Identification of cellular proteome using two-dimensional difference gel electrophoresis in ST cells infected with transmissible gastroenteritis coronavirus

Xin Zhang,Zhong-Tian Wang,Da Shi,Hong-Wu Lang,Jian-Fei Chen,Hong-Yan Shi,Li Feng

doi:10.1186/1477-5956-11-31

Abstract

BackgroundTransmissible gastroenteritis coronavirus (TGEV) is an enteropathogenic coronavirus that causes diarrhea in pigs, which is correlated with high morbidity and mortality in suckling piglets. Information remains limited about the comparative protein expression of host cells in response to TGEV infection. In this study, cellular protein response to TGEV infection in swine testes (ST) cells was analyzed, using the proteomic method of two-dimensional difference gel electrophoresis (2D DIGE) coupled with MALDI-TOF-TOF/MS identification.Results33 differentially expressed protein spots, of which 23 were up-regulated and 10 were down-regulated were identified. All the protein spots were successfully identified. The identified proteins were involved in the regulation of essential processes such as cellular structure and integrity, RNA processing, protein biosynthesis and modification, vesicle transport, signal transduction, and the mitochondrial pathway. Western blot analysis was used to validate the changes of alpha tubulin, keratin 19, and prohibitin during TGEV infection.ConclusionsTo our knowledge, we have performed the first analysis of the proteomic changes in host cell during TGEV infection. 17 altered cellular proteins that differentially expressed in TGEV infection were identified. The present study provides protein-related information that should be useful for understanding the host cell response to TGEV infection and the underlying mechanism of TGEV replication and pathogenicity.

Highlights

Transmissible gastroenteritis coronavirus (TGEV) is an enteropathogenic coronavirus that causes diarrhea in pigs, which is correlated with high morbidity and mortality in suckling piglets
Confirmation of TGEV propagation in swine testes (ST) cells To obtain a detailed comparison of differences in protein expression, the cellular proteins were extracted at 24 h, 48 h and 72 h p.i. from the TGEV-infected and mockinfected ST cells and were identified by IFA and Western blot analysis using monoclonal antibody (mAb) to N protein of TGEV as primary antibody
IFA and Western blot analysis revealed that the ST cells infected with TGEV could be recognized with mAb to N protein of the TGEV at 24 h, 48 h and 72 h p.i. (Figure 1)

Summary

Introduction

Transmissible gastroenteritis coronavirus (TGEV) is an enteropathogenic coronavirus that causes diarrhea in pigs, which is correlated with high morbidity and mortality in suckling piglets. Information remains limited about the comparative protein expression of host cells in response to TGEV infection. TGEV is an enveloped virus with a positivesense RNA genome of 28.5 kb. The genome of TGEV encodes four structural proteins: spike (S), membrane (M,), minor envelope (E), and nucleocapsid (N) proteins. The N protein, an internal phosphoprotein [8], interacts with the TGEV genomic RNA to form viral nucleocapsid [5,9], and may disrupt host cell division [10]. There is limited information about host cell responses to TGEV infection

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