Project description:Transmissible gastroenteritis virus (TGEV; Coronaviridae family) causes huge economic losses to the swine industry. MicroRNAs (miRNAs) play a regulatory role in viral infection and may be involved in the mammalian immune response. Here, we report a comprehensive analysis of host miRNA expression in TGEV-infected swine testis (ST) cells. Deep sequencing generated 3,704,353 and 2,763,665 reads from uninfected ST cells and infected ST cells, respectively. The reads were aligned to known Sus scrofa pre-miRNAs in miRBase 19, identifying 284 annotated miRNAs. Certain miRNAs were differentially regulated during TGEV infection, which confirmed the hypothesis that specific miRNAs play a regulatory role in virus-host interactions. 59 unique miRNAs displayed significant differentially expression between the normal and TGEV-infected ST cell samples: 15 miRNAs were significantly up-regulated and 44 were significantly down-regulated. Stem-loop RT-PCR was carried out to determine the expression levels of specific miRNAs in the two samples, and the results were consistent with those of sequencing. Gene ontology enrichment analysis of host target genes demonstrated that the differentially expressed miRNAs are involved in regulatory networks, including cellular process, metabolic process, immune system process. This is the first report of the identification of ST cell miRNAs and the comprehensive analysis of the miRNA regulatory mechanism during TGEV infection, which revealed the miRNA molecular regulatory mechanisms for the viral infection, expression of viral genes and the expression of immune-related genes. The results presented here will aid research on the prevention and treatment of viral diseases. 2 ST(Porcine testicular cells) cell samples, ST: normal ST cell sample (contro sample), TGEV: TGEV (Transmissible gastroenteritis virus) infected ST cell samples
Project description:Transmissible gastroenteritis virus (TGEV; Coronaviridae family) causes huge economic losses to the swine industry. MicroRNAs (miRNAs) play a regulatory role in viral infection and may be involved in the mammalian immune response. Here, we report a comprehensive analysis of host miRNA expression in TGEV-infected swine testis (ST) cells. Deep sequencing generated 3,704,353 and 2,763,665 reads from uninfected ST cells and infected ST cells, respectively. The reads were aligned to known Sus scrofa pre-miRNAs in miRBase 19, identifying 284 annotated miRNAs. Certain miRNAs were differentially regulated during TGEV infection, which confirmed the hypothesis that specific miRNAs play a regulatory role in virus-host interactions. 59 unique miRNAs displayed significant differentially expression between the normal and TGEV-infected ST cell samples: 15 miRNAs were significantly up-regulated and 44 were significantly down-regulated. Stem-loop RT-PCR was carried out to determine the expression levels of specific miRNAs in the two samples, and the results were consistent with those of sequencing. Gene ontology enrichment analysis of host target genes demonstrated that the differentially expressed miRNAs are involved in regulatory networks, including cellular process, metabolic process, immune system process. This is the first report of the identification of ST cell miRNAs and the comprehensive analysis of the miRNA regulatory mechanism during TGEV infection, which revealed the miRNA molecular regulatory mechanisms for the viral infection, expression of viral genes and the expression of immune-related genes. The results presented here will aid research on the prevention and treatment of viral diseases.
Project description:Transmissible gastroenteritis virus (TGEV), a member of coronavirus, is the pathogen of TGE. We previously found 123 circular RNAs (circRNAs) were differential expression during activation of mitochondrial permeability transition in porcine intestinal epithelial cells-jejunum 2 cell line (IPEC-J2) in response to TGEV infection. Mitochondrial permeability transition pore (mPTP) is a transmembrane pore of mitochondria and plays a key role in MPT. mPTP abnormal opening causes MPT. Therefore, we postulated that circRNAs might be related to mPTP abnormal opening induced by TGEV. In this study, we found that circBIRC6-2 could inhibit the mPTP abnormal opening induced by TGEV. Interestingly, circBIRC6-2 encodes protein BIRC6-236aa. An open reading frame (ORF) and internal ribosomal entrance site (IRES) of circBIRC6-2 were identified. We also found that BIRC6-236aa rather than circBIRC6-2 inhibited the mPTP opening by overexpression of circBIRC6-2 related vectors. We obtained 91 proteins that interacted with BIRC6-236aa using immunoprecipitation-mass spectrometry (IP-MS). The interaction between voltage-dependent anion-selective channel protein 1 (VDAC1) and BIRC6-236aa was demonstrated through co-immunoprecipitation (Co-IP). Moreover, BIRC6-236aa could inhibit the formation of VDAC1 and Cyclophilin D (CypD) complexes. Overall, these results indicated that BIRC6-236aa antagonized mPTP opening by interacting with VDAC1 to weaken the extent of interaction between VDAC1 and CypD.
Project description:Transmissible gastroenteritis virus (TGEV) is a member of Coronaviridae family. Our previous research showed that TGEV infection could induce mitochondrial dysfunction and up-regulat miR-222 level. Therefore, we presumed that miR-222 might be implicated in regulating mitochondrial dysfunction induced by TGEV infection. To verify the hypothesis, the effect of miR-222 on mitochondrial dysfunction was detected and showed that miR-222 attenuated TGEV-induced mitochondrial dysfunction. To investigate the underlying molecular mechanism of miR-222 in TGEV-induced mitochondrial dysfunction, a quantitative proteomic analysis of PK-15 cells that were transfected with miR-222 mimics and infected with TGEV was performed. In total, 4151 proteins were quantified and 100 differentially expressed proteins were obtained (57 up-regulated, 43 down-regulated), among which thrombospondin-1 (THBS1) and cluster of differentiation 47 (CD47) were down-regulated. THBS1 was identified as the target of miR-222. Knockdown of THBS1 and CD47 increased mitochondrial Ca2+ level and decreased mitochondrial membrane potential (MMP) level. Together, our data establish a significant role of miR-222 in regulating mitochondrial dysfunction in response to TGEV infection.
Project description:Transmissible gastroenteritis virus (TGEV) is an infective coronavirus (CoV) that causes diarrhea-related morbidity and mortality in piglets. For the first time, a natural recombination strain of a TGEV Anhui Hefei (AHHF) virus between the Purdue and the Miller clusters was isolated from the small intestine content of piglets in China. A phylogenetic tree based on a complete genome sequence placed the TGEV AHHF strain between the Purdue and the Miller clusters. The results of a computational analysis of recombination showed that the TGEV AHHF strain is a natural recombinant strain between these clusters. Two breakpoints located in the open reading frame 1a (ORF1a) and spike (S) genes were identified. The pathogenicity of the TGEV AHHF strain was evaluated in piglets, and the results show that TGEV AHHF is an enteric pathogenic strain. These results provide valuable information about the recombination and evolution of CoVs and will facilitate future investigations of the molecular pathogenesis of TGEV.
Project description:Transmissible gastroenteritis virus strain AYU was isolated in Shanghai. The complete genome has a length of 28,582 bp and contains seven open reading frames. Sequence analysis suggested that Shanghai strain AYU and U.S. strain Purdue P115 are derived from a common ancestor, as they have 99.6% similarity at the nucleotide level.