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Calcium/calmodulin-dependent kinases can regulate the TSH expression in the rat pituitary

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Abstract

Purpose

The endocrine secretion of TSH is a finely orchestrated process controlled by the thyrotropin-releasing hormone (TRH). Its homeostasis and signaling rely on many calcium-binding proteins belonging to the “EF-hand” protein family. The Ca2+/calmodulin (CaM) complex is associated with Ca2+/CaM-dependent kinases (Ca2+/CaMK). We have investigated Ca2+/CaMK expression and regulation in the rat pituitary.

Methods

The expression of CaMKII and CaMKIV in rat anterior pituitary cells was shown by immunohistochemistry. Cultured anterior pituitary cells were stimulated by TRH in the presence and absence of KN93, the pharmacological inhibitor of CaMKII and CaMKIV. Western blotting was then used to measure the expression of these kinases and of the cAMP response element-binding protein (CREB). TSH production was measured by RIA after time-dependent stimulation with TRH. Cells were infected with a lentiviral construct coding for CaMKIV followed by measurement of CREB phosphorylation and TSH.

Results

Our study shows that two CaM kinases, CaMKII and CaMKII, are expressed in rat pituitary cells and their phosphorylation in response to TRH occurs at different time points, with CaMKIV being activated earlier than CaMKII. TRH induces CREB phosphorylation through the activity of both CaMKII and CaMKIV. The activation of CREB increases TSH gene expression. CaMKIV induces CREB phosphorylation while its dominant negative and KN93 exert the opposite effects.

Conclusion

Our data indicate that the expression of Ca2+/CaMK in rat anterior pituitary are correlated to the role of CREB in the genetic regulation of TSH, and that TRH stimulation activates CaMKIV, which in turn phosphorylates CREB. This phosphorylation is linked to the production of thyrotropin.

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Acknowledgements

We thank N. Furente, for help in setting up experiments during his apprenticeship for B.Sc., AR Means (Duke University Medical Center) for providing constructs and Jiro Kasahara for the kind gift of CaMKIV antibody. Partial financial support was received from Immunocytochemistry Advanced Course, University of Naples Federico II.

Funding

The research was supported by funds of Immunocytochemistry Advanced Course, Federico II University of Naples.

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Authors and Affiliations

  1. Department of Advanced Biomedical Sciences, Medical School, “Federico II” University of Naples, Naples, Italy

    G. G. Altobelli, D. Sorriento & V. Cimini

  2. Department of Histopathology, Imperial College London, Hammersmith Hospital, London, UK

    S. Van Noorden

  3. Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, University of Campania Luigi Vanvitelli, Caserta, Italy

    D. Cimini

  4. Department of Public Health, Medical School, “Federico II” University of Naples, Naples, Italy

    M. Illario

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  1. G. G. Altobelli
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  2. S. Van Noorden
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  3. D. Cimini
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  4. M. Illario
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  5. D. Sorriento
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  6. V. Cimini
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Contributions

GGA and VC conceived the study and designed and performed the experiments. DC and GGA interpreted data. GGA and VC wrote the first draft of the manuscript. DS and MI assisted with important, definitive experiments. SVN revised the manuscript critically for important intellectual content. All authors revised and read the manuscript and approved the submitted final version.

Corresponding authors

Correspondence to G. G. Altobelli or V. Cimini.

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Animals were cared for according to the “Guide for the Care and Use of Laboratory Animals” by the US National Institutes of Health (NIH Publication No. 85.23, revised 1996) and to institutional rules for the care and handling of experimental animals of University of Naples Federico II, Italy.

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Altobelli, G.G., Van Noorden, S., Cimini, D. et al. Calcium/calmodulin-dependent kinases can regulate the TSH expression in the rat pituitary . J Endocrinol Invest 44, 2387–2394 (2021). https://doi.org/10.1007/s40618-021-01545-0

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  • DOI: https://doi.org/10.1007/s40618-021-01545-0

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