The Essential Role of MEKK3 Signaling in Angiotensin II-induced Calcineurin/Nuclear Factor of Activated T-cells Activation

The nuclear factor of activated T-cells (NFATp) is a phosphorylated transcription factor that resides in the cytoplasm of unactivated T-cells. T-cell activation results in the activation of the phosphatase calcineurin (CaN), which leads to the dephosphorylation and subsequent nuclear localization of NFATp. We have investigated the role of kinases in the phosphorylation state and subcellular localization of NFATp. The phosphorylation state and nuclear/cytoplasmic location of NFATp were determined in unstimulated murine HT-2 cells treated with a panel of kinase inhibitors. Two of the seven kinase inhibitors, staurosporine (St) and bisindolylmaleimide I (BI), resulted in the dephosphorylation and nuclear localization of NFATp. These St-induced effects were inhibited by pretreatment with FK506, indicating that CaN activity was required for the observed effects on NFATp. Treatment of cells with ionomycin resulted in NFATp dephosphorylation and nuclear localization. Removal of ionomycin from the cells resulted in the reappearance of phosphorylated NFATp in the cytosol. St and BI also inhibited the re-accumulation of NFATp in the cytoplasm and its re-phosphorylation after ionomycin removal. The re-accumulation of NFATp in the cytosol after ionomycin withdrawal was shown to be energy- and temperature-dependent. Taken together, these results suggest that in unstimulated cells NFATp is actively maintained in the cytoplasm by kinases acting in opposition to basal CaN activity.

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Mice Lacking the Calcineurin Inhibitor Rcan2 Have an Isolated Defect of Osteoblast Function

Endocrinology ◽

10.1210/en.2011-1814 ◽

2012 ◽

Vol 153 (7) ◽

pp. 3537-3548 ◽

Cited By ~ 14

Author(s):

J. H. Duncan Bassett ◽

John G. Logan ◽

Alan Boyde ◽

Moira S. Cheung ◽

Holly Evans ◽

...

Keyword(s):

T Cells ◽

Bone Development ◽

Skeletal Development ◽

Dominant Negative ◽

Nuclear Factor ◽

Activated T Cells ◽

Osteoblast Function ◽

And Function

Calcineurin-nuclear factor of activated T cells signaling controls the differentiation and function of osteoclasts and osteoblasts, and regulator of calcineurin-2 (Rcan2) is a physiological inhibitor of this pathway. Rcan2 expression is regulated by T3, which also has a central role in skeletal development and bone turnover. To investigate the role of Rcan2 in bone development and maintenance, we characterized Rcan2−/− mice and determined its skeletal expression in T3 receptor (TR) knockout and thyroid-manipulated mice. Rcan2−/− mice had normal linear growth but displayed delayed intramembranous ossification, impaired cortical bone formation, and reduced bone mineral accrual during development as well as increased mineralization of adult bone. These abnormalities resulted from an isolated defect in osteoblast function and are similar to skeletal phenotypes of mice lacking the type 2 deiodinase thyroid hormone activating enzyme or with dominant-negative mutations of TRα, the predominant TR isoform in bone. Rcan2 mRNA was expressed in primary osteoclasts and osteoblasts, and its expression in bone was differentially regulated in TRα and TRβ knockout and thyroid-manipulated mice. However, in primary osteoblast cultures, T3 treatment did not affect Rcan2 mRNA expression or nuclear factor of activated T cells c1 expression and phosphorylation. Overall, these studies establish that Rcan2 regulates osteoblast function and its expression in bone is regulated by thyroid status in vivo.

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Human T-Cell Lymphotropic Virus Type 1 p12I Expression Increases Cytoplasmic Calcium To Enhance the Activation of Nuclear Factor of Activated T Cells

Journal of Virology ◽

10.1128/jvi.76.20.10374-10382.2002 ◽

2002 ◽

Vol 76 (20) ◽

pp. 10374-10382 ◽

Cited By ~ 61

Author(s):

Wei Ding ◽

Björn Albrecht ◽

Robert E. Kelley ◽

Natarajan Muthusamy ◽

Seung-Jae Kim ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Calcium Release ◽

Nuclear Factor ◽

Cytoplasmic Calcium ◽

Activated T Cells ◽

Human T Cell ◽

Nfat Activation

ABSTRACT Human T-cell lymphotropic virus type 1 (HTLV-1) establishes persistent infection and is associated with lymphoproliferative or neurodegenerative diseases. As a complex retrovirus, HTLV-1 contains typical structural and enzymatic genes, as well as regulatory and accessory genes encoded in the pX region. The early events necessary for HTLV-1 to establish infection in lymphocytes, its primary target cells, remain unresolved. Recent studies have demonstrated the importance of regulatory and accessory gene products in determining this virus-host interaction. Among these, pX open reading frame I, which encodes two proteins, p12I and p27I, is required for establishing persistent infection in vivo and for infection in quiescent primary lymphocytes. In addition, p12I localizes in the endoplasmic reticulum (ER) and cis-Golgi apparatus and associates with a calcium binding protein, calreticulin. We recently reported that p12I expression induces the calcium-responsive T-cell transcription factor, nuclear factor of activated T cells (NFAT), in the presence of phorbol ester activation. Based on these studies, we hypothesize that p12I may modulate calcium release from the ER. Here, we report that p12I expression increases basal cytoplasmic calcium and concurrently diminishes calcium available for release from the ER stores. Overexpression of calreticulin, a calcium buffer protein, blocked p12I-mediated NFAT activation independently of its ability to bind p12I. Chemical inhibition studies using inhibitors of inositol 1,4,5-triphosphate receptor and calcium release-activated calcium channels suggest that inositol 1,4,5-triphosphate receptor in the ER membrane and calcium release-activated calcium channels in the plasma membrane contribute to p12I-mediated NFAT activation. Collectively, our results are the first to demonstrate the role of p12I in elevating cytoplasmic calcium, an antecedent to T-cell activation, and further support the important role of this accessory protein in the early events of HTLV-1 infection.

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