Exploring a Family Relationship

Herlocher has sought to find what features of the structure of this cold-adapted virus could explain why it lives happily only in nasal climes while its parent thrives in the moist, warm folds of the lungs. In particular, her work focuses on the eight strands of genetic material -- composed of ribonucleic acid (RNA) -- packed within the flu particle's protein and lipid outer shell. These eight genes are comprised of long strands of thousands of chemical bases, and the sequence of these bases determines what amino acids will be synthesized to form the viral proteins. The long RNA strands flop around and become looped, folded structures, and the shape of these complex folds may play a role in the activity of the virus.

"We're looking at whether we can attach function to folding patterns," she says. "For instance, we want to determine how each viral particle can package its genetic material and how the RNA fold might accomplish this function. In addition, the RNA folding may affect the transport of genetic material in and out of a host cell's nucleus where the viral RNA is reproduced." It may turn out, Herlocher says, that the cold-adapted virus loses its virulence because it's transported or packaged poorly at the higher temperature of the lungs.

In Dr. Robert Webster's laboratory at St. Jude, Herlocher did a sequence analysis of the genes in the cold-adapted virus and its parent, comparing all the chemical bases (10,398) with the corresponding bases in the other. She found only four differences or mutations in the cold-adapted virus. She then did detailed RNA-folding calculations at Pittsburgh. Using the CRAY Y-MP and a program that predicts RNA folding developed by Michael Zuker of the National Research Council of Canada, Herlocher did the folding simulation for all eight genes of the cold-adapted virus and its parent at body temperature (37 degrees C). Results showed different folding in two genes, and one of them (PB2) is known to control temperature sensitivity. Though this gene contained two of the four mutations, it synthesizes the same amino acids as the parent. This suggests, Herlocher says, that the RNA folds rather than purely the amino-acid sequence have a significant role in cold adaptation.

Herlocher is now doing laboratory analysis to confirm the RNA-folding predicted by her calculations. This work required 30 hours of CRAY Y-MP time and, says Herlocher, wouldn't have been practical without supercomputing. "It takes 12 hours to do a half gene on a VAX, and this system doesn't have enough memory to do a full gene." Laboratory methods are also limited. "It's impossible to determine every possible RNA fold in the laboratory," and as compared to months and years of lab work, the CRAY requires only 30 minutes to determine the RNA fold of a single gene.

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