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Lwoff's Pathways - Viral Replication

CellLwoff's Pathways - Viral Replication

Chris Evers
Document originally from Genentech's Access Excellence

Virus is from the Latin word for "poison." The term remains an apt one since viruses constitute one of the greatest threats to human health. One group of viruses, Herpes simplex (HSV), is the most commonly transmitted pathogen in industrialized countries. One type of HSV causes genital herpes - an incurable, sexually transmitted disease suffered by 30 million people in the United States.

HSV illustrates an interesting property of certain viruses - it can take both active and latent forms. During the active phase, the virus interferes with the cell's normal metabolism, causing the symptoms associated with the disease - including painful genital blisters. During the latent phase, it's as if the virus has gone to sleep. Although the host cells remain infected, the host is a symptom-free carrier of the disease.

We now know the difference between the active and latent manifestations of viral infection is a result of a switch in viral replication patterns. Some viruses can only replicate by what is called the lytic pathway. They enter and inject a host cell with DNA, forcing it to make new viruses. At some point bursting at the seams with the viruses, the host breaks open, releasing new pathogens into the environment. Other viruses operate differently: they enter and inject their DNA into the host cell - but instead of taking over the host cell and using it to make viruses, the injected DNA can become inactive for some time, until the appropriate cellular event triggers its awakening. This latter pathway is called the temperate or lysogenic pathway.

The lysogenic pathway was discovered in bacterial viruses (bacteriophages) in the early 1920s, but was not really understood until the 1950s, when it was explored at the cellular level by Andre' Lwoff, a French scientist. At that time, it was known that some bacterial cultures that grew normally and otherwise seemed perfectly healthy were actually infected by phage. Although the phage didn't seem to interfere with the host bacteria, such cultures had the ability to cause the lysis or rupture of other bacteria. Thus, the culture was described as "lysogenic."

It was not clear why such cultures were lethal to other bacteria. The lysogenic effect didn't seem to stem from phage particles floating around in the culture, since the cultures remained lethal even after treatment by methods that removed any free floating phage. Nor was the effect due to some large reserve of phage stored within the host cells - no phage were released when the cells of a lysogenic culture were artificially caused to burst open.

Lwoff's definitive studies on the subject relied on the time-honored technique of persistent, careful observations. He observed the growth of single bacterial cells of Bacillus megaterium, a really large bacterium, in tiny droplets of medium. Close observation revealed the secret - although free phage particles were never found floating around in droplets that contained only single cells, they were found in the small colonies derived from single cells. Where did they come from? Lwoff actually observed the answer - occasionally, while a culture in a droplet was being watched, a single cell in it would spontaneously burst, releasing about 100 phage.

Lwoff concluded that the host cells were not really entirely immune to the phage. When a phage did become active in a bacterium in the lysogenic culture, it forced the host to manufacture more phage, eventually killing the host, and releasing new phage when the cell burst open. However, the switch from the lysogenic to the lytic pathway was the exception rather than the rule. Most of the time, in most of the bacterial host cells, the phage was in an inactive form. Later, Lwoff found that it was possible to artificially induce all the cells in a lysogenic culture to enter the lytic pathway simultaneously by exposing the cultures to UV light, or X-rays.

The lytic and the lysogenic pathways of viral replication are illustrated.

Figure Legend:

Lytic Pathway:
In lysis, the host cell's membrane is damaged and the cytoplasm is able to leak out. The cell dies rapidly, releasing virus particles.

Lysogenic Pathway:
The lysogenic path is one of several temperate pathways in which the host cell is not killed outright, but is occupied by the virus and used as a factory for replicating the viral genes. The viral infection enters a latent period during this process.

In the illustrated instance, genetic recombination occurs: The viral enzyme cuts the host chromosome and integrates viral genes into it. When the recombinant DNA is replicated, it passes the viral DNA onto all of its descendents. If the viral genes later undergo lysis and leave the cell, a new cycle of infection will begin.

Lwoff's careful experiments laid the groundwork for further research on the interaction between the phage and their hosts in lysogenic cultures - research that turned out to have far-reaching implications. It was by studying this interaction that he made an important discovery: that the genes injected into a bacterium by a phage can actually become integrated into the host's genome - a discovery that is the basis for much of genetic engineering.

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