Role of the human T-cell leukemia virus type 1 PTAP motif in Gag targeting and particle release.
Human T-cell leukemia virus type 1 (HTLV-1) Gag is targeted to the plasma membrane for particle assembly and release. How HTLV-1 Gag targeting occurs is not well understood. The PPPY and PTAP motifs were previously shown to be involved in HTLV-1 particle release with PTAP playing a more subtle role in virus budding. These L domains function through the interaction with host cellular proteins normally involved in multivesicular body (MVB) morphogenesis. The plasma membrane pathway rather than the MVB pathway was found to be the primary pathway for HTLV-1 particle release in HeLa cells. Intriguingly, disruption of the PTAP motif led to a defect in the targeting of Gag from the plasma membrane to CD63-positive MVBs. Particles or particle buds were observed to be associated with MVBs by electron microscopy, implying that Gag targeting to the MVB resulted in particle budding. Blocking clathrin-dependent endocytosis was found not to influence localization of the HTLV-1 Gag PTAP mutant, indicating that Gag did not reach the MVBs through clathrin-dependent endocytosis. Our observations imply that the interaction between Gag and TSG101 is not required for Gag targeting to the MVB. Overexpression of dynamitin p50 increased particle release, suggesting that there was an increase in the intracellular transport of MVBs to the cell periphery by the utilization of the dynein-dynactin motor complex. Intriguingly, virus particle release with this mutant was reduced by 20-fold compared to that of wild type in HeLa cells, which is in marked contrast to the less-than-twofold defect observed for particle production of the HTLV-1 Gag PTAP mutant from 293T cells. These results indicate that the role of the PTAP motif in L domain function is cell type dependent.
Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA.
Irene J
IJ
Dorweiler
Susan J
SJ
Ruone
Huating
H
Wang
Richard W
RW
Burry
Louis M
LM
Mansky
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000378:metabolism; D015703:Antigens, CD / Q000378:metabolism / Q000648:ultrastructure; D001324:Autoantigens; D002462:Cell Membrane / Q000378:metabolism / Q000821:virology; D000072159:Dynactin Complex; D019084:Fluorescent Antibody Technique, Indirect; D015683:Gene Products, gag / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D005819:Genetic Markers; D006367:HeLa Cells; D015368:Human T-lymphotropic virus 1 / Q000737:chemistry / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D006801:Humans; D007150:Immunohistochemistry; D008565:Membrane Proteins / Q000378:metabolism / Q000648:ultrastructure; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009154:Mutation; D010957:Plasmids; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D014162:Transfection; D014168:Transferrin / Q000378:metabolism; D033921:Vesicular Transport Proteins; D014771:Virion / Q000502:physiology* / Q000648:ultrastructure; D019065:Virus Assembly
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D001324:Autoantigens; C000606361:DCTN2 protein, human; D000072159:Dynactin Complex; D015683:Gene Products, gag; D005819:Genetic Markers; C097741:Golgin subfamily A member 2; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; D014168:Transferrin; D033921:Vesicular Transport Proteins; C093846:early endosome antigen 1