Transcriptomic analysis of the dialogue between Pseudorabies virus and porcine epithelial cells during infection.
BACKGROUND
Transcriptomic approaches are relevant for studying virus-host cell dialogues to better understand the physiopathology of infection and the immune response at the cellular level. Pseudorabies virus (PrV), a porcine Alphaherpesvirus, is a good model for such studies in pig. Since PrV displays a strong tropism for mucous epithelial cells, we developed a kinetics study of PrV infection in the porcine PK15 epithelial cell line. To identify as completely as possible, viral and cellular genes regulated during infection, we simultaneously analyzed PrV and cellular transcriptome modifications using two microarrays i.e. a laboratory-made combined SLA/PrV microarray, consisting of probes for all PrV genes and for porcine genes contained in the Swine Leukocyte Antigen (SLA) complex, and the porcine generic Qiagen-NRSP8 oligonucleotide microarray. We confirmed the differential expression of a selected set of genes by qRT-PCR and flow cytometry.


RESULTS
An increase in the number of differentially expressed cellular genes and PrV genes especially from 4 h post-infection (pi) was observed concomitantly with the onset of viral progeny while no early global cellular shutoff was recorded. Many cellular genes were down-regulated from 4 h pi and their number increased until 12 h pi. UL41 transcripts encoding the virion host shutoff protein were first detected as differentially expressed at 8 h pi. The viral gene UL49.5 encoding a TAP inhibitor protein was differentially expressed as soon as 2 h pi, indicating that viral evasion via TAP inhibition may start earlier than the cellular gene shutoff. We found that many biological processes are altered during PrV infection. Indeed, several genes involved in the SLA class I antigenic presentation pathway (SLA-Ia, TAP1, TAP2, PSMB8 and PSMB9), were down-regulated, thus contributing to viral immune escape from this pathway and other genes involved in apoptosis, nucleic acid metabolism, cytoskeleton signaling as well as interferon-mediated antiviral response were also modulated during PrV infection.


CONCLUSIONS
Our results show that the gene expression of both PrV and porcine cells can be analyzed simultaneously with microarrays, providing a chronology of PrV gene transcription, which has never been described before, and a global picture of transcription with a direct temporal link between viral and host gene expression.
Flori
Laurence
L
INRA, DGA, UMR 314, Laboratoire de Radiobiologie et d'Etude du Génome, Jouy-en-Josas, F-78350 France. laurence.flori@jouy.inra.fr
Rogel-Gaillard
Claire
C
Cochet
Marielle
M
Lemonnier
Gaetan
G
Hugot
Karine
K
Chardon
Patrick
P
Robin
Stéphane
S
Lefèvre
François
F
D000818:Animals; D017209:Apoptosis / Q000276:immunology; D004847:Epithelial Cells / Q000378:metabolism* / Q000821:virology*; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000276:immunology*; D011558:Herpesvirus 1, Suid; D008285:Major Histocompatibility Complex / Q000276:immunology; D020411:Oligonucleotide Array Sequence Analysis; D011557:Pseudorabies / Q000276:immunology / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013552:Swine; D013553:Swine Diseases / Q000276:immunology / Q000378:metabolism / Q000821:virology*