Analyses Of The Feather Pulp Proteome Of Chickens Infected With Marek's Disease Virus

Gallid herpesvirus 2(GaHV-2),commonly known as Marek's disease virusserotype-1(MDV-1),causes T-cell lymphomas, immunosuppression and neurological disorder inchickens. MDV-1 is classified in the genus Mardivirus of the subfamily Alphaherpesvirinae along withtwo other non-oncogenic poultry viruses,Gallid herpesvirus 3(GaHV-3 or MDV serotype 2)andMelea-grid herpesvirus 1(MeHV-1,MDV serotype 3 or Turkey herpesvirus). This vaccine preventscancer in chickens that become infected with MDV, making it the first vaccine used to prevent cancer inany animal.Of the tissues that become infected with MDV,only skin and the associated feather follicleepithelium (FFE) permit MDV envelope formation leading to infectious virus formation.Envelopeformation and shedding of infectious MDV from feathers are critical steps for transmission of thevirus.Importantly,althoμgh the currently used MD vaccines are effective against development of clinicalsigns of disease,they are not capable of significantly reducing MDV replication and genomeaccumulation in feathers and also virus shedding to the environment..For this reason,the feather pulp isimportant in transmission of MDV and,hence,epidemiology of MD. But some study before is knownfrom DNA or mRNA level.In this paper,we identified differentially expressed proteins in the featherpulp of chickens infected with the virulent MDV (vMDV) GA strain and vaccine strain CVI988,usingtwo-dimensional gel electrophoresis on samples from protein level. The present findings provide a basisfor investigating production of enveloped fully infectious virus and pathogenesis of MDV.1. Development of two-dimensional gel electrophoresis for proteomic of the feather pulp.The first dimension was isoelectric focusing in immobilized pH gradients(IPG)strip and the seconddimension was SDS-PAGE on vertical electrophoresis units.The conditions were optimized,such asisoelectric focusing, different pH range IPG, proteins loaded, staining,and other steps.The results wereanalyzed with the software ImageMaster 2D Platinum version 6.0.On the 13cm IPG strip(pH3–10),more than 700 spots were detected in the feather pulp tissue throμgh this method,and the matchrate of protein spots was 80%. as well as develop two-dimensional electrophoresis(2-DE) with goodrepetition and high resolution for the proteomic analysis of the feather pulp,in this study,develope a2-DE method for proteomic of the feather pulp.2. Analyses of the feather pulp proteome of chickens infected with virulent Marek's disease virusIn the present study,changes in the FFE proteome at 4,7,14 and 21 days post-infection in responseto MDV GA strain infection were studied using two-dimensional gel electrophoresis and subsequentlythe protein spots were identified by MALDI-TOF/TOF-MS/MS.In total,29 host proteins were detectedas either quantitatively or qualitatively differentially expressed at least once during different samplingpoints.Database queries confirmed that these proteins were mainly associated with signaltransduction,immunology,cell proliferation and apoptosis,formation of the cytoskeleton,cellularmetabolism,signal transduction and regulation of translation.The present findings provide a basis for investigating production of enveloped fully infectious virus and pathogenesis of MDV.3. Analyses of the feather pulp proteome of chickens infected with differently virulent Marek'sdisease virusThe present study was designed to study comparative changes in the feather pulp proteomes of thedifferent virulent MDV infection.The the feather pulp infected with GA strain andvaccine strain CVI988 proteomes were examined at 4,7,14 and 21 days post-infection (d.p.i.) usingtwo-dimensional gel electrophoresis and subsequently the protein spots were identified byMALDI-TOF/TOF-MS/MS. There was no spot which differed quantitatively in their expressionbetween infected with GA strain and vaccine strain CVI988 at 4 d.p.i.and individually 4,6,5 spots eachat the rest of the time points. The differentially expressed proteins identified in the present study areLASP-1,heterogeneous nuclear ribonucleoprotein D-like,triosephosphate isomerase, L-lactatedehydrogenase and S-methyl-5'-thioadenosine phosphorylase.These findings shed light on some of theunderlying processes of different virulent virus pathogenic mechanism to MD.