Role Of Lipid Raft In Assembly Of Human Herpesvirus-6 And Effect On Virus Infection

Human Herpesvirus 6(HHV-6) is a neotype herpesvirus which wasfirstly isolated from the patients of AIDS and lymphoproliferativedisorders. It is very common for human infection the HHV-6, whichmostly are silent infection and virus might incubate in host for a long time.When human suffer the nonspecific stimulation and resistance attenuateand immune function descent, latent HHV-6 will be actived and becomethe lasting infection. About 60-80% children and adult serum can bedetected the HHV-6 antibody. HHV-6 can be classified two types:HHV-6A and HHV-6B by the virus DNA restriction enzyme analysis,nucleotide sequence analysis, the reaction to the monoclonal antibody andthe growth in different T lymph corpuscles. HHV-6B is the pathogen ofthe exanthema subitum (rose anthema).And it has been comfirmed relatedwith lymphoproliferating disease, auto-immune disease and immunologicdeficiency disease. With the development of the organ transplant and thegrowth number of the AIDS patients, HHV-6 infection becomes importantincreasingly. But the fitting process of virus in the susceptible cell andbudding releasing are still unclear.Recently, the tradition conception, which presumed the plasmamembrane as a homogeneous 2-dimensional liquid phase and membraneprotein uniformly distributed in it, underwent the severe challenge. Studyshowed that the distribution of lipid bimolecular leaflet in membrane isnot symmetry and aequalis. There are some high density, sequence andespecial local constructions at the plasma membrane. These membranemicro-constructions rich of sphingolipid and cholesterin with the abilityof recruited some protein selectively, and possess special chemicalcomposition and characteristic, so called them as lipid microdomains. Thecholesterin plays an important role in maintaining the lipid raftconstruction and its functional status. Respecting the micro-constructionsdistribution in membrane present as island, just like the raft floating in thesea, thus it is named as lipid raft. The raft floats in the derangementmatrix membrane lipid and is formed a lipid ordered phase. It takes partin some of the important biological process such as the transmembranesignal transduction, T cell activation, virus assembly, budding and so on.Majority of the enveloped virus budding releases from the hostmembrane and by means of it to obtain lipid bilayer from the cell;partialvirus buds from the endoplasmic reticulum, and mostly virus use the lipidraft construction during the budding. The lipid raft provids the platformfor the assembly and budding in human immunodeficiency virus (HIV-1).Moreover, the assembly and budding processes of the influenza virus andmeasles virus depend on the lipid raft. The main enveloped glycoproteinhemagglutinin (HA) and neuramidinase (NA) of the influenza virus arealso relative of the lipid raft construction.The enveloped virus depends on the lipid membrane infestation intothe susceptible cell. The process of enveloped virus infecting the host cellis the fusion process of the viral envelope and host lipid membrane, and itpromotes the virion invading in the host cell (the viral genetic materialenter in by the manner of neucleocapsid). The fusion process is mediatedby the glycoprotein in the virus surface recognizing and coupling thereciprocal specific receptors. The lipid raft plays an important role in thisprocedure. The fusion between SFV (Semliki forest virus) and cellularmembrane depends on the cholesterin (lipid raft) on the targeted cellularmembrane;the invading of HIV-1 and HSV (herpes simplex virus) alsoneed the lipid raft composition lying on the viral envelope and cellularmembrane;moreover, the lipid raft composition plays a crucial role infusion of influenza virus.HHV-6 invading in the host cell is a complex procedure, and itscombination between the viral envelope glycoprotein and the surfacereceptors on the susceptible cell is the first step of the viral adsorptionand initiated invading. CD46 is the receptor of the HHV-6, and HHV-6triggering the fusion depends on the CD46 expressing on the target cellsurface. The viral glycoprotein complex gH-gL-gQ1-gQ2 combines withthe CD46. Release the viral genome into the host cell to infect by the cellfusion.In order to investigate the effect of the lipid raft on the assembly inHHV-6 and lipid raft of the viral envelope on the HHV-6 invading thesusceptible cell, our researches are as follow:We used the HHV-6 GS strain to infect the HSB2 cell, extracted thelipid raft composition by using non-ion decontaminant Triton-X 100, andanalyzed the relationship between the HHV-6 envelope glycoprotein andlipid raft by Western Blot. The results showed that HHV-6 envelopeglycoprotein B,H,L,Q1 and Q2 (gB,gH,gL,gQ and gQ2 )distributedat the lipid raft site. In particularly, 80KDa gQ1(gQ1-80K)and 37KDagQ2(gQ-37K)protein, residing in the mature virion, also distributed inthe lipid raft. Being the non-structural protein, the immediate earlyprotein, IE1 distributed in the soluble the non-lipid raft composition.We researched the relationship of expression and distributionbetween the HHV-6 glycoprotein B(gB) and GPI anchored protein CD59and ganglioside GM1 by immunofluorescence double labeling.Weobserved that CD59 and HHV-6 envelope glycoprotein B had the samedistribution through the confocolmicroscope. In addition,we detected thatthe raft maker GM1 in purified HHV-6 paricle.To approach the effect of the raft of viral envelope on the viralinfectional susceptible cell, we used the mythyl-β-cyclodextrin (MβCD),which could removal the cholesterin in raft from the HHV-6, to decreasethe content of the cholesterin and with the concentration of the MβCDpresenting positive correlation. With the decreasting of the cholesterincontent, it significantly influenced the infectivity of the virus, and viralinfectivity presented the decreasing in a manner of the dose-depended.Meanwhile, virus infection could not cause cell fusion. Nevertherless,supplement of exogenous cholesterin could recover the viral infectivityand cell fusion.The virus, deleted cholesterin after treating with the MβCD, still hadthe ability of absorbing to the cell surface in target cell absorption assay,but it could not enter into cell interior. It showed that cholesterin in viralenvelope played a key role in virus entering into cell.The combination between tetramer complex gH-gL-gQ1-gQ2 andCD46 molecular played a crucial role in cell fusion. Treated virus withMβCD, complex gH-gL-gQ1-gQ2 might combine to the CD46 molecularbut the ability is lower. The cholesterin of HHV-6 envelope played acrucial role in the key step-cell fusion of virus invading into the cell.Our results showed that lipid raft provided the platform for HHV-6assembly and HHV-6 entering the cell depended on the rich cholesterin ofthe viral envelope, and was the results that virus selectively budded by therafts.