Strict Conservation of the Retroviral Nucleocapsid Protein Zinc Finger Is Strongly Influenced by Its Role in Viral Infection Processes: Characterization of HIV-1 Particles Containing Mutant Nucleocapsid Zinc-Coordinating Sequences

Virology. 1999 Mar 30;256(1):92-104. doi: 10.1006/viro.1999.9629.

Abstract

The retroviral nucleocapsid (NC) protein contains highly conserved amino acid sequences (-Cys-X2-Cys-X4-His-X4-Cys-) designated retroviral (CCHC) Zn2+ fingers. The NC protein of murine leukemia viruses contains one NC Zn2+ finger and mutants that were competent in metal binding (CCCC and CCHH) packaged wild-type levels of full-length viral RNA but were not infectious. These studies were extended to human immunodeficiency virus type 1 (HIV-1), a virus with two NC Zn2+ fingers. Viruses with combinations of CCHC, CCCC, and CCHH Zn2+ fingers in each position of HIV-1 NC were characterized. Mutant particles contained the normal complement of processed viral proteins. Four mutants packaged roughly wild-type levels of genomic RNA, whereas the remaining mutants packaged reduced levels. Virions with mutated C-terminal position NC fingers were replication competent. One interesting mutant, containing a CCCC Zn2+ finger in the N-terminal position of NC, packaged wild-type levels of viral RNA and showed approximately 5% wild-type levels of infectivity when examined in CD4-expressing HeLa cells containing an HIV-1 LTR/beta-galactosidase construct. However, this particular mutant was replication defective in H9 cells; all other mutants were replication defective over the 8-week course of the assay. Two long terminal repeat viral DNA species could be detected in the CCCC mutant but not in any of the other replication-defective mutants. These studies show that the N-terminal Zn2+ finger position is more sensitive to alterations than the C-terminal position with respect to replication. Additionally, the retroviral (CCHC) NC Zn2+ finger is required for early infection processes. The evolutionary pressure to maintain CCHC NC Zn2+ fingers depends mainly on its function in infection processes, in addition to its function in genome packaging.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Amino Acid Substitution
  • Base Sequence
  • Cell Line
  • Conserved Sequence
  • DNA Primers
  • HIV Long Terminal Repeat
  • HIV-1 / genetics
  • HIV-1 / pathogenicity*
  • HIV-1 / physiology*
  • HeLa Cells
  • Humans
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Nucleocapsid / chemistry
  • Nucleocapsid / genetics
  • Nucleocapsid / metabolism*
  • Polymerase Chain Reaction
  • Transfection
  • Virus Replication
  • Zinc Fingers

Substances

  • DNA Primers