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Differential roles of interleukin 15 mRNA isoforms generated by alternative splicing in immune responses in vivo.

Nishimura H, Yajima T, Naiki Y, Tsunobuchi H, Umemura M, Itano K, Matsuguchi T, Suzuki M, Ohashi PS, Yoshikai Y - J. Exp. Med. (2000)

Bottom Line: At least two types of interleukin (IL)-15 mRNA isoforms are generated by alternative splicing at the 5' upstream of exon 5 in mice.On the other hand, a large amount of intracellular IL-15 protein was detected but hardly secreted extracellularly in alternative IL-15 Tg mice.Thus, two groups of IL-15 Tg mice may provide information concerning the different roles of IL-15 isoforms in the immune system in vivo.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Host Defense & Germfree Life, Research Institute for Disease Mechanism and Control, Nagoya University School of Medicine, Nagoya 466, Japan. nishihit@med.nagoya-u.ac.jp

ABSTRACT
At least two types of interleukin (IL)-15 mRNA isoforms are generated by alternative splicing at the 5' upstream of exon 5 in mice. To elucidate the potential roles of IL-15 isoforms in immune responses in vivo, we constructed two groups of transgenic mice using originally described IL-15 cDNA with a normal exon 5 (normal IL-15 transgenic [Tg] mice) and IL-15 cDNA with an alternative exon 5 (alternative IL-15 Tg mice) under the control of an MHC class I promoter. Normal IL-15 Tg mice constitutionally produced a significant level of IL-15 protein and had markedly increased numbers of memory type (CD44(high) Ly6C(+)) of CD8(+) T cells in the LN. These mice showed resistance to Salmonella infection accompanied by the enhanced interferon (IFN)-gamma production, but depletion of CD8(+) T cells exaggerated the bacterial growth, suggesting that the IL-15-dependent CD8(+) T cells with a memory phenotype may serve to protect against Salmonella infection in normal IL-15 Tg mice. On the other hand, a large amount of intracellular IL-15 protein was detected but hardly secreted extracellularly in alternative IL-15 Tg mice. Although most of the T cells developed normally in the alternative IL-15 Tg mice, they showed impaired IFN-gamma production upon TCR engagement. The alternative IL-15 transgenic mice were susceptible to Salmonella accompanied by impaired production of endogenous IL-15 and IFN-gamma. Thus, two groups of IL-15 Tg mice may provide information concerning the different roles of IL-15 isoforms in the immune system in vivo.

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Cell surface markers on CD44+CD8+ T cells in normal IL-15 Tg mice. (A) Expression of activation markers on the CD8+ T cells from the LN of normal IL-15 Tg mice was examined by a flow cytometer. The cells were stained with FITC-CD44, Biotin-CD8α, and PE-mAbs against various markers, and then they were analyzed by a flow cytometer, and the analysis gate was set on CD8+ cells. Data are representative of three independent experiments using pooled cells from three mice and are shown as typical two-color profiles. (B) Expression of IL-2 receptor subunits on CD8+ or CD4+ T cells from the LN of normal IL-15 Tg mice were examined. The cells were stained with PE-CD44, Cy-Chrome–CD4, or biotin-CD8α, and FITC-mAbs against IL-2 receptor subunits, and they were then analyzed by a flow cytometer, and the analysis gate was set on CD44+ or CD44− on CD4+ or CD8+ cells. Data are representative of three independent experiments using pooled cells from three mice and are shown as typical single-color profiles.
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Figure 4: Cell surface markers on CD44+CD8+ T cells in normal IL-15 Tg mice. (A) Expression of activation markers on the CD8+ T cells from the LN of normal IL-15 Tg mice was examined by a flow cytometer. The cells were stained with FITC-CD44, Biotin-CD8α, and PE-mAbs against various markers, and then they were analyzed by a flow cytometer, and the analysis gate was set on CD8+ cells. Data are representative of three independent experiments using pooled cells from three mice and are shown as typical two-color profiles. (B) Expression of IL-2 receptor subunits on CD8+ or CD4+ T cells from the LN of normal IL-15 Tg mice were examined. The cells were stained with PE-CD44, Cy-Chrome–CD4, or biotin-CD8α, and FITC-mAbs against IL-2 receptor subunits, and they were then analyzed by a flow cytometer, and the analysis gate was set on CD44+ or CD44− on CD4+ or CD8+ cells. Data are representative of three independent experiments using pooled cells from three mice and are shown as typical single-color profiles.

