Hepatitis C Virus Takes Charge, Changes Shape of Cellular Machinery

NYS DOH Wadsworth Center Finding May Aid Design of New Drugs to Fight the Disease

Albany, March 9, 2001 – A major advance in understanding hepatitis C infection may facilitate the design of new drugs to treat the disease, which affects nearly 4 million Americans and 170 million people worldwide. Research in the health department's Wadsworth Center laboratories has shown that hepatitis C virus hijacks a host cell's protein factory by changing its shape in a way that favors viral proteins being produced, rather than host proteins. This is the first time the genetic material of a virus has been captured in the process of reprogramming the cell in this way. The finding is published in the March 9 issue of Science.

The groundbreaking work was done by Dr. Joachim Frank, a Howard Hughes Medical Institute investigator at Wadsworth Center, in collaboration with researchers at Yale University. Dr. Frank studies ribosomes, cellular machinery that carries out the synthesis of all proteins. He has pioneered the use of cryo–electron microscopy and his own image processing software to reconstruct maps of large biological molecules. This is the first picture of viral genetic material bound to a ribosome at the start of protein synthesis.

"Dr. Frank's research epitomizes the quality of scientific research conducted in New York State and illustrates how cutting–edge basic science can address a problem of pressing public health importance," State Health Commissioner Dr. Antonia C. Novello said.

The majority of people infected with hepatitis C (HCV) develop chronic liver disease, cirrhosis or liver cancer. The Centers for Disease Control and Prevention estimates that HCV costs the U.S. $600 million a year in health–care costs and lost wages. Available therapies fail in many cases, resulting in 10,000 deaths annually in this country. A drug that attacks molecular targets presented by the virus, without harming the tissues and organs the virus infects, would be a significant advance. Ribosomes may be just such a target.

Ribosomes read the genetic code and translate its message into proteins, the molecular workhorses of life. Ribosomes are themselves composed of proteins and ribonucleic acid (RNA). Earlier three–dimensional reconstruction efforts by Dr. Frank and others showed the structure of the two unequal subunits of this giant macromolecule. In the case of eukaryotes (organisms whose genetic material in enclosed in a nucleus), these are 40S and 60S subunits.

The present work shows a reconstruction of HCV RNA bound to the 40S ribosomal subunit from rabbits. The detailed images obtained by the Frank group show how the virus directs the cell to synthesize its own proteins for constructing more viruses, leading to the eventual destruction of the infected cell.

In normal cells, messenger RNAs that code for proteins are recognized by a collection of helper molecules, named initiation factors, which bind them to ribosomes and instruct the ribosomes where to begin translating their sequence into proteins.

Hepatitis C RNA, by contrast, contains a complex structure at one end that allows it to directly interact with ribosomes, dispensing with the need for most initiation factors. This structure, an internal ribosomal entry site (IRES), gives the viral RNAs an unfair advantage over normal cellular RNAs, allowing it to subvert the cell's machinery for its own purpose. The virus gains a competitive edge by forcing a dramatic change in the shape of the ribosome, placing itself in the optimal position to then be decoded into a protein.

"Before, this process was a complete puzzle," says Dr. Frank. "People knew that viruses would introduce their messenger RNAs, but how could that be done to the exclusion of host messenger RNAs being expressed? Our reconstruction shows how the host's mechanism is completely disabled."

Dr. Frank's combination of cryo–electron microscopy (cryo–EM) and image processing captures ribosomes and other macromolecular machines in their native, dynamic states. Thousands of ribosomes are rapidly frozen and their images obtained by an electron microscope with a low–intensity beam. Using the SPIDER software that he developed, thousands of the cryo–EM images are transformed into a three–dimensional map. A complementary structural technique, x–ray crystallography, provides atomic–level detail of static, biological molecules. Together, they can point pharmaceutical researchers to a molecule's active site so that drugs can be developed to initiate or inhibit activity.

Co–authors of the Science paper are Christian Spahn, Robert Grassucci and Pawel Penczek of the Wadsworth Center, and Jeffrey S. Kieft, Kaihong Zhou and Jennifer Doudna of Yale University. The work was supported by the Howard Hughes Medical Institute and grants from the National Institutes of Health and the National Science Foundation.

Wadsworth Center is the public health and research laboratory of the New York State Department of Health. Wadsworth's role in detecting and responding to disease threats, and ensuring the quality of laboratories services received by state residents, is complemented by a longstanding commitment to biomedical and environmental research. The Center also houses the Resource for the Visualization of Biological Complexity, a National Institutes of Health–supported biotechnology facility directed by Dr. Frank.

3/9/01–21 OPA