TMV, BACTERIOPHAGE, LIFE CYCLE OF PLANT VIRUS, LIFE CYCLE OF BACTERIOPHAGE

TOBACCO MOSAIC VIRUS (TMV)

            SYSTEMATIC POSITION

            Division: Vira

                Sub-division: Ribovira

                     Class: Ribohelica

                          Order: Virales

                                Sub-order: Phytophagineae

                                      Family: Marmoraceae

                                           Genus: Marmor

            ULTRASTRUCTURE OF TMV

            Tobacco Mosaic Virus (TMV), the fine structure of which has been extensively studied, is taken as a type to illustrate the ultrastructure of plant viruses.

            (a) Structure:- The virus particles or virons of TMV are straight, rigid cylindrical rods with a helical structure and remarkably uniform outer measurements of 18nm in diameter and 300 nm in length. The rods are hollow. The hollow core or cavity which is 4 nm wide extends the entire length of the virus particle (viron) and is bounded by 14 nm thick walls. Thus in a cross section, the viron consists of two parts, namely – (i) an outer protective protein coat or shell called the capsid, and (ii) the inner infective agent, the genetic material or the nucleic acid. The capsid is built of about 2200 small, similar, protein subunits called capsomers. They are closely packed and arranged in a regular left handed helical around the RNA helix. The nucleic acid in plant virus is generally, a single stranded RNA which in TMV is in the form of a long helix. It extends the entire length of the helix.

           (b) Composition:- Virus protein comprises 95% of the total substance of the virus particle. The remaining 5% is RNA. Both protein and RNA are co-axially inter-winged to form a rod shaped virons with helical architecture. The molecular weight of RNA molecule is about 2 millions. It provides a code which controls the amino acid sequence in the capsid.

            The virus protein which forms the capsid is similar to other plant proteins and is made up of usual amino acids joined by the peptide links. The virus proteins, however, are unusual in their size and are of high molecular weight. The molecular weight of TMV protein is nearly 40 million. Each capsomere has a molecular weight of 17,400 and is formed by the condensation of about 158 amino acid residues. The capsid forms a tightly fitting layer. The virus particles remain ineffective for about 50 years in dried tobacco leaves. They can stand boiling for 10 minutes.

            LIFE CYCLE OF PLANT VIRUSES

            The life cycle of a typical plant virus such as TMV comprises 4 stages –

            Stage1. Transmission of virus particles (Virions):- The rod shaped Tobacco Mosaic Virus (TMV) infects only the leaves of tobacco plants. It is transmitted from one host plant to another by insect vectors such as different aphids, grasshoppers, etc. The virus can also be transmitted by mechanical means such as rubbing of virus on the leaves and through handling of tobacco plants at transplanting. The virus is extremely contagious. A person who has handled cigarettes containing infected tobacco leaves may transmit the virus to the healthy plants.

            Stage2. Infection:- Infection takes place with the whole virus particle. The protein coat i.e., capsid alone is not capable of producing the infection. The RNA alone, however, is infectious but much less than that of the intact virus particle. But when the protein coat is placed in a solution of viral RNA, the viral particles of TMV are reconstituted. They were able to re-infect tobacco leaves. It shows that the nucleic acid alone carries full instructions to form new virus particle.

            Stage3. Multiplication:- After infection, the virus particle disappears from view. It ceases to exist as an organized unit and breaks up into its two components parts in the host cytoplasm. The viral RNA is thus released from the protein coat. Uncoating of the viral RNA is the initial step towards viral multiplication. It takes place within minutes after the viron gains entry into the host cell. This is known as eclipse stage. Thereafter starts the multiplication process. Viral protein coat remains in the host cell cytoplasm. The RNA component moves from cell to cell causing the production of new virus components which assemble to form more virus particle.

            Stage4. Sites of Viral RNA and Viral Protein synthesis:- There are various views regarding the synthesis of viral RNA and protein. The widely held view is that the virus protein and RNA takes control of the cell machinery and uses it to produce the viral components instead of the host cell parts. The viral components are then assembled into new virus particles.

            One view suggests that the viral nucleic acid combines with or replaces part of that of the host cell. Thereafter the host cell produces the viral components at the expense of its own.

            The other commonly held view is that the DNA of the host cell uses viral RNA to elaborate more viral RNA and protein. They are elaborated in the cytoplasm of the host cell, first the viral RNA and then the viral protein.

            The widely accepted view is that viral components are formed by two separate systems. The release viral RNA reaches the nucleus and takes control of the host cell machinery and uses it to produce viral components instead of host cell parts. The nucleolus in the nucleus produces the new viral RNA which escapes to the cytoplasm around the nucleus through a channel present in the nuclear membrane. The TMV protein is synthesized in the cytoplasm very near to the nucleus on cytoplasmic ribosomes.

