Systems Biology Investigation On The Evolutional Network Of Protein Molecules In The Pathogenesis Of Piglets Experimentally Infected With HuN4HP-PRRSV

HP-PRRS has brought enormous economic loss for the swine industry in China.However, the pathogenesis mechanism of which was still not clear. The present studyanalyzed protein profile changes of lungs, mandibular lymph nodes and serums from pigletsinfected with HuN4stain of HP-PRRSV at different time points, aimed to screen dynamicchanges in the process of HP-PRRS development, to look for and identify proteins or peptidesrelated to the virus infection and pathogenesis, and to analyse the differentially expressedproteins (DEPs) using bioinformatics methods. The above work would help to understandpathogenesis mechanism and develop new vaccines/drugs and breeding methods for diseaseresistance.We analyzed proteins from lungs, the target organ of PRRSV, from piglets inoculatedwith HuN4at6time points by means of2DE and mass spectrometry.20DEPs were foundand17were successfully identified with MALDI-TOF/TOF, which stand for14geneproducts. These DEPs were functionally classified into cytoskeleton, redox enzymes,biosynthesis, cell metabolism, adipose differentiation-related protein, albumin, immuneresponse, posttranslational modification and transcription modulating proteins. All of thecytoskeletons and metabolism enzymes were down-regulated; indicating that normalfunction of pig lungs was seriously affected. Bioinformatics analysis showed that Q31068(MHC PD14transplantation antigen) played central function in protein-protein interaction(PPI) network of pig lungs. Cytoskeletal proteins Q2XQY5and B5APU3and tumorsuppressor B8XSI6were all node proteins in the PPI network of pig lungs. Variable levels ofthese DEPs offered clues to understand respiratory diseases caused by PRRSV.Lymph nodes are organs where immune response occurs. Mandibular lymph nodes andmesenteric lymph nodes from piglets inoculated with HuN4showed swelling andhemorrhages. We compared the mandibular lymph nodes from piglets inoculated with HuN4for1d,3d and7d with those from healthy controls. Among22DEPs,20of which stand for19protein products were successfully identified. Three cytoskeletons (plasma gelsolinprecursor, actin related protein2/3complex subunit3and actin-related protein2-like protein)were all down-regulated, indicating that the structure and function of lymphcytes wereseriously harmed. Three of the HSP family members HSP60, HSP70and gp96were all up-regulated, indicating that strong immune responses were induced in lymph organs afterHuN4infection. In addition, many DEPs involving in transcriptional modulation andpost-translational modification played important roles in regulating gene expression. ProteinMx1with broad anti-virus activities was down-regulated. Five sera proteins were alsoinvolved in DEPs, of which P01965(Porcine Hemoglobin) indicated by bioinformaticsanalysis played central role in PPI of pig mandibular lymph nodes. The dynamic changes ofthe above proteins not only showed serious damage and lesion of lymph nodes, but alsooffered clues for immune response research on HP-PRRSV infection.We selected a farrow of piglets in order to reduce the genetic background ofexperimental animals. Serums were obtained at1,3,7and14days post inoculation of HuN4.2D-DIGE and mass spectrometry were used to analyze protein changes in serums. The resultsindicated that HuN4infection induced increasing complement C1s subunit,α1-antichimotrypsin family members, haptoglobin and decreased apolipoprotein. Most ofthese varied proteins belong to acute phase proteins (APPs), which play broad roles inactivities related to immunity and anti-inflammation. Bioinformatics analysis indicated thatcomplement proteins and haptoglobin have played central roles in serum PPI. The aboveresults of protein changes of host serum after infection offered useful information to observeserum changes in PRRSV infection.The present study is the most comprehensive understanding of systems biological studyon PRRSV infected piglets so far, which offered protein profile changes of lungs, mandibularlymph nodes and serums from piglets inoculated with HuN4stain of HP-PRRSV at differenttime points, and investigated the functions of DEPs. Many node proteins were found in thePPI network constructed for all DEPs from the lungs, mandibular lymph nodes and serums,among which MHC transplantation antigen and complement play core roles in the immuneregulation; Haptoglobin, Transferrin and Hemoglobin might affect the energy system throughmodulating the oxygen and iron balance in the body; Cytoskeletal related proteins areinvolved in the cell dynamics regulation, and are intimately related to the integrity of cellstructure, materials transportation and energy system; Transcription regulating proteins areinvolved in controlling synthesis of many genes. These node proteins that possess multiplefunctions of modulation have offered entrance to understand the PPI during the pathogenesisof piglets infected with HuN4.