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miR-155 Controls Lymphoproliferation in LAT Mutant Mice by Restraining T-Cell Apoptosis via SHIP-1/mTOR and PAK1/FOXO3/BIM Pathways.

Rouquette-Jazdanian AK, Kortum RL, Li W, Merrill RK, Nguyen PH, Samelson LE, Sommers CL - PLoS ONE (2015)

Bottom Line: Using miR-155-deficient, LAT mutant T cells as a discovery tool, we found two connected pathways that impact the nuclear translocation and activation of FOXO3 in T cells.This study establishes a role for PAK1 in T cell apoptosis, which contrasts to its previously identified role in T cell proliferation.Furthermore, miR-155 regulates the delicate balance between PAK1-mediated proliferation and apoptosis in T cells impacting lymphoid organ size and function.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cellular and Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.

ABSTRACT
Linker for Activation of T cells (LAT) is an adapter protein that is essential for T cell function. Knock-in mice with a LAT mutation impairing calcium flux develop a fatal CD4+ lymphoproliferative disease. miR-155 is a microRNA that is correlated with hyperproliferation in a number of cancers including lymphomas and leukemias and is overexpressed in mutant LAT T cells. To test whether miR-155 was merely indicative of T cell activation or whether it contributes to lymphoproliferative disease in mutant LAT mice, we interbred LAT mutant and miR-155-deficient mice. miR-155 deficiency markedly inhibited lymphoproliferative disease by stimulating BIM-dependent CD4+ T cell apoptosis, even though ERK activation and T cell proliferation were increased in double mutant CD4+ T cells. Bim/Bcl2l11 expression is activated by the forkhead transcription factor FOXO3. Using miR-155-deficient, LAT mutant T cells as a discovery tool, we found two connected pathways that impact the nuclear translocation and activation of FOXO3 in T cells. One pathway is mediated by the inositide phosphatase SHIP-1 and the serine/threonine kinases AKT and PDK1. The other pathway involves PAK1 and JNK kinase activation. We define crosstalk between the two pathways via the kinase mTOR, which stabilizes PAK1. This study establishes a role for PAK1 in T cell apoptosis, which contrasts to its previously identified role in T cell proliferation. Furthermore, miR-155 regulates the delicate balance between PAK1-mediated proliferation and apoptosis in T cells impacting lymphoid organ size and function.

No MeSH data available.


Related in: MedlinePlus

miR-155 deficiency results in increased basal CD4+ T cell activation and proliferation in LAT-KI mice.(A) CD5 and CD69 surface marker expression as measured by flow cytometry in CD4+ lymph node T cells of mice of the indicated genotypes. Ages of the mice were the same as in Fig 1C. The results are representative of 9 experiments. (B) In vivo BrdU labeling. CD4+ lymph node T cells from mice previously injected with BrdU. Ages of the mice were 10 wks (WT) and 8 wks (other 3 genotypes). The results are representative of 3 experiments. (C) Negatively selected CD4+ T cells from the indicated genotypes were stimulated with αCD3ε/CD4 (10 μg/ml, 3 min.). Ages of the mice were 10 wks (WT, miR155-/-), 11 wks (LAT-KI), and 12 wks (DM). SDS whole cell lysates (WCLs) were analyzed by western blotting (WB). A representative WB is shown (n = 3).
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pone.0131823.g002: miR-155 deficiency results in increased basal CD4+ T cell activation and proliferation in LAT-KI mice.(A) CD5 and CD69 surface marker expression as measured by flow cytometry in CD4+ lymph node T cells of mice of the indicated genotypes. Ages of the mice were the same as in Fig 1C. The results are representative of 9 experiments. (B) In vivo BrdU labeling. CD4+ lymph node T cells from mice previously injected with BrdU. Ages of the mice were 10 wks (WT) and 8 wks (other 3 genotypes). The results are representative of 3 experiments. (C) Negatively selected CD4+ T cells from the indicated genotypes were stimulated with αCD3ε/CD4 (10 μg/ml, 3 min.). Ages of the mice were 10 wks (WT, miR155-/-), 11 wks (LAT-KI), and 12 wks (DM). SDS whole cell lysates (WCLs) were analyzed by western blotting (WB). A representative WB is shown (n = 3).

Mentions: We previously reported that LAT-KI mice show elevated basal ERK activation that is instrumental to their lymphoproliferative phenotype, and that introduction of mutations of either Bam32 or RasGRP lead to reduced lymphoproliferative disease in LAT-KI mice [5, 6]. These two molecules are components of two separate pathways that lead to ERK activation in LAT-KI T cells. In double mutant BAM32-/-LAT-KI [5]or RASGRP1-/-LAT-KI T cells [6], proliferation and elevated basal ERK activity were decreased as were levels of markers of T cell activation, CD5 and CD69, when compared to LAT-KI T cells. In contrast, the DM T cells analyzed here showed a completely different phenotype. LAT-KI CD4+ T cells expressed elevated levels of CD5 and higher percentages of CD69hi cells than WT CD4+ T cells [2, 3]. Surprisingly, although there are fewer CD4+ T cells in DM compared to LAT-KI mice, DM CD4+ T cells expressed even higher levels of CD5 and higher percentages of CD69hi cells than CD4+ T cells from age-matched LAT-KI mice (Fig 2A). We also observed a higher percentage of proliferating CD4+ T cells from DM mice compared to age-matched LAT-KI mice as measured by in vivo BrdU incorporation (Fig 2B). Therefore, the reduced number of CD4+ T cells in DM mice appeared to be more activated and were proliferating at a higher rate than CD4+ T cells from LAT-KI mice. These paradoxical observations led us to attempt to characterize signaling pathways downstream of miR-155 that might affect lymphoproliferative disease.


miR-155 Controls Lymphoproliferation in LAT Mutant Mice by Restraining T-Cell Apoptosis via SHIP-1/mTOR and PAK1/FOXO3/BIM Pathways.

