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MICROBIOLOGY
SARS Cellular Receptor DiscoveredFindings Could Lead Quickly to Drug and Vaccine Therapies Against the DiseaseFor a few panicked months last winter, SARS seemed to displace HIV as reigning viral uber-menace. By the time the last outbreak was over in late spring, the SARS virus had killed more than 750 people and had debilitated nearly 10 times as many. Now, HMS researchers, working with scientists at other institutions, have identified a cellular receptor that enables the virus to gain a foothold in the human body, a discovery that may lead to drug and vaccine strategies to quell the upstart virus. The findings appear in the Nov. 27 Nature.
Work on HIV gave Michael Farzan (second from right) and colleagues a head start on probing the SARS virus. With him (left to right) are co-authors Michael Moore, a technician in Farzan's lab, Hyeryun Choe, and Wenhui Li. (Photo by Graham Ramsay) The receptor, ACE2, belongs to a well-known family of proteins, the angiotensin-converting enzymes, involved in the contraction of smooth vascular muscle and the resulting rise in blood pressure. Though not as famous as its cousin ACE1, which is targeted by blood pressure- lowering ACE inhibitors, ACE2 has been garnering interest for its potential role in hypertension and inflammation. The pharmaceutical industry is developing peptides and small molecules against the enzyme, and anti-ACE2 antibodies already exist. In fact, in their experiments, Michael Farzan, Wenhui Li, Hyeryun Choe, and their colleagues found that they could slow replication of the SARS virus by exposing infected cells to these antibodies.
"It may be that the S protein we are working with every day in the lab could function as a SARS vaccine. In general, a SARS vaccine should be significantly easier to develop than one against HIV," said Farzan. HIV's LeadThe researchers' familiarity with HIV entry--Farzan and Choe, HMS assistant professor of pediatrics at Children's, have been working on the problem for more than eight years-- may have given the team an edge in cracking the SARS mystery. When the SARS virus was first identified in March, little was known about it other than that it belongs to the coronavirus family. "We were watching this from afar and did not have much to do with it until the genome became available in late April," said Farzan. Intrigued by the similarities between HIV and coronavirus entry--both use fusion proteins--Li, an HMS research fellow in medicine at BWH, Choe, Farzan, and colleagues set out to build the gene for the SARS virus S protein, which has an unusually hefty 1,250 amino acids. "That is larger than most people have built," said Farzan. Boosted by a combination of "late nights plus luck," and the personal motivation of Li, who hails from China, the epicenter of the epidemic, the researchers had the gene by the end of May.Synthesizing the protein, though comparatively easy, would be the critical step. "Building the S protein was what put us ahead," Farzan said. With the protein in hand, the researchers embarked on a series of experiments. They mixed the S protein with a line of monkey kidney cells known to host SARS infection. The protein bound to the cells. To see exactly which receptor it was binding, they used antibodies to pull the S protein out of the cells in hopes that the receptor would remain attached and be dragged out as well. They pulled out three protein fragments and, thanks to the human genome project and advances in mass spectrometry, had them identified by the end of July. "Everything that took a month here took a year in the context of AIDS," Farzan commented. Only one of the proteins, ACE2, was expressed in the right location, on the cell surface, and in the right tissues, namely those susceptible to SARS--the lungs, kidneys, gastrointestinal tract, and heart. The receptor would go on to pass a whole series of tests. The first was a fusion assay. Occasionally, proteins that mediate fusion between patches of viral and cellular membrane can cause whole cells to fuse, creating multinucleated clusters, or syncytia. When mixed with cells expressing the SARS S protein, ACE2-expressing cells formed syncytia. ACE2 also passed the infection assay. In this test, cells were transfected with ACE2, mixed with SARS S protein, and checked several days later for signs of viral replication. "There was a ton more virus coming out of the ACE2-transfected cells compared with those without the receptor," said Farzan. Though some ACE2-transfected cells were found to produce as much as ten thousand-fold more SARS virus, this replication could be slowed by the simple application of anti-ACE2, but not anti-ACE1, antibodies. Picturing Viral FusionWhat makes the ACE2-bearing cells such efficient SARS-churning machines--and why is the virus so deadly in humans? One possibility is that the human ACE2 receptor is just more efficient at letting the virus in.Farzan and his colleagues are hoping to get a better handle on these and other questions about SARS entry. If the pace of discovery keeps up, insights from his lab might reflect back the other way, from SARS to HIV. "The two viruses exhibit certain similarities regarding entry--this very complex fusion, which is what we are most interested in," Farzan said. "They both do it in slightly different ways, but probably very similar ways. By looking at both of them, we hope to get a better picture." --Misia Landau |
