viral species has 50nm a very limited host range infecting Figure 3.11: Viruses diversity. Diagrams of adenoviruse (a), related species, even related cell T-even bacteriophages (b) HIV (c) and influenza virus (d). types. Viruses are species-specific since viruses that infect chimpanzee cells for instance, do not infect human cells and vice versa (exceptions are possible). The virus-host cell specificity is even more restricted since certain viruses infect specific organs (e.g. lungs), tissues (e.g. epithelia), cell type or even sub-cell type. For instance, HIV virus specifically infects T4-lymphocytes but not other lymphocytes. The specificity of infection is due to requirement of specific receptor at the host cell surface which mediates viral entry into the host and infection. The virus-host specificity is, however, not absolute, for instance, influenza viruses may switch among different species provided that they recognize a specific receptor at the host cell surface. Most viruses undergo permanent change in their properties as they enter and leave their hosts. For instance, influenza viruses are divided into three types (A, B and C), each of these is present in different strains. For instance, many strains of influenza virus are known and continue to modify their make up every year. The H5N1 virus is a strain of the A-type which infects both birds and human beings. H1 N1 could infect humans and pig. "H" and "N" refer to the hemagglutinin and neuraminidase antigens that are present in the viral envelope. 1. Structure and classification of viruses The viral nucleic acids may be DNA or RNA and encode up to several tens of protein types including the capsid proteins and the polymerases that are necessary for viral nucleic acid replication and transcription. The viral capsid results from polymerization of Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  49 many protein subunits and may be enveloped by a lipid bilayer (containing proteins) in some types such as HIV, influenza and smallpox viruses. Capsid symmetry may be helical (example of TMV), polyhedral (example of adenovirus) or complex (example of bacteriophage). Certain virues have no defined symmetry. The capsid determines viral shape which may be either rod-like (e.g. tobacco mosaic virus, TMV, Figure 3.12), globular, spherical (e.g. HIV), polyhedral (e.g. adenovirus), helical or filamentous. Moreover, there are complex viruses that are tailed­ polyhedral such as bacteriophages T2, T4 and T7, Figure 3.11. These viruses have a tail-like Protein coat (capsid) structure that emerges from the polyhedral head (a) Nucleic acid which houses the viral nucleic acids. The tail consists of a sheath of proteins arranged in a helical pattern. Tail extremity shows specialized appendages (tail fibers) which are involved in attachment of the virus to the receptor at the host cell surface. Viruses may be classified into families depending on their size, shape, symmetry mode, the presence of lipid bilayer,... Moreover, viruses may be classified depending on the nature of their nucleic acid content (DNA or RNA). Such nucleic acid-based classification divides viruses into DNA-containing viruses and RNA-containing (b) 50nm Figure 3.12: Tobacco Mosaic Virus scheme viruses; each of these two broad classes and TEM image. comprises several families. Retroviruses such as the HIV (containing one RNA type) is an example of class of RNA-containing viruses, also influenza viruses contain RNA (8 different types) but it is not a retrovirus and belongs to the family of orthomyxoviruses. The family of adenoviruses (contains DNA) comprises many types, some of them cause diseases in the respiratory system and in the eyes. Since the aim of this chapter is not to describe all families of viruses, only certain examples of viral families are presented. Finally, a host cell-based classification divides viruses into bacteriophages and viruses which infect prokaryotes and eukaryotes, respectively. Viruses that infect prokaryotes are also referred to as phages. 2. Proliferation of viruses In spite of the structural differences between viruses, nearly all viruses follow the same principles to infect and proliferate inside host cells. All viruses reproduce exclusively within host cells and exploit the host cell metabolism. After recognition between the viral capsid (or membrane when it is present) and a specific receptor at the host surface, the virus (phage) adheres to the host surface and injects its nucleic acid inside the host cell. Once the viral nucleic acid is delivered to the cytoplasm of the host, it chooses between the lytic or lysogenic cycles (strategies) that are presented in the next paragraph. Ultimately, hundreds or thousands of new viruses (phages) are produced by the host cell which will eventually die. So, the first step in infection process is adhesion of the viral particle on its host cell surface (Figure 3.13). This adhesion involves viral proteins belonging to the capsid or the lipid bilayer, and receptors exposed at the host cell surface. For instance, the membrane protein CD4 of T4 lymphocytes is the receptor for HIV and thus mediates the infection process. It is a glycoprotein of the viral envelope that binds to CD4 receptor. Hemagglutinin of influenza virus mediates recognition with a specific receptor (glycophorin A) at the surface of host cell membrane in the respiratory tract epithelium. Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  50 The second step of the viral cycle aims at introducing the viral nucleic acid and the associated proteins (e.g. polymerases, integrases) into the cytoplasm. Different modes of injection are possible. The viral nucleic acid holds the information required for viral reproduction. Once a virus attaches to its host cell, it injects its nucleic acid into the cytoplasm. In case of viruses whose capsid is enveloped by a lipid bilayer, fusion between cell membrane and viral membrane releases the whole capsid in the cytoplasm where it dissociates in order to deliver the viral nucleic acid (Figure 3.14). If the virus lacks a lipid bilayer, the capsid is kept outside the host cell and only the nucleic acid is injected into the H ,ds are pack d w th DNA, Bacterial cell lyHs, r, I lng compl t d LYTIC I f tiv ph CYCLE o Th ph DNA dir ct > ch s to the cell's metabolism to produce viral r It on compon nt -prot in another bacterial cell ndcopl ofph @----....