Mentions: To investigate T cell development in the IL-15 Tg mice, thymocytes and peripheral T cells from Tg or wild-type animals were analyzed by flow cytometry using antibodies specific for a variety of developmentally regulated T cell surface antigens. Thymi from newborn to 4-wk-old normal IL-15 Tg mice possessed an ∼25% higher level of cells than did control mice, whereas the level in thymi from alternative IL-15 Tg mice was very similar to that from control C57BL/6 mice. The thymi from both IL-15 Tg mice contained normal proportions of double negative CD4−CD8−, double positive CD4+CD8+, and single positive CD4+ or CD8+ cells. Levels of CD3 and TCR-α/β expression were also indistinguishable among wild-type and both IL-15 Tg mice, as were the expression levels of TCR-γ/δ and heat-stable antigen (data not shown). Thus, overexpression of the normal or alternative IL-15 transgene did not appear to perturb thymocyte ontogeny. The populations of CD44+CD8+ T cells in the spleen and the lymph node were threefold higher in normal IL-15 Tg mice (Table and Fig. 3). The CD44+CD8+ T cells showed a memory phenotype such as CD25−, CD69−, Ly-6C+, CD62L+, and CD45 RB+ ( Fig. 4a and Fig. b). The expression of IL-2Rα (CD25) was undetectable on CD44+CD8+ T cells, but IL-2/IL-15R β and γ chains were highly expressed on memory-phenotype CD44+CD8+ T cells. On the other hand, the proportion of memory-type CD8+ T cells in alternative IL-15 Tg mice was very similar to that in control mice. However, it should be noted that NK1.1+ cells in the spleen of alternative IL-15 Tg mice were significantly reduced in number compared with control mice ( Fig. 3).


Differential roles of interleukin 15 mRNA isoforms generated by alternative splicing in immune responses in vivo.

Nishimura H, Yajima T, Naiki Y, Tsunobuchi H, Umemura M, Itano K, Matsuguchi T, Suzuki M, Ohashi PS, Yoshikai Y - J. Exp. Med. (2000)

