Wip1 phosphatase-deficient mice exhibit defective T cell maturation due to sustained p53 activation.
ABSTRACT The PP2C phosphatase Wip1 dephosphorylates p38 and blocks UV-induced p53 activation in cultured human cells. Although the level of TCR-induced p38 MAPK activity is initially comparable between Wip1-/- and wild-type thymocytes, phosphatase-deficient cells failed to down-regulate p38 MAPK activity after 6 h. Analysis of young Wip1-deficient mice showed that they had fewer splenic T cells. Their thymi were smaller, contained significantly fewer cells, and failed to undergo age-dependent involution compared with wild-type animals. Analysis of thymocyte subset numbers by flow cytometry suggested that cell numbers starting at the double-negative (DN)4 stage are significantly reduced in Wip1-deficient mice, and p53 activity is elevated in cell-sorted DN4 and double-positive subpopulations. Although apoptosis and proliferation was normal in Wip1-/- DN4 cells, they appeared to be in cell cycle arrest. In contrast, a significantly higher percentage of apoptotic cells were found in the double-positive population, and down-regulation of thymocyte p38 MAPK activation by anti-CD3 was delayed. To examine the role of p38 MAPK in early thymic subpopulations, fetal thymic organ cultures cultured in the presence/absence of a p38 MAPK inhibitor did not correct the thymic phenotype. In contrast, the abnormal thymic phenotype of Wip1-deficient mice was reversed in the absence of p53. These data suggest that Wip1 down-regulates p53 activation in the thymus and is required for normal alphabeta T cell development.
SourceAvailable from: Alice Gentil Dit Maurin[Show abstract] [Hide abstract]
ABSTRACT: The production of T cell receptor αβ(+) (TCRαβ(+) ) T lymphocytes in the thymus is a tightly regulated process that can be monitored by the regulated expression of several surface molecules, including CD4, CD8, cKit, CD25 and the TCR itself, after TCR genes have been assembled from discrete V, D (for TCRβ) and J gene segments by a site-directed genetic recombination. Thymocyte differentiation is the result of a delicate balance between cell death and survival: developing thymocytes die unless they receive a positive signal to proceed to the next stage. This equilibrium is altered in response to various physiological or physical stresses such as ionizing radiation, which induces a massive p53-dependent apoptosis of CD4(+) CD8(+) double positive (DP) thymocytes. Interestingly, these cells are actively rearranging their TCRα chain genes. To unravel an eventual link between V(D)J recombination activity and thymocyte radiosensitivity, we analyzed the dynamics of thymocyte apoptosis and regeneration following exposure of wild type and p53-deficient mice to different doses of γ-radiation. p53-dependent radiosensitivity was already found high in immature CD4(-) CD8(-) (double negative, DN) cKit(+) CD25(+) thymocytes, where TCRβ gene rearrangement is initiated. However, TCRαβ(-) CD8(+) immature single positive thymocytes, an actively cycling intermediate population between the DN and DP stages, are the most radiosensitive cells in the thymus, even though their apoptosis is only partially p53-dependent. Within the DP population, TCRαβ(+) thymocytes that completed TCRα gene recombination are more radioresistant than their TCRαβ(-) progenitors. Finally, we found no correlation between p53 activation and thymocyte sensitivity to radiation-induced apoptosis.Clinical & Experimental Immunology 03/2014; DOI:10.1111/cei.12329 · 3.28 Impact Factor
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ABSTRACT: Diabetes is a growing health care issue, and pre-diabetes has been established as a risk factor for type 2 diabetes. Pre-diabetes is characterized by deregulated glucose control, and elucidating pathways which govern this process is critical. We have identified the WIP1 phosphatase as a regulator of glucose homeostasis. Initial characterization of insulin signaling in WIP1 knockout (WIP1(KO)) MEFs demonstrated reduced insulin-mediated AKT activation. In order to assess the role of WIP1 in glucose homeostasis we performed metabolic analysis on mice on a low fat chow diet (LFD) and high fat diet (HFD). We observed increased expression of pro-inflammatory cytokines in WIP1(KO) MEFs, and WIP1(KO) mice fed a LFD and a HFD. WIP1(KO) mice exhibited glucose intolerance and insulin intolerance on a LFD and HFD. However, the effects of WIP1 deficiency causes different metabolic defect in mice on a LFD and a HFD. WIP1(KO) mice on a LFD develop hepatic insulin resistance, while this is not observed in HFD-fed mice. Mouse body weights and food consumption increase slightly over time in LFD-fed wildtype and WIP1(KO) mice. Leptin levels are increased in LFD-fed WIP1(KO) mice, compared to wildtype. In contrast, HFD-fed WIP1(KO) mice are resistant to HFD-induced obesity, have decreased levels of food consumption, and decreased leptin levels compared to HFD-wildtype mice. WIP1 has been shown to regulate the NF-kΒ pathway, loss of which leads to increased inflammation. We propose that this increased inflammation triggers insulin resistance in WIP1(KO) mice on LFD and HFD.Molecular Endocrinology 11/2014; DOI:10.1210/me.2014-1136 · 4.20 Impact Factor
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ABSTRACT: Wip1 (protein phosphatase Mg (2+)/Mn (2+)-dependent 1D, Ppm1d) is a nuclear serine/threonine protein phosphatase that is induced by p53 following the activation of DNA damage response (DDR) signaling. Ppm1d(-/-) mouse embryonic fibroblasts (MEFs) exhibit premature senescence under conventional culture conditions; however, little is known regarding the role of Wip1 in regulating cellular senescence. In this study, we found that even at a representative physiological concentration of 3% O 2, Ppm1d(-/-) MEFs underwent premature cellular senescence that depended on the functional activation of p53. Interestingly, Ppm1d(-/-) MEFs showed increased H2AX phosphorylation levels without increased levels of reactive oxygen species (ROS) or DNA base damage compared with wild-type (Wt) MEFs, suggesting a decreased threshold for DDR activation or sustained DDR activation during recovery. Notably, the increased H2AX phosphorylation levels observed in Ppm1d(-/-) MEFs were primarily associated with S-phase cells and predominantly dependent on the activation of ATM. Moreover, these same phenotypes were observed when Wt and Ppm1d(-/-) MEFs were either transiently or chronically exposed to low levels of agents that induce replication-mediated double-stranded breaks. These findings suggest that Wip1 prevents the induction of cellular senescence at physiological oxygen levels by attenuating DDR signaling in response to endogenous double-stranded breaks that form during DNA replication.Cell cycle (Georgetown, Tex.) 01/2014; 13(6). DOI:10.4161/cc.27920 · 5.01 Impact Factor