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Cloning, expression, and tissue distribution of bovine interleukin-21
Yoshihiro Muneta1, Reiko Kikuma, Kazuhiro Yoshihara
1National Institute of Animal Health, 3-1-5 Kannondai Tsukuba, Ibaraki 305-0856, Japan. ymuneta@affrc.go.jp
Insights
Researchers cloned and sequenced bovine interleukin-21 (IL-21) from lymphocytes. The recombinant bovine IL-21 enhanced human cell proliferation, suggesting potential for boosting innate immunity in cows.
Area of Science:
- Immunology
- Molecular Biology
- Veterinary Science
Background:
- Interleukin-21 (IL-21) is a cytokine crucial for immune responses.
- Understanding bovine IL-21 is vital for advancing cattle immunology and health.
Purpose of the Study:
- To clone and characterize the bovine IL-21 cDNA.
- To express and functionally analyze recombinant bovine IL-21.
- To investigate the tissue-specific mRNA expression of bovine IL-21.
Main Methods:
- Cloning and sequencing of bovine IL-21 cDNA from stimulated peripheral blood lymphocytes (PBLs).
- Expression of recombinant bovine IL-21 using a baculovirus system.
- Functional assay using human IL-2-dependent cells (ILT-MAT) to assess proliferation.
- mRNA expression analysis in various bovine tissues via RT-PCR (implied).
Main Results:
- Bovine IL-21 cDNA sequence determined, encoding 152 amino acids with significant homology to human and murine IL-21.
- Recombinant bovine IL-21 was successfully expressed, processed to its mature form, and secreted.
- Recombinant bovine IL-21 induced proliferation in human IL-2-dependent cells.
- Bovine IL-21 mRNA was detected in the spleen but not in other tested organs.
Conclusions:
- The study successfully characterized bovine IL-21, providing a valuable resource for immunological research.
- Recombinant bovine IL-21 demonstrates biological activity, stimulating immune cell proliferation.
- The findings suggest a potential role for bovine IL-21 in enhancing innate immunity in cattle.
Abstract:
Bovine interleukin-21 (IL-21) cDNA was cloned and sequenced from bovine peripheral blood lymphocytes (PBLs) stimulated with 10 microg/ml concanavalin A (ConA), 10 microg/ml phytohemagglutinin (PHA), and 50 ng/ml phorbol 12-myristate 13-acetate (PMA) for 48 h. The open reading frame of the bovine IL-21 cDNA is 459 bp in length and encodes 152 amino acids. The predicted amino acid sequence is 78.2 and 58.5% homologous to the human and murine IL-21 amino acid sequences, respectively. Recombinant bovine IL-21 was expressed by a baculovirus expression system. The bovine IL-21 was processed to the mature form in insect cells and secreted to the supernatant confirmed by N-terminal amino acid sequencing. The recombinant bovine mature IL-21 induced the proliferation of human IL-2-dependent cells, ILT-MAT. The mRNA expression for bovine IL-21 was observed in the spleen, but not in the brain, heart, lung, liver, and kidney. The bovine IL-21 identified in this study may provide new methods for the enhancement of innate immunity in cows.
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