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Characterization of the proline-rich region of murine leukemia virus envelope protein
B Weimin Wu1, P M Cannon, E M Gordon
1Gene Therapy Laboratories, Norris Cancer Center, University of Southern California School of Medicine, Los Angeles, California 90033, USA.
Abstract:
Mammalian type C retroviral envelope proteins contain a variable proline-rich region (PRR), located between the N-terminal receptor-binding domain and the more highly conserved C-terminal portion of the surface (SU) subunit. We have investigated the role of the PRR in the function of murine leukemia virus (MuLV) envelope protein. In the MuLVs, the PRR contains a highly conserved N-terminal sequence and a hypervariable C-terminal sequence. Despite this variability, the amphotropic PRR could functionally substitute for the ecotropic PRR. The hypervariable region of the PRR was not absolutely required for envelope protein function. However, truncations in this region resulted in decreased levels of both the SU and TM proteins in viral particles and increased amounts of the uncleaved precursor protein, Pr85. In contrast, the N-terminal conserved region was essential for viral infectivity. Deletion of this region prevented the stable incorporation of envelope proteins into viral particles in spite of normal envelope protein processing, wild-type levels of cell surface expression, and a wild-type ability to induce syncytia in an XC cell cocultivation assay. However, higher levels of the SU protein were shed into the supernatant, suggesting a defect in SU-TM interactions. Our data are most consistent with a role for the PRR in stabilizing the overall structure of the protein, thereby affecting the proper processing of Pr85, SU-TM interactions, and the stable incorporation of envelope proteins into viral particles. In addition, we have demonstrated that the PRR can tolerate the insertion of a peptide-binding domain, making this a potentially useful site for constructing targetable retroviral vectors.
Insights
The proline-rich region (PRR) in murine leukemia virus (MuLV) envelope proteins is crucial for viral infectivity by stabilizing protein structure and ensuring proper processing and incorporation. This region can be engineered for targeted retroviral vectors.
Area of Science:
- Molecular Biology
- Virology
- Protein Engineering
Background:
- Mammalian type C retroviral envelope proteins feature a variable proline-rich region (PRR) between the receptor-binding domain and the surface (SU) subunit.
- The PRR's role in murine leukemia virus (MuLV) envelope protein function was investigated, noting its conserved N-terminal and hypervariable C-terminal sequences.
Purpose of the Study:
- To elucidate the specific functions of the proline-rich region (PRR) in murine leukemia virus (MuLV) envelope protein.
- To determine the necessity of the PRR's conserved and hypervariable regions for viral infectivity and protein stability.
Main Methods:
- Functional analysis of MuLV envelope protein mutants with deletions or alterations in the PRR.
- Assessment of protein processing, cell surface expression, syncytia induction, and viral particle incorporation.
- Investigation of SU-TM interactions and protein shedding.
Main Results:
- The conserved N-terminal region of the PRR is essential for viral infectivity, with its deletion preventing stable envelope protein incorporation.
- Truncations in the hypervariable C-terminal region reduced SU and TM protein levels in viral particles and increased uncleaved precursor protein (Pr85).
- Deletion of the N-terminal PRR region led to increased SU shedding, indicating a defect in SU-TM interactions.
Conclusions:
- The PRR plays a critical role in stabilizing the overall structure of MuLV envelope proteins, influencing Pr85 processing, SU-TM interactions, and stable incorporation into viral particles.
- The PRR is essential for maintaining viral infectivity, despite normal processing and cell surface expression when its conserved N-terminal region is deleted.
- The PRR's ability to tolerate peptide-binding domain insertion offers potential for developing targeted retroviral vectors.