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Epithelia Use Butyrophilin-like Molecules to Shape Organ-Specific γ δ T Cell Compartments

View Article: PubMed Central - PubMed

ABSTRACT

Many body surfaces harbor organ-specific γδ T cell compartments that contribute to tissue integrity. Thus, murine dendritic epidermal T cells (DETCs) uniquely expressing T cell receptor (TCR)-Vγ5 chains protect from cutaneous carcinogens. The DETC repertoire is shaped by Skint1, a butyrophilin-like (Btnl) gene expressed specifically by thymic epithelial cells and suprabasal keratinocytes. However, the generality of this mechanism has remained opaque, since neither Skint1 nor DETCs are evolutionarily conserved. Here, Btnl1 expressed by murine enterocytes is shown to shape the local TCR-Vγ7+ γδ compartment. Uninfluenced by microbial or food antigens, this activity evokes the developmental selection of TCRαβ+ repertoires. Indeed, Btnl1 and Btnl6 jointly induce TCR-dependent responses specifically in intestinal Vγ7+ cells. Likewise, human gut epithelial cells express BTNL3 and BTNL8 that jointly induce selective TCR-dependent responses of human colonic Vγ4+ cells. Hence, a conserved mechanism emerges whereby epithelia use organ-specific BTNL/Btnl genes to shape local T cell compartments.

No MeSH data available.


Related in: MedlinePlus

Inducible Btnl1 Transgene Expression and Its Impact in Adult Mice, Related to Figure 5(A) Schematic representation of the WT Btnl1 locus (top) and TRE-Btnl1 transgene construct (bottom). Grey: unstranslated region; green: translated region; orange: upstream-tetracycline response element/CMV promoter and downstream-β-globulin/polyA. (B) Southern blot to detect transgene insertion. Genomic DNA was digested with EcoR1 as indicated (arrowheads) and a probe (blue bar) targeting the indicated region (Exon3/4 boundary in ORF) was generated to detect the WT and targeted allele (n = 2). C-F) W7-13 (ADULT) mice of indicated genotypes on a Btnl1−/− background were administered doxycycline water (1mg/ml Dox, 2% suchrose) or ctrl water (2% suchrose) for 1-2 weeks. (C) Gene expression by qRT.PCR in proximal small intestine of adult mice following the indicated treatment. (D) γδIEL composition (left) and absolute cell counts (right) assessed by flow cytometry in adult mice following the indicated treatment (sugar, n = 3-5; rest, n = 4-10). (E) Ki67 and cell surface CD122 expression in Vγ7+ IEL from adult mice following the indicated treatment. (F) Ki67 expression in Vγ7+ versus Vγ7- and TCRβ+ IEL from adult mice following the indicated treatment (n = 5-9). Data are representative of 2 (B), or 3 independent experiments. Some panels include results pooled from 2 (C) or ≥ 3 (D,F) independent experiments. All error bars represent mean ± SD.
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figs5: Inducible Btnl1 Transgene Expression and Its Impact in Adult Mice, Related to Figure 5(A) Schematic representation of the WT Btnl1 locus (top) and TRE-Btnl1 transgene construct (bottom). Grey: unstranslated region; green: translated region; orange: upstream-tetracycline response element/CMV promoter and downstream-β-globulin/polyA. (B) Southern blot to detect transgene insertion. Genomic DNA was digested with EcoR1 as indicated (arrowheads) and a probe (blue bar) targeting the indicated region (Exon3/4 boundary in ORF) was generated to detect the WT and targeted allele (n = 2). C-F) W7-13 (ADULT) mice of indicated genotypes on a Btnl1−/− background were administered doxycycline water (1mg/ml Dox, 2% suchrose) or ctrl water (2% suchrose) for 1-2 weeks. (C) Gene expression by qRT.PCR in proximal small intestine of adult mice following the indicated treatment. (D) γδIEL composition (left) and absolute cell counts (right) assessed by flow cytometry in adult mice following the indicated treatment (sugar, n = 3-5; rest, n = 4-10). (E) Ki67 and cell surface CD122 expression in Vγ7+ IEL from adult mice following the indicated treatment. (F) Ki67 expression in Vγ7+ versus Vγ7- and TCRβ+ IEL from adult mice following the indicated treatment (n = 5-9). Data are representative of 2 (B), or 3 independent experiments. Some panels include results pooled from 2 (C) or ≥ 3 (D,F) independent experiments. All error bars represent mean ± SD.

Mentions: To attempt to restore IEL selection, we rendered Btnl1−/− mice transgenic for Btnl1 expressed from a doxycycline (Dox)-inducible promoter (Figures S5A and S5B) and crossed them onto Btnl1−/− mice, into which was introduced a Dox-responsive trans-activator (rtTA) regulated by the ubiquitously expressed Rosa26 promoter. As reported, adding low-dose Dox to drinking water had little overt impact on the gut over a several-week period (Roth et al., 2009). Therefore, this system offered a means to globally induce Btnl1 de novo in Btnl1−/− bitransgenic (BiTg) mice that inherited both rtTA and the inducible Btnl1 transgene. Conversely, BiTg mice administered sugar water and Dox-treated single-transgenic (SiTg) Btnl1−/− mice (that inherited only the Btnl1 transgene) served as controls. Btnl1 induction in mice of appropriate genotypes was validated by qPCR (Figure S5C).


