A membrane leucine heptad contributes to trafficking, signaling, and transformation by latent membrane protein 1.
Latent membrane protein 1 (LMP1) of Epstein Barr virus (EBV) is important for maintaining proliferation of EBV-infected B cells. LMP1, unlike its cellular counterpart, CD40, signals without a ligand and is largely internal to the plasma membrane. In order to understand how LMP1 initiates its ligand-independent signaling, we focused on a leucine heptad in LMP1's first membrane-spanning domain that was shown to be necessary for LMP1's signaling through NF-kappaB. LZ1EBV, a recombinant EBV genetically altered to express LZ1, a derivative of LMP1 in which a leucine heptad was replaced with alanines, transformed B cells with 56% of wild-type (wt) EBV's efficiency, demonstrating the importance of this heptad. To elucidate the mechanism by which this domain contributes to the functions of LMP1, the properties of the wt and LZ1 were compared in transfected cells. LZ1 failed to home to lipid rafts as efficiently as did wt LMP1. The distribution of tagged derivatives of LZ1 also differed from that of wt LMP1 in transfected cells. LZ1's defect in homing to lipid rafts and altered trafficking likely underlie the defect in transformation of LZ1EBV. While the third and fourth membrane-spanning domains of LMP1 foster its trafficking to the Golgi, the leucine heptad within the first membrane-spanning domain contributes to its trafficking, particularly to internal rafts. B cells that are successfully transformed by LZ1EBV have the same average number of viral genomes and the same fraction of cells with capped LZ1 at the cell surface but express 50% more of the LZ1 allele than wt infected cells.
Lee
Jisook
J
McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, WI 53706, USA.
Sugden
Bill
B
D019943:Amino Acid Substitution; D001402:B-Lymphocytes / Q000821:virology; D002460:Cell Line; D002472:Cell Transformation, Viral; D004279:DNA, Viral / Q000032:analysis; D004854:Herpesvirus 4, Human / Q000502:physiology*; D006801:Humans; D021962:Membrane Microdomains / Q000737:chemistry; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D016297:Mutagenesis, Site-Directed; D016328:NF-kappa B / Q000032:analysis; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D015398:Signal Transduction; D014763:Viral Matrix Proteins / Q000235:genetics* / Q000378:metabolism*
D004279:DNA, Viral; C037662:EBV-associated membrane antigen, Epstein-Barr virus; D016328:NF-kappa B; D014763:Viral Matrix Proteins