- wall, penetrates it. DNA. ndln DNA, Phag ONA in rts it If (as a prophage) into bact rial chromosom Bin ry fission I completed; each cell has th ph g ONA prior to binary fission. incorporated. Figure 3.13: Basic principle of viral proliferation (lytic and lysogenic cycles). host cell through specific channel (e.g. tail of bacteriophage T4). For most viruses, the third step will prepare the nucleic acid for one of the two possible cycles, lysogenic or lytic. If the viral nucleic acid is an RNA it may be retrotranscribed into DNA by a reverse transcriptase usually associated with the viral genomic RNA in retroviruses (Figure 3.14). Therefore, after infection, viruses as well as retroviruses have DNA genomes named provirus. If reverse transcription is not possible (reverse transcriptase not available), the viral RNA serves as template for replication by RNA­ dependent RNA polymerases, also named replicases. The choice between lytic and lysogenic pathways is determined by specific viral proteins but also it depends on the host cell conditions. If the lysogenic pathway is adopted, the viral DNA will integrate (insert) in the host's chromosome by a process of recombination. Integration is catalyzed by integrases which are encoded by genes present in most viral genomes. The host cell is referred to as lysogenic for the virus in question when a copy of the viral DNA is integrated inside it. Such integrated DNA is silent without any active effect. The cells seem to be normal and continue their life normally. As the lysogenic cell divides, the integrated DNA replicates along with the host chromosome and will be transmitted to the daughter cells silently. Thus, the virus is reproducing indirectly through cell division since all daughter cells will be lysogenic. This is a kind of silent reproduction for the virus (or phage). If the lytic strategy is preferred by the virus, integration may occur or not according to the virus type. But the main event is that cell metabolism is deviated by some specific viral Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  51 proteins so that viral genes are preferably transcribed and the produced RNA translated into proteins (e.g. capsid proteins) by the host cell equipments. Afterward, the viral genome is reproduced by replication or by transcription (depending on whether the viral genome is DNA or RNA). Now that all viral Retrovirus particle components are ready RNAgenome (viral capsid proteins and viral nucleic acids), new viral particles result from assembly between viral nucleic acids and capsid proteins that polymerize forming a cage all around the nucleic acid. Viruses that adopt a DNA provirus lysogenic strategy do not stay forever in that state. In fact, the lysogenic cells can revert to the lytic cycle at any time in response to diverse signals from the Figure 3.14: Reproduction of retroviruses surrounding medium (e.g. UV irradiation). After the switch to the lytic pathway, the virus behaves in the same way as previously described to produce new viral particles. The fifth step is the release of the newly produced viruses (Figure 3.13 to Figure 3.15). Those that are devoid of lipid envelope (e.g. bacteriophage T4, bacteriophage A) are released by destruction of the host cell envelope by specific enzymes produced by the virus thereby causing cell lysis. However, viruses that were initially enveloped by a lipid bilayer will bud from the host cell surface. In fact, during formation of new nucleocapsids in the cytoplasm, other viral proteins are produced and exported to the host cell plasma membrane where they are inserted. Viral membrane proteins are gathered at a specific pole of the cell membrane Figure 3.15: TEM images of phages adhered to their host (left) and retrovirus where newly (right) released from their host cells. produced nucleocapsids move and interact with the inserted viral membrane proteins (Figure 3.14). The interaction causes every capsid to be enveloped by a membrane fragment, forms a bud and detaches from the cell surface. Therefore, the host cell is a factory that Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)  52 continuously produces viruses. Nevertheless, such a cell will ultimately die since its metabolism is seriously disturbed. It is worth mentioning that certain visuses that adopt lysogenic state in animal cells may lead to cell transformation and cancer development. V. PRIONS Prions are unusual pathogen agents since they consist of proteins only. In other words, prions, unlike other pathogenic agents (such as viruses, certain bacteria, certain protozoa and certain fungi) do not have nucleic acids. A prion could be defined as a protease­ resistant protein that has an abnormal conformation and which can transmit its abnormal conformation to other copies of proteins inside the cell. Since prions are Figure 3.16: Secondary and tertiary structures Protease-resistant, they accumulate inside the of the normal PrPc and PrPres. cell and lead to toxicity by interfering with diverse cell functions such as membrane polarity, signal transduction, ion transport,... Transmissible spongiform encephalopathies (TSE) are prion-caused diseases of humans and animals. Such diseases are characterized by fatal neurodegeneration (neuron death). Although some TSE, like scrapie in sheep, have been known to exist for centuries, bovine spongiform encephalopathy (BSE) was recognized in the late 20 th century. The new variant Creutzfeldt-Jacob disease in humans is probably caused by consumption of BSE-infected food. The infectious agent is devoid of nucleic acids and consists at least in part of an abnormal form of a host protein named PrPc (PrPc for prion­ related protein-cellular) which is naturally produced in nerve and blood cells. The hallmark of these progressive neurodegenerative diseases is the accumulation of the protease­ resistant, pathologic conformation of prion protein (named PrPres, "res" for resistant) in the central nervous system (Figure 3.16). Moreover, PrPc is sensitive to proteolysis, but PrPres is protease-resistant, which forms insoluble fibrils. The 3D structure change is thought to be propagated by a template-like effect in which a normal PrPc interacts with an abnormal PrPres isoform and assumes the abnormal pathologic conformation which is resistant to protease. Lebanese University-FS1. B1100 - Cell Biology by Pr Ziad ABDEL-RAZZAK (2022)