Epithelia Use Butyrophilin-like Molecules to Shape Organ-Specific γ δ T Cell Compartments
Inducible Btnl1 Transgene Expression and Its Impact in Adult Mice, Related to Figure 5(A) Schematic representation of the WT Btnl1 locus (top) and TRE-Btnl1 transgene construct (bottom). Grey: unstranslated region; green: translated region; orange: upstream-tetracycline response element/CMV promoter and downstream-β-globulin/polyA. (B) Southern blot to detect transgene insertion. Genomic DNA was digested with EcoR1 as indicated (arrowheads) and a probe (blue bar) targeting the indicated region (Exon3/4 boundary in ORF) was generated to detect the WT and targeted allele (n = 2). C-F) W7-13 (ADULT) mice of indicated genotypes on a Btnl1−/− background were administered doxycycline water (1mg/ml Dox, 2% suchrose) or ctrl water (2% suchrose) for 1-2 weeks. (C) Gene expression by qRT.PCR in proximal small intestine of adult mice following the indicated treatment. (D) γδIEL composition (left) and absolute cell counts (right) assessed by flow cytometry in adult mice following the indicated treatment (sugar, n = 3-5; rest, n = 4-10). (E) Ki67 and cell surface CD122 expression in Vγ7+ IEL from adult mice following the indicated treatment. (F) Ki67 expression in Vγ7+ versus Vγ7- and TCRβ+ IEL from adult mice following the indicated treatment (n = 5-9). Data are representative of 2 (B), or 3 independent experiments. Some panels include results pooled from 2 (C) or ≥ 3 (D,F) independent experiments. All error bars represent mean ± SD.
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figs5: Inducible Btnl1 Transgene Expression and Its Impact in Adult Mice, Related to Figure 5(A) Schematic representation of the WT Btnl1 locus (top) and TRE-Btnl1 transgene construct (bottom). Grey: unstranslated region; green: translated region; orange: upstream-tetracycline response element/CMV promoter and downstream-β-globulin/polyA. (B) Southern blot to detect transgene insertion. Genomic DNA was digested with EcoR1 as indicated (arrowheads) and a probe (blue bar) targeting the indicated region (Exon3/4 boundary in ORF) was generated to detect the WT and targeted allele (n = 2). C-F) W7-13 (ADULT) mice of indicated genotypes on a Btnl1−/− background were administered doxycycline water (1mg/ml Dox, 2% suchrose) or ctrl water (2% suchrose) for 1-2 weeks. (C) Gene expression by qRT.PCR in proximal small intestine of adult mice following the indicated treatment. (D) γδIEL composition (left) and absolute cell counts (right) assessed by flow cytometry in adult mice following the indicated treatment (sugar, n = 3-5; rest, n = 4-10). (E) Ki67 and cell surface CD122 expression in Vγ7+ IEL from adult mice following the indicated treatment. (F) Ki67 expression in Vγ7+ versus Vγ7- and TCRβ+ IEL from adult mice following the indicated treatment (n = 5-9). Data are representative of 2 (B), or 3 independent experiments. Some panels include results pooled from 2 (C) or ≥ 3 (D,F) independent experiments. All error bars represent mean ± SD.
Mentions: To attempt to restore IEL selection, we rendered Btnl1−/− mice transgenic for Btnl1 expressed from a doxycycline (Dox)-inducible promoter (Figures S5A and S5B) and crossed them onto Btnl1−/− mice, into which was introduced a Dox-responsive trans-activator (rtTA) regulated by the ubiquitously expressed Rosa26 promoter. As reported, adding low-dose Dox to drinking water had little overt impact on the gut over a several-week period (Roth et al., 2009). Therefore, this system offered a means to globally induce Btnl1 de novo in Btnl1−/− bitransgenic (BiTg) mice that inherited both rtTA and the inducible Btnl1 transgene. Conversely, BiTg mice administered sugar water and Dox-treated single-transgenic (SiTg) Btnl1−/− mice (that inherited only the Btnl1 transgene) served as controls. Btnl1 induction in mice of appropriate genotypes was validated by qPCR (Figure S5C).

View Article: PubMed Central - PubMed

ABSTRACT

Many body surfaces harbor organ-specific γδ T cell compartments that contribute to tissue integrity. Thus, murine dendritic epidermal T cells (DETCs) uniquely expressing T cell receptor (TCR)-Vγ5 chains protect from cutaneous carcinogens. The DETC repertoire is shaped by Skint1, a butyrophilin-like (Btnl) gene expressed specifically by thymic epithelial cells and suprabasal keratinocytes. However, the generality of this mechanism has remained opaque, since neither Skint1 nor DETCs are evolutionarily conserved. Here, Btnl1 expressed by murine enterocytes is shown to shape the local TCR-Vγ7+ γδ compartment. Uninfluenced by microbial or food antigens, this activity evokes the developmental selection of TCRαβ+ repertoires. Indeed, Btnl1 and Btnl6 jointly induce TCR-dependent responses specifically in intestinal Vγ7+ cells. Likewise, human gut epithelial cells express BTNL3 and BTNL8 that jointly induce selective TCR-dependent responses of human colonic Vγ4+ cells. Hence, a conserved mechanism emerges whereby epithelia use organ-specific BTNL/Btnl genes to shape local T cell compartments.

No MeSH data available.


Related in: MedlinePlus