Rouquette-Jazdanian AK, Kortum RL, Li W, Merrill RK, Nguyen PH, Samelson LE, Sommers CL - PLoS ONE (2015)

miR-155 deficiency results in increased basal CD4+ T cell activation and proliferation in LAT-KI mice.(A) CD5 and CD69 surface marker expression as measured by flow cytometry in CD4+ lymph node T cells of mice of the indicated genotypes. Ages of the mice were the same as in Fig 1C. The results are representative of 9 experiments. (B) In vivo BrdU labeling. CD4+ lymph node T cells from mice previously injected with BrdU. Ages of the mice were 10 wks (WT) and 8 wks (other 3 genotypes). The results are representative of 3 experiments. (C) Negatively selected CD4+ T cells from the indicated genotypes were stimulated with αCD3ε/CD4 (10 μg/ml, 3 min.). Ages of the mice were 10 wks (WT, miR155-/-), 11 wks (LAT-KI), and 12 wks (DM). SDS whole cell lysates (WCLs) were analyzed by western blotting (WB). A representative WB is shown (n = 3).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4487994&req=5

pone.0131823.g002: miR-155 deficiency results in increased basal CD4+ T cell activation and proliferation in LAT-KI mice.(A) CD5 and CD69 surface marker expression as measured by flow cytometry in CD4+ lymph node T cells of mice of the indicated genotypes. Ages of the mice were the same as in Fig 1C. The results are representative of 9 experiments. (B) In vivo BrdU labeling. CD4+ lymph node T cells from mice previously injected with BrdU. Ages of the mice were 10 wks (WT) and 8 wks (other 3 genotypes). The results are representative of 3 experiments. (C) Negatively selected CD4+ T cells from the indicated genotypes were stimulated with αCD3ε/CD4 (10 μg/ml, 3 min.). Ages of the mice were 10 wks (WT, miR155-/-), 11 wks (LAT-KI), and 12 wks (DM). SDS whole cell lysates (WCLs) were analyzed by western blotting (WB). A representative WB is shown (n = 3).
Mentions: We previously reported that LAT-KI mice show elevated basal ERK activation that is instrumental to their lymphoproliferative phenotype, and that introduction of mutations of either Bam32 or RasGRP lead to reduced lymphoproliferative disease in LAT-KI mice [5, 6]. These two molecules are components of two separate pathways that lead to ERK activation in LAT-KI T cells. In double mutant BAM32-/-LAT-KI [5]or RASGRP1-/-LAT-KI T cells [6], proliferation and elevated basal ERK activity were decreased as were levels of markers of T cell activation, CD5 and CD69, when compared to LAT-KI T cells. In contrast, the DM T cells analyzed here showed a completely different phenotype. LAT-KI CD4+ T cells expressed elevated levels of CD5 and higher percentages of CD69hi cells than WT CD4+ T cells [2, 3]. Surprisingly, although there are fewer CD4+ T cells in DM compared to LAT-KI mice, DM CD4+ T cells expressed even higher levels of CD5 and higher percentages of CD69hi cells than CD4+ T cells from age-matched LAT-KI mice (Fig 2A). We also observed a higher percentage of proliferating CD4+ T cells from DM mice compared to age-matched LAT-KI mice as measured by in vivo BrdU incorporation (Fig 2B). Therefore, the reduced number of CD4+ T cells in DM mice appeared to be more activated and were proliferating at a higher rate than CD4+ T cells from LAT-KI mice. These paradoxical observations led us to attempt to characterize signaling pathways downstream of miR-155 that might affect lymphoproliferative disease.

Bottom Line: Using miR-155-deficient, LAT mutant T cells as a discovery tool, we found two connected pathways that impact the nuclear translocation and activation of FOXO3 in T cells.This study establishes a role for PAK1 in T cell apoptosis, which contrasts to its previously identified role in T cell proliferation.Furthermore, miR-155 regulates the delicate balance between PAK1-mediated proliferation and apoptosis in T cells impacting lymphoid organ size and function.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cellular and Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.

ABSTRACT
Linker for Activation of T cells (LAT) is an adapter protein that is essential for T cell function. Knock-in mice with a LAT mutation impairing calcium flux develop a fatal CD4+ lymphoproliferative disease. miR-155 is a microRNA that is correlated with hyperproliferation in a number of cancers including lymphomas and leukemias and is overexpressed in mutant LAT T cells. To test whether miR-155 was merely indicative of T cell activation or whether it contributes to lymphoproliferative disease in mutant LAT mice, we interbred LAT mutant and miR-155-deficient mice. miR-155 deficiency markedly inhibited lymphoproliferative disease by stimulating BIM-dependent CD4+ T cell apoptosis, even though ERK activation and T cell proliferation were increased in double mutant CD4+ T cells. Bim/Bcl2l11 expression is activated by the forkhead transcription factor FOXO3. Using miR-155-deficient, LAT mutant T cells as a discovery tool, we found two connected pathways that impact the nuclear translocation and activation of FOXO3 in T cells. One pathway is mediated by the inositide phosphatase SHIP-1 and the serine/threonine kinases AKT and PDK1. The other pathway involves PAK1 and JNK kinase activation. We define crosstalk between the two pathways via the kinase mTOR, which stabilizes PAK1. This study establishes a role for PAK1 in T cell apoptosis, which contrasts to its previously identified role in T cell proliferation. Furthermore, miR-155 regulates the delicate balance between PAK1-mediated proliferation and apoptosis in T cells impacting lymphoid organ size and function.

No MeSH data available.


Related in: MedlinePlus