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Transmissible gastroenteritis virus (TGEV) infection alters the expression of cellular microRNA species that affect transcription of TGEV gene 7.

Song X, Zhao X, Huang Y, Xiang H, Zhang W, Tong D - Int. J. Biol. Sci. (2015)

Bottom Line: The results showed TGEV infection caused the change of miRNAs profile.Then we selected miR-4331 for further analysis and subsequently identified cell division cycle-associated protein 7 (CDCA7) as the target of miR-4331.Moreover, miR-4331 showed the ability to inhibit transcription of TGEV gene 7 (a non-structure gene) via directly targeting CDCA7.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, P.R. China.

ABSTRACT
Transmissible gastroenteritis virus (TGEV) is a member of Coronaviridae family. TGEV infection has emerged as a major cause of severe gastroenteritis and leads to alterations of many cellular processes. Meanwhile, the pathogenic mechanism of TGEV is still unclear. microRNAs (miRNAs) are a novel class of small non-coding RNAs which are involved in the regulation of numerous biological processes such as viral infection and cell apoptosis. Accumulating data show that miRNAs are involved in the process of coronavirus infection such as replication of severe acute respiratory syndrome coronavirus (SARS-CoV). However, the link between miRNAs and TGEV infection is unknown. In this study, we performed microRNA microarray assay and predicted targets of altered miRNAs. The results showed TGEV infection caused the change of miRNAs profile. Then we selected miR-4331 for further analysis and subsequently identified cell division cycle-associated protein 7 (CDCA7) as the target of miR-4331. Moreover, miR-4331 showed the ability to inhibit transcription of TGEV gene 7 (a non-structure gene) via directly targeting CDCA7. In conclusion, differentially expressed miR-4331 that is caused by TGEV infection can suppress transcription of TGEV gene 7 via targeting cellular CDCA7. Our key finding is that TGEV selectively manipulates the expression of some cellular miRNAs to regulate its subgenomic transcription.

No MeSH data available.


Related in: MedlinePlus

miRNAs expression profile in TGEV-infected PK-15 cells. (A) Heat map analysis showing differentially expressed miRNAs in PK-15 cells infected with TGEV at 24 h. Red indicates higher expression and green indicates lower expression. Each colored block represents the expression of one miRNA (labeled on the right) in the indicated sample. The map showed all significantly differential expressed miRNAs in three independent samples (p < 0.01). (B) Verification of miRNAs microarray by real-time PCR. The fold change was determined using the 2-ΔΔCt method and all miRNAs expression values were normalized to endogenous U6. Data from real-time PCR were shown as mean ±standard deviation (S.D.) of three independent experiments. Similar results were obtained in three independent experiments.
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Figure 1: miRNAs expression profile in TGEV-infected PK-15 cells. (A) Heat map analysis showing differentially expressed miRNAs in PK-15 cells infected with TGEV at 24 h. Red indicates higher expression and green indicates lower expression. Each colored block represents the expression of one miRNA (labeled on the right) in the indicated sample. The map showed all significantly differential expressed miRNAs in three independent samples (p < 0.01). (B) Verification of miRNAs microarray by real-time PCR. The fold change was determined using the 2-ΔΔCt method and all miRNAs expression values were normalized to endogenous U6. Data from real-time PCR were shown as mean ±standard deviation (S.D.) of three independent experiments. Similar results were obtained in three independent experiments.

Mentions: To gain the overview of the impact of TGEV infection on host cellular miRNAs expression, PK-15 cells were infected with TGEV Shaanxi strain at a multiplicity of infection (MOI) of 1.0 for microarray. The microarray was performed with the total RNA extracted from PK-15 cells infected with TGEV at 24 hpi. The differential expression of multiple miRNAs in TGEV-infected cells in comparison with mock-infected cells was observed in Fig. 1(A). The expression levels of 21 miRNAs were changed remarkably (fold change > 1.5, and p < 0.01). Among them 13 miRNAs were up-regulated and 8 miRNAs were down-regulated. To validate the microarray results of differentially expressed miRNAs, we tested the expression levels of them using real-time PCR. The fold changes of 21 miRNAs in TGEV-infected cells were referred to that in mock-infected cells. The miRNAs levels were normalized to U6. The results were correlated with microarray (Fig. 1B). A total of 21 differentially expressed miRNAs were chosen for target prediction. The potential targets of 21 differentially expressed miRNAs were predicted with TargetScan and miRanda and 7,844 potential target genes were gained (Supplementary Table S1).


