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HIV Pathogenesis: In vivo Findings Suggests New Model of HIV-1 Nef Protein Downregulation of CD4 <p></b>
Prepared by AIDS Weekly editors from staff and other reports
January 22, 2001
-- Gladstone Institute, San Francisco, researchers have developed a new in vivo co-immunoprecipitation assay to investigate the mechanism by which the HIV-1 Nef protein binds to CD4 cells.

Based on the results of their assay, they suggest that surface downregulation by Nef occurs via collaboration of endocytosis motifs of both CD4 di-leucine and Nef.

Wes Yonemoto and Warner C. Greene reported their findings at the 40th annual meeting of the American Society for Cell Biology, held December 9-13, 2000, in San Francisco, California. The title of their presentation was "Biochemical analysis of the in vivo interaction between HIV-1 Nef and CD4."

CD4 is a component of the HIV-1 co-receptor complex, Yonemoto and Greene pointed out. "We have previously characterized a leucine-based endocytosis motif within the C-terminal region of Nef, which is critical for adaptor binding and CD4 regulation," they said.

They also referred to other research, conducted in vitro, that identified a di-leucine sequence (LL413/414) that occurs within CD4's cytoplasmic tail and which serves as Nef's binding site.

"This result correlates with previous work identifying LL413/414 as CD4 residues essential for downregulation by Nef. It is also important to note that LL413/414 have also been shown to function as a leucine-based endocytosis motif, in the absence of Nef," Yonemoto told the audience.

The in vivo co-immunoprecipitation assay the team developed uses wild-type and mutant CD4 proteins, they explained. Using it, they determined that, contrary to in vitro study findings, LL413/414 isn't necessary for Nef binding to occur.

"In addition," Yonemoto and Greene reported, "analysis of a panel of Nef mutants indicates that residues WL57/58, identified to bind in vitro to LL413/414, are not required for interaction with CD4.

"Therefore," they concluded, "our data indicate that the presumed site(s) of in vivo interaction between HIV-1 Nef and CD4 is incorrect. Instead, these results suggest a new model for targeting of CD4 by Nef, in which the CD4 di-leucine endocytosis motif works in concert with the endocytosis motif found within Nef, to achieve surface downregulation."

Wes Yonemoto was the presenting author of this study to the conference. His address is Virology and Immunology, Gladstone Institutes, PO Box 419100, San Francisco, CA 94114, USA.

Key points reported in this study include:

  • Previous research findings, based on in vitro studies, suggested that the di-leucine sequence, LL413/414, in CD4 cells is necessary for surface downregulation by the HIV-1 Nef protein

  • The current study was conducted in vivo, and used a new co-immunoprecipitation assay based on wild-type and mutant CD4 proteins

  • The in vivo study results suggest, contrary to in vitro studies, that LL413/414 is not required for binding to Nef

  • Instead, the CD4 di-leucine endocytosis motif and the Nef endocytosis motif may work together for surface downregulation to occur

This article was prepared by AIDS Weekly editors from staff and other reports.



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