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ADAM17 deletion in thymic epithelial cells alters aire expression without affecting T cell developmental progression.

Gravano DM, McLelland BT, Horiuchi K, Manilay JO - PLoS ONE (2010)

Bottom Line: However, knockdown of Adam17 results in multisystem developmental defects and perinatal lethality, which has made study of the role of Adam17 in specific cell types difficult.Surprisingly, we have uncovered a novel genetic link between Adam17and Aire expression in vivo.The cell type in which ADAM17 mediates its non-cell autonomous impact and the mechanisms by which it regulates intrathymic T cell development remain to be identified.

View Article: PubMed Central - PubMed

Affiliation: School of Natural Sciences, University of California at Merced, Merced, California, United States of America.

ABSTRACT

Background: Cellular interactions between thymocytes and thymic stromal cells are critical for normal T cell development. Thymic epithelial cells (TECs) are important stromal niche cells that provide essential growth factors, cytokines, and present self-antigens to developing thymocytes. The identification of genes that mediate cellular crosstalk in the thymus is ongoing. One candidate gene, Adam17, encodes a metalloprotease that functions by cleaving the ectodomain of several transmembrane proteins and regulates various developmental processes. In conventional Adam17 knockout mice, a non-cell autonomous role for ADAM17 in adult T cell development was reported, which strongly suggested that expression of ADAM17 in TECs was required for normal T cell development. However, knockdown of Adam17 results in multisystem developmental defects and perinatal lethality, which has made study of the role of Adam17 in specific cell types difficult. Here, we examined T cell and thymic epithelial cell development using a conditional knockout approach.

Methodology/principal findings: We generated an Adam17 conditional knockout mouse in which floxed Adam17 is deleted specifically in TECs by Cre recombinase under the control of the Foxn1 promoter. Normal T cell lineage choice and development through the canonical αβ T cell stages was observed. Interestingly, Adam17 deficiency in TECs resulted in reduced expression of the transcription factor Aire. However, no alterations in the patterns of TEC phenotypic marker expression and thymus morphology were noted.

Conclusions/significance: In contrast to expectation, our data clearly shows that absence of Adam17 in TECs is dispensable for normal T cell development. Differentiation of TECs is also unaffected by loss of Adam17 based on phenotypic markers. Surprisingly, we have uncovered a novel genetic link between Adam17and Aire expression in vivo. The cell type in which ADAM17 mediates its non-cell autonomous impact and the mechanisms by which it regulates intrathymic T cell development remain to be identified.

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T cell developmental progression and thymocyte cellularity are unaltered in Adam17/Foxn1 mice.(A) Representative image of the thymus and spleen extracted from 4-week old Control or Adam17/Foxn1 mice. (B) Total cell numbers from individual mice of the indicated genotype. Data represent mean + SD; Control, n = 6; Adam17/Foxn1, n = 8. p>0.05 for all comparisons between Control and Adam17/Foxn1 thymocyte populations. (C) Representative flow cytometry data from Control and Adam17/Foxn1 mice showing CD4 versus CD8 profiles and DN1 through DN4 stages, as assessed by CD44 and CD25 staining within the DN gate. Control: fl/+ or fl/fl; Adam17/Foxn1: Cre fl/fl.
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pone-0013528-g002: T cell developmental progression and thymocyte cellularity are unaltered in Adam17/Foxn1 mice.(A) Representative image of the thymus and spleen extracted from 4-week old Control or Adam17/Foxn1 mice. (B) Total cell numbers from individual mice of the indicated genotype. Data represent mean + SD; Control, n = 6; Adam17/Foxn1, n = 8. p>0.05 for all comparisons between Control and Adam17/Foxn1 thymocyte populations. (C) Representative flow cytometry data from Control and Adam17/Foxn1 mice showing CD4 versus CD8 profiles and DN1 through DN4 stages, as assessed by CD44 and CD25 staining within the DN gate. Control: fl/+ or fl/fl; Adam17/Foxn1: Cre fl/fl.

Mentions: Conventional Adam17 KO mice exhibit a non-cell autonomous block in T cell development from the DN to DP stage, which indicated a likely role for ADAM17 on TECs [25]. However, in our TEC-specific Adam17/Foxn1 knockout mice, the thymus and spleen are of normal size (Figure 2A). Likewise, analysis of intrathymic populations revealed indistinguishable numbers of DN, DP, CD4SP, and CD8SP thymocytes between control and Adam17/Foxn1 mice (Figure 2B). Furthermore, the frequencies and absolute numbers of DN1 through DN4 stages of thymocyte development were unaltered in the absence of ADAM17 on TECs (Figure 2C, and data not shown). Likewise, analysis of regulatory T cells showed no alterations in the frequency of CD4SP thymocytes expressing FoxP3 or CD25 nor CD4SP splenocytes expressing FoxP3 (Figure S1). These results are not consistent with the hypothesis that the ADAM17 non-cell autonomous role in T cell development is localized to the TECs.