Transmissible gastroenteritis virus (TGEV) infection alters the expression of cellular microRNA species that affect transcription of TGEV gene 7.

Song X, Zhao X, Huang Y, Xiang H, Zhang W, Tong D - Int. J. Biol. Sci. (2015)

miRNAs expression profile in TGEV-infected PK-15 cells. (A) Heat map analysis showing differentially expressed miRNAs in PK-15 cells infected with TGEV at 24 h. Red indicates higher expression and green indicates lower expression. Each colored block represents the expression of one miRNA (labeled on the right) in the indicated sample. The map showed all significantly differential expressed miRNAs in three independent samples (p < 0.01). (B) Verification of miRNAs microarray by real-time PCR. The fold change was determined using the 2-ΔΔCt method and all miRNAs expression values were normalized to endogenous U6. Data from real-time PCR were shown as mean ±standard deviation (S.D.) of three independent experiments. Similar results were obtained in three independent experiments.
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Related In: Results  -  Collection

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Figure 1: miRNAs expression profile in TGEV-infected PK-15 cells. (A) Heat map analysis showing differentially expressed miRNAs in PK-15 cells infected with TGEV at 24 h. Red indicates higher expression and green indicates lower expression. Each colored block represents the expression of one miRNA (labeled on the right) in the indicated sample. The map showed all significantly differential expressed miRNAs in three independent samples (p < 0.01). (B) Verification of miRNAs microarray by real-time PCR. The fold change was determined using the 2-ΔΔCt method and all miRNAs expression values were normalized to endogenous U6. Data from real-time PCR were shown as mean ±standard deviation (S.D.) of three independent experiments. Similar results were obtained in three independent experiments.
Mentions: To gain the overview of the impact of TGEV infection on host cellular miRNAs expression, PK-15 cells were infected with TGEV Shaanxi strain at a multiplicity of infection (MOI) of 1.0 for microarray. The microarray was performed with the total RNA extracted from PK-15 cells infected with TGEV at 24 hpi. The differential expression of multiple miRNAs in TGEV-infected cells in comparison with mock-infected cells was observed in Fig. 1(A). The expression levels of 21 miRNAs were changed remarkably (fold change > 1.5, and p < 0.01). Among them 13 miRNAs were up-regulated and 8 miRNAs were down-regulated. To validate the microarray results of differentially expressed miRNAs, we tested the expression levels of them using real-time PCR. The fold changes of 21 miRNAs in TGEV-infected cells were referred to that in mock-infected cells. The miRNAs levels were normalized to U6. The results were correlated with microarray (Fig. 1B). A total of 21 differentially expressed miRNAs were chosen for target prediction. The potential targets of 21 differentially expressed miRNAs were predicted with TargetScan and miRanda and 7,844 potential target genes were gained (Supplementary Table S1).

Bottom Line: The results showed TGEV infection caused the change of miRNAs profile.Then we selected miR-4331 for further analysis and subsequently identified cell division cycle-associated protein 7 (CDCA7) as the target of miR-4331.Moreover, miR-4331 showed the ability to inhibit transcription of TGEV gene 7 (a non-structure gene) via directly targeting CDCA7.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, P.R. China.

ABSTRACT
Transmissible gastroenteritis virus (TGEV) is a member of Coronaviridae family. TGEV infection has emerged as a major cause of severe gastroenteritis and leads to alterations of many cellular processes. Meanwhile, the pathogenic mechanism of TGEV is still unclear. microRNAs (miRNAs) are a novel class of small non-coding RNAs which are involved in the regulation of numerous biological processes such as viral infection and cell apoptosis. Accumulating data show that miRNAs are involved in the process of coronavirus infection such as replication of severe acute respiratory syndrome coronavirus (SARS-CoV). However, the link between miRNAs and TGEV infection is unknown. In this study, we performed microRNA microarray assay and predicted targets of altered miRNAs. The results showed TGEV infection caused the change of miRNAs profile. Then we selected miR-4331 for further analysis and subsequently identified cell division cycle-associated protein 7 (CDCA7) as the target of miR-4331. Moreover, miR-4331 showed the ability to inhibit transcription of TGEV gene 7 (a non-structure gene) via directly targeting CDCA7. In conclusion, differentially expressed miR-4331 that is caused by TGEV infection can suppress transcription of TGEV gene 7 via targeting cellular CDCA7. Our key finding is that TGEV selectively manipulates the expression of some cellular miRNAs to regulate its subgenomic transcription.

No MeSH data available.


Related in: MedlinePlus