원문정보
초록
영어
Interleukin 11 (IL-11) is a secretory cytokine with pleotropic properties, including anti-inflammatory and antiapoptotic functions. This study aimed to functionally characterize a teleostean IL-11a ortholog from redlip mullet (LhIL-11a) through bioinformatic analysis, transcriptional expression profiling and protein function assays. The deduced LhIL-11a protein sequence is 200 amino acids long, with a predicted molecular weight of 23.168 kDa. Multiple sequence alignment indicates that LhIL-11a has a typical four-bundle architecture of α- helixes as observed in other IL-11s. The identity–similarity matrix show a higher identity between LhIL-11a and other fish IL-11a sequences. Phylogenetic analysis demonstrated that LhIL-11a falls within a clade including other fish counterparts. In the tissue distribution analysis, the highest constitutive expression of LhIL-11a mRNA was observed in the mullet gastrointestinal tract and brain tissues. Following the challenges with LPS, poly I:C and Lactococcus garvie, the transcription levels of LhIL-11a were significantly upregulated in both PBCs and liver. In the biological functional assay, recombinant LhIL-11a protein showed strong activities of suppressing proinflammatory cytokines and apoptotic gene expression in mullet kidney cells and reducing LPS stimulated NO production in murine macrophage cells. Overall, the findings in this study provide the experimental clues to understanding the functional roles of fish IL-11a in inflammation and apoptosis regulation during host defense against invading microbial pathogens.
목차
1. Introduction
2. Materials and methods
2.1. Rearing of fish and tissue collection
2.2. Transcriptomic database construction
2.3. Bioinformatics analysis
2.4. Immune challenge experiment
2.5. Total RNA isolation and cDNA synthesis
2.6. Analysis of spatial and temporal IL-11expression variation by real-timePCR (qRT-PCR)
2.7. Plasmid construction, overexpression, and purification of rLhIL-11a
2.8. rLhIL-11a treatment in mullet kidney cells
2.9. NO production assay
3. Results and discussion
3.1. LhIL-11a sequence analysis
3.2. Sequence alignment and phylogenetic relationship of LhIL-11a withother homologs
3.3. Three dimensional (3D) modeling of LhIL-11a tertiary structure
3.4. Expression of LhIL-11a in different redlip mullet tissues
3.5. LhIL-11a expression under simulated pathogenic stress in mullet
3.6. Biological activity of rLhIL-11a on red lip mullet kidney cells
4. Conclusion
Acknowledgments
Appendix A. Supplementary data
References