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. 2019 Jun 5;14(6):e0217438.
doi: 10.1371/journal.pone.0217438. eCollection 2019.

RNA-seq Profiles of Chicken Type II Pneumocyte in Response to Escherichia Coli Infection

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Free PMC article

RNA-seq Profiles of Chicken Type II Pneumocyte in Response to Escherichia Coli Infection

Lu-Yuan Peng et al. PLoS One. .
Free PMC article

Abstract

Avian pathogenic Escherichia coli (APEC) causes great economic loss to the poultry industry worldwide. Chicken type II pneumocytes (CP II cells) secrete surfactants and modulate lung immunity to decrease the infection of the invading pathogen. Nevertheless, the pathogenesis of CP II cells to APEC infection remains poorly understood. Therefore, we conducted global gene expression profiling of CP II cells after APEC-O78 infection to explore the host-pathogen interaction. The differentially expressed genes of CP II cells to APEC infection were characterized by RNA-seq with EB-seq algorithm. In consequence, the mRNA of 18996 genes was identified, and CP II cells responded to APEC infection with marked changes in the expression of 1390 genes. Among them, there are 803 down-regulated mRNAs and 587 up-regulated mRNAs. The KEGG prediction and Gene Ontology terms analysis revealed that the major enriched pathways were related to NF-κB signaling pathway, apoptosis pathway, tight junction, and cytokine-cytokine receptor interaction and other pathways. We adopted qRT-PCR to verify the validity of the selected gene expression. The fold induction of qPCR was similar to the RNA-seq results. These results provide a better understanding of the pathogenesis of APEC, especially apoptosis pathway involved in APEC infection.

Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Volcano plot of differentially expressed genes of infected cells (APEC-O78 infection) and non-infected cells (Control).
Red dots represent up-regulated genes, green, blue and black dots represent non-differentially expressed genes (Fold Change>1.5 or Fold Change<0.667, FDR<0.05). The horizontal axis represents the fold change between infected and control cells. The vertical axis represents the FDR‐value of the multiple‐test for the differences between samples.
Fig 2
Fig 2. Correlation between RNA-seq and qRT-PCR data on 15 selected genes with various fold changes.
Plotted by the logarithm of ratios of means between infected and uninfected samples. Primers are listed in S1 Table.
Fig 3
Fig 3. Gene ontology (GO) analysis of regulated genes in chicken type II pneumocyte in response to APEC-O78 infection.
GO terms in biological process, molecular function and cellular components. The black line stands for “p = 0.05”.
Fig 4
Fig 4. Pathway analysis of differentially expressed genes according to the KEGG database.
Top-ranking regulated pathways identified by KEGG according to P-value <0.01 and FDR <0.01.
Fig 5
Fig 5. Gene-Act network of differential genes according to pathways in the database.
Red dots represent up-regulated genes and green dots represent down-regulated genes. The arrows indicate the connection and regulatory relationship between two genes. Genes that have more connections with other genes have a higher degree score.
Fig 6
Fig 6. Electron micrographs of CP II cells.
(A) Viable cell (untreated); (B) Apoptotic cell (treated with APEC for 4h [MOI = 100]); (C) Necrotic cells (treated with APEC for 4h [MOI = 100]); (D) Cells treated with DH5α for 4h [MOI = 100].
Fig 7
Fig 7. The apoptosis rate of APEC-O78-induced CP II cells.
The CP II cells infected by APEC-O78 and DH5α were at a MOI of 100 for 4h. Then the cells were stained using the TransDetect Annexin V-FITC/PI Cell Apoptosis Detection Kit. Finally, the cells were analyzed with a FACSCalibur flow cytometer.
Fig 8
Fig 8. Changes in the expression levels of related immune resistance genes of CP II cells infected by APEC- O78.
After DH5α, APEC-O78 and heat inactivated APEC-O78 treated for 4 h with CP II cells, the expression levels of related immune resistance genes of CP II cells was detected by qRT-PCR. *P < 0.05 and **P < 0.01 are significantly different from the DH5α and heat inactivated APEC-O78 group.

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This work was supported by the National Natural Science Foundation of China (no. 31372470).
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