ADAM17 deletion in thymic epithelial cells alters aire expression without affecting T cell developmental progression.

Gravano DM, McLelland BT, Horiuchi K, Manilay JO - PLoS ONE (2010)

T cell developmental progression and thymocyte cellularity are unaltered in Adam17/Foxn1 mice.(A) Representative image of the thymus and spleen extracted from 4-week old Control or Adam17/Foxn1 mice. (B) Total cell numbers from individual mice of the indicated genotype. Data represent mean + SD; Control, n = 6; Adam17/Foxn1, n = 8. p>0.05 for all comparisons between Control and Adam17/Foxn1 thymocyte populations. (C) Representative flow cytometry data from Control and Adam17/Foxn1 mice showing CD4 versus CD8 profiles and DN1 through DN4 stages, as assessed by CD44 and CD25 staining within the DN gate. Control: fl/+ or fl/fl; Adam17/Foxn1: Cre fl/fl.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2958126&req=5

pone-0013528-g002: T cell developmental progression and thymocyte cellularity are unaltered in Adam17/Foxn1 mice.(A) Representative image of the thymus and spleen extracted from 4-week old Control or Adam17/Foxn1 mice. (B) Total cell numbers from individual mice of the indicated genotype. Data represent mean + SD; Control, n = 6; Adam17/Foxn1, n = 8. p>0.05 for all comparisons between Control and Adam17/Foxn1 thymocyte populations. (C) Representative flow cytometry data from Control and Adam17/Foxn1 mice showing CD4 versus CD8 profiles and DN1 through DN4 stages, as assessed by CD44 and CD25 staining within the DN gate. Control: fl/+ or fl/fl; Adam17/Foxn1: Cre fl/fl.
Mentions: Conventional Adam17 KO mice exhibit a non-cell autonomous block in T cell development from the DN to DP stage, which indicated a likely role for ADAM17 on TECs [25]. However, in our TEC-specific Adam17/Foxn1 knockout mice, the thymus and spleen are of normal size (Figure 2A). Likewise, analysis of intrathymic populations revealed indistinguishable numbers of DN, DP, CD4SP, and CD8SP thymocytes between control and Adam17/Foxn1 mice (Figure 2B). Furthermore, the frequencies and absolute numbers of DN1 through DN4 stages of thymocyte development were unaltered in the absence of ADAM17 on TECs (Figure 2C, and data not shown). Likewise, analysis of regulatory T cells showed no alterations in the frequency of CD4SP thymocytes expressing FoxP3 or CD25 nor CD4SP splenocytes expressing FoxP3 (Figure S1). These results are not consistent with the hypothesis that the ADAM17 non-cell autonomous role in T cell development is localized to the TECs.

Bottom Line: However, knockdown of Adam17 results in multisystem developmental defects and perinatal lethality, which has made study of the role of Adam17 in specific cell types difficult.Surprisingly, we have uncovered a novel genetic link between Adam17and Aire expression in vivo.The cell type in which ADAM17 mediates its non-cell autonomous impact and the mechanisms by which it regulates intrathymic T cell development remain to be identified.

View Article: PubMed Central - PubMed

Affiliation: School of Natural Sciences, University of California at Merced, Merced, California, United States of America.

ABSTRACT

Background: Cellular interactions between thymocytes and thymic stromal cells are critical for normal T cell development. Thymic epithelial cells (TECs) are important stromal niche cells that provide essential growth factors, cytokines, and present self-antigens to developing thymocytes. The identification of genes that mediate cellular crosstalk in the thymus is ongoing. One candidate gene, Adam17, encodes a metalloprotease that functions by cleaving the ectodomain of several transmembrane proteins and regulates various developmental processes. In conventional Adam17 knockout mice, a non-cell autonomous role for ADAM17 in adult T cell development was reported, which strongly suggested that expression of ADAM17 in TECs was required for normal T cell development. However, knockdown of Adam17 results in multisystem developmental defects and perinatal lethality, which has made study of the role of Adam17 in specific cell types difficult. Here, we examined T cell and thymic epithelial cell development using a conditional knockout approach.

Methodology/principal findings: We generated an Adam17 conditional knockout mouse in which floxed Adam17 is deleted specifically in TECs by Cre recombinase under the control of the Foxn1 promoter. Normal T cell lineage choice and development through the canonical αβ T cell stages was observed. Interestingly, Adam17 deficiency in TECs resulted in reduced expression of the transcription factor Aire. However, no alterations in the patterns of TEC phenotypic marker expression and thymus morphology were noted.

Conclusions/significance: In contrast to expectation, our data clearly shows that absence of Adam17 in TECs is dispensable for normal T cell development. Differentiation of TECs is also unaffected by loss of Adam17 based on phenotypic markers. Surprisingly, we have uncovered a novel genetic link between Adam17and Aire expression in vivo. The cell type in which ADAM17 mediates its non-cell autonomous impact and the mechanisms by which it regulates intrathymic T cell development remain to be identified.

Show MeSH
Related in: MedlinePlus