            The assembly of new TMV particles occurs in the cytoplasm on endoplasmic reticulum. The eclipse stage last for about 6 hours. By that time the first progeny of TMV occurs in the cytoplasm. The result of experiments with fluorescent microscopy reveals that TMV protein and RNA both are synthesized within the host cell nucleus. The assembled TMV virons or particles escape into the host cell cytoplasm where they accumulate.

BACTERIOPHAGE

            SYSTEMATIC POSITION

            Division: Vira

                 Sub-division: Deoxyvira

                       Class: Deoxybinala

                             Order: Virales

                                    Sub-order: Phagineae

                                         Common Name: Bacteriophage

            ULTRASTRUCTURE OF BACTERIOPHAGE

            The bacteriophage (T-even phage) particle resembles a tiny sperm. It has a head and a tail. In addition there is some sort of attachment region adapted to stick to the surface of the host cell. The virus particle of T-even phage is about 200 to 280µm in length. The virus particle is thus too small to be seen with the best of light microscope. Pictures of virus particles are obtained by the use of electron microscope.

The head of T-even phage is hexagonal in outline and bears numerous facets. It consists of protein coat surrounding a core of genetic material which is a DNA molecule. The coiled coil of a single thread-like double-stranded macro molecule of DNA is packed tightly in the head. It is about 50 microns long. The phage DNA is said to lack base cytosine. Instead it has another base hydromethylcytosine (HMC).

            The cylindrical tail entirely consists of a protein sheath surrounding an empty core. The tail sheath can contract longitudinally. The attachment apparatus of the phage consists of six long, slender protein fibres known as tail or caudal fibres. They arise from a plate at the basal end of the tail. The tail fibres normally remain twined inside the core of the tail. Extended they help to attach the phage particles to the coli cell.

            LIFE CYCLE OF BACTERIOPHAGE

            Bacteriophages exhibit two different types of life cycle – lytic cycle and lysogenic cycle. In the virulent or lytic cycle, intracellular multiplication of the phage culminates in the lysis of the host bacterium and the release of progeny virions. In the temperate or lysogenic cycle, the phage DNA becomes integrated with the bacterial genome, replicating synchronously with it, causing no harm to the host cell.

            1. LYTIC CYCLE:- Lytic cycle of virulent phage can be considered in the following stages – adsorption, penetration, synthesis of phage components, assembly, maturation and release of the progeny phage particles.

            In adsorption, the phage particles come in contact with the bacterial cell by random collision. A phage attaches to the surface of a susceptible bacterium by its tail. Under optimum condition, adsorption is very rapid process, being complete within minutes. Bacterial protoplasts, which are devoid of cell wall components, cannot adsorb phages and therefore will not be infected. Host specificity of phages is determined at the level of adsorption.

            Adsorption is followed by penetration of the phage nucleic acid (DNA) into the bacterial cell. The process of penetration resembles injection through a syringe. The base plate and tail fibres are held firmly against the cell causing the hollow core to pierce through the cell wall. The phage DNA is injected into the bacterial body through the hollow core. After injection of the DNA, the empty head and tail of the phage remain outside the bacterium as the shell or ‘ghost’.

            Immediately after the penetration of the phage nucleic acid, the synthesis of the phage components is initiated. The first products to be synthesized are the enzymes necessary for building of the complex molecules peculiar to the phage. Subsequently, the protein subunits of the phage head and tail are synthesized. Phage DNA, head protein and tail protein are synthesized separately in the bacterial cell.

            This is followed by assembly of the phage components. The DNA is condensed into a compact polyhedron and packaged into the head and finally, the tail structures are added. This assembly of the phage components into the mature infective phage particle is known as maturation.

            Release of the mature progeny phages typically occurs by lysis of the bacterial cell. The bacterial cell wall bursts or lyse, resulting in the release of mature daughter phages.

            The interval between the entry of the phage DNA into the bacterial cell and the appearance of the first infectious intracellular phage particle is known as eclipse phase. It represents the time required for the synthesis of phage components and their assembly into mature phage particles. The interval between the infection of a bacterial cell and the first release of infectious phage particles is known as the latent period.

            2. LYSOGENIC CYCLE:- In this cycle the pacteriophage enter into a symbiotic relationship with their host cells without destroying them. Following entry into the host cell, the bacteriophage DNA becomes integrated with the bacterial chromosome. The integrated phage nucleic acid is known as prophage. The prophage behaves like a segment of host chromosome and replicates synchronously with it. This phenomenon is called lysogeny and a bacterium that carries a prophage within its genome is called lysogenic bacterium.

            During the multiplication of lysogenic bacteria, the prophage may become excised from occasional cells. The excised prophage initiates lytic replication and the daughter prophage particles are released which infect the other bacterial cells and render them lysogenic. The lysogenic bacteria in a population can be induced to shift to the lytic cycle by exposure to certain physical and chemical agents. Such inducing agents include UV rays, hydrogen peroxide and nitrogen mustard.

            The lysogenic bacterium is resistant to reinfection by the same or related phages. This is known as superinfection immunity.

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