title: Inhibition of porcine transmissible gastroenteritis virus infection in porcine kidney cells using short hairpin RNAs targeting the membrane gene.
abstract: The membrane (M) protein is the most abundant component of the porcine transmissible gastroenteritis virus (TGEV) particle. To exploit the possibility of using RNA interference (RNAi) as a strategy against TGEV infection, three plasmids (pRNAT-1, pRNAT-2, and pRNAT-3) expressing short hairpin RNAs were designed to target three different coding regions of the M gene of TGEV. The plasmids were constructed and transiently transfected into a porcine kidney cells, PK-15, to determine whether these constructs inhibited TGEV production. The analysis of cytopathic effects demonstrated that pRNAT-2 and pRNAT-3 could protect PK-15 cells against pathological changes specifically and efficiently. Additionally, indirect immunofluorescence and 50% tissue culture infectious dose (TCID50) assays showed that pRNAT-2 and pRNAT-3 inhibited the multiplication of the virus at the protein level effectively. Quantitative real-time PCR further confirmed that the amounts of viral RNAs in cell cultures pre-transfected with the three plasmids were reduced by 13, 68, and 70%, respectively. This is the first report showing that RNAi targeting of the M gene. Our results could promote studies of the specific function of viral genes associated with TGEV infection and might provide a theoretical basis for potential therapeutic applications.
authors.affiliation: Department of Veterinary Medicine, Southwest University, Rongchang Campus, Chongqing, 402460, People's Republic of China.
authors.forename: Li
authors.initials: L
authors.lastname: Wang
authors.affiliation: Department of Veterinary Medicine, Southwest University, Rongchang Campus, Chongqing, 402460, People's Republic of China.
authors.forename: Xianjin
authors.initials: X
authors.lastname: Dai
authors.affiliation: Department of Veterinary Medicine, Southwest University, Rongchang Campus, Chongqing, 402460, People's Republic of China.
authors.forename: Han
authors.initials: H
authors.lastname: Song
authors.affiliation: Department of Veterinary Medicine, Southwest University, Rongchang Campus, Chongqing, 402460, People's Republic of China.
authors.forename: Peng
authors.initials: P
authors.lastname: Yuan
authors.affiliation: Department of Veterinary Medicine, Southwest University, Rongchang Campus, Chongqing, 402460, People's Republic of China.
authors.forename: Zhou
authors.initials: Z
authors.lastname: Yang
authors.affiliation: Department of Veterinary Medicine, Southwest University, Rongchang Campus, Chongqing, 402460, People's Republic of China.
authors.forename: Wei
authors.initials: W
authors.lastname: Dong
authors.affiliation: Department of Veterinary Medicine, Southwest University, Rongchang Campus, Chongqing, 402460, People's Republic of China. szh7678@126.com.
authors.forename: Zhenhui
authors.initials: Z
authors.lastname: Song
mesh_terms: D000818:Animals; D005761:Gastroenteritis, Transmissible, of Swine / Q000235:genetics* / Q000628:therapy / Q000821:virology; D007668:Kidney / Q000473:pathology / Q000821:virology; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000235:genetics*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D013552:Swine / Q000235:genetics / Q000821:virology; D005760:Transmissible gastroenteritis virus / Q000235:genetics* / Q000472:pathogenicity; D014779:Virus Replication / Q000235:genetics
chemical_list: D008565:Membrane Proteins; D034741:RNA, Small Interfering
keywords: Membrane gene; Porcine kidney cell; RNA interference; Short hairpin RNA; Transmissible gastroenteritis virus