Cell surface markers on CD44+CD8+ T cells in normal IL-15 Tg mice. (A) Expression of activation markers on the CD8+ T cells from the LN of normal IL-15 Tg mice was examined by a flow cytometer. The cells were stained with FITC-CD44, Biotin-CD8α, and PE-mAbs against various markers, and then they were analyzed by a flow cytometer, and the analysis gate was set on CD8+ cells. Data are representative of three independent experiments using pooled cells from three mice and are shown as typical two-color profiles. (B) Expression of IL-2 receptor subunits on CD8+ or CD4+ T cells from the LN of normal IL-15 Tg mice were examined. The cells were stained with PE-CD44, Cy-Chrome–CD4, or biotin-CD8α, and FITC-mAbs against IL-2 receptor subunits, and they were then analyzed by a flow cytometer, and the analysis gate was set on CD44+ or CD44− on CD4+ or CD8+ cells. Data are representative of three independent experiments using pooled cells from three mice and are shown as typical single-color profiles.
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Figure 4: Cell surface markers on CD44+CD8+ T cells in normal IL-15 Tg mice. (A) Expression of activation markers on the CD8+ T cells from the LN of normal IL-15 Tg mice was examined by a flow cytometer. The cells were stained with FITC-CD44, Biotin-CD8α, and PE-mAbs against various markers, and then they were analyzed by a flow cytometer, and the analysis gate was set on CD8+ cells. Data are representative of three independent experiments using pooled cells from three mice and are shown as typical two-color profiles. (B) Expression of IL-2 receptor subunits on CD8+ or CD4+ T cells from the LN of normal IL-15 Tg mice were examined. The cells were stained with PE-CD44, Cy-Chrome–CD4, or biotin-CD8α, and FITC-mAbs against IL-2 receptor subunits, and they were then analyzed by a flow cytometer, and the analysis gate was set on CD44+ or CD44− on CD4+ or CD8+ cells. Data are representative of three independent experiments using pooled cells from three mice and are shown as typical single-color profiles.
Mentions: To investigate T cell development in the IL-15 Tg mice, thymocytes and peripheral T cells from Tg or wild-type animals were analyzed by flow cytometry using antibodies specific for a variety of developmentally regulated T cell surface antigens. Thymi from newborn to 4-wk-old normal IL-15 Tg mice possessed an ∼25% higher level of cells than did control mice, whereas the level in thymi from alternative IL-15 Tg mice was very similar to that from control C57BL/6 mice. The thymi from both IL-15 Tg mice contained normal proportions of double negative CD4−CD8−, double positive CD4+CD8+, and single positive CD4+ or CD8+ cells. Levels of CD3 and TCR-α/β expression were also indistinguishable among wild-type and both IL-15 Tg mice, as were the expression levels of TCR-γ/δ and heat-stable antigen (data not shown). Thus, overexpression of the normal or alternative IL-15 transgene did not appear to perturb thymocyte ontogeny. The populations of CD44+CD8+ T cells in the spleen and the lymph node were threefold higher in normal IL-15 Tg mice (Table and Fig. 3). The CD44+CD8+ T cells showed a memory phenotype such as CD25−, CD69−, Ly-6C+, CD62L+, and CD45 RB+ ( Fig. 4a and Fig. b). The expression of IL-2Rα (CD25) was undetectable on CD44+CD8+ T cells, but IL-2/IL-15R β and γ chains were highly expressed on memory-phenotype CD44+CD8+ T cells. On the other hand, the proportion of memory-type CD8+ T cells in alternative IL-15 Tg mice was very similar to that in control mice. However, it should be noted that NK1.1+ cells in the spleen of alternative IL-15 Tg mice were significantly reduced in number compared with control mice ( Fig. 3).

Bottom Line: At least two types of interleukin (IL)-15 mRNA isoforms are generated by alternative splicing at the 5' upstream of exon 5 in mice.On the other hand, a large amount of intracellular IL-15 protein was detected but hardly secreted extracellularly in alternative IL-15 Tg mice.Thus, two groups of IL-15 Tg mice may provide information concerning the different roles of IL-15 isoforms in the immune system in vivo.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Host Defense & Germfree Life, Research Institute for Disease Mechanism and Control, Nagoya University School of Medicine, Nagoya 466, Japan. nishihit@med.nagoya-u.ac.jp

ABSTRACT
At least two types of interleukin (IL)-15 mRNA isoforms are generated by alternative splicing at the 5' upstream of exon 5 in mice. To elucidate the potential roles of IL-15 isoforms in immune responses in vivo, we constructed two groups of transgenic mice using originally described IL-15 cDNA with a normal exon 5 (normal IL-15 transgenic [Tg] mice) and IL-15 cDNA with an alternative exon 5 (alternative IL-15 Tg mice) under the control of an MHC class I promoter. Normal IL-15 Tg mice constitutionally produced a significant level of IL-15 protein and had markedly increased numbers of memory type (CD44(high) Ly6C(+)) of CD8(+) T cells in the LN. These mice showed resistance to Salmonella infection accompanied by the enhanced interferon (IFN)-gamma production, but depletion of CD8(+) T cells exaggerated the bacterial growth, suggesting that the IL-15-dependent CD8(+) T cells with a memory phenotype may serve to protect against Salmonella infection in normal IL-15 Tg mice. On the other hand, a large amount of intracellular IL-15 protein was detected but hardly secreted extracellularly in alternative IL-15 Tg mice. Although most of the T cells developed normally in the alternative IL-15 Tg mice, they showed impaired IFN-gamma production upon TCR engagement. The alternative IL-15 transgenic mice were susceptible to Salmonella accompanied by impaired production of endogenous IL-15 and IFN-gamma. Thus, two groups of IL-15 Tg mice may provide information concerning the different roles of IL-15 isoforms in the immune system in vivo.

Show MeSH
Related in: MedlinePlus