Sarah Dremier

Activation of cyclic AMP-dependent kinase is required but may not be sufficient to mimic cyclic AMP-dependent DNA synthesis and thyroglobulin expression in dog thyroid cells.

Thyrotropin (TSH), via a cyclic AMP (cAMP)-dependent pathway, induces cytoplasmic retractions, proliferation, and differentiation expression in dog thyroid cells. The role of cAMP-dependent protein kinase (PKA) in the induction of these events was assessed by microinjection into living cells. Microinjection of the heat-stable inhibitor of PKA (PKI) inhibited the effects of TSH, demonstrating that activation of PKA was required in this process. Overexpression of the catalytic (C) subunit of PKA brought about by microinjection of the expression plasmid pC alpha ev or of purified C subunit itself was sufficient to mimic the cAMP-dependent cytoplasmic changes and thyroperoxidase mRNA expression but not to induce DNA synthesis and thyroglobulin (Tg) expression. The cAMP-dependent morphological effect was not observed when C subunit was coinjected with the regulatory subunit (RI or RII subunit) of PKA. To mimic the cAMP-induced PKA dissociation into free C and R subunits, the C subunit was coinjected with the regulation-deficient truncated RI subunit (RIdelta1-95) or with wild-type RI or native RII subunits, followed by incubation with TSH at a concentration too low to stimulate the cAMP-dependent events by itself. Although the cAMP-dependent morphology changes were still observed, neither DNA synthesis nor Tg expression was stimulated in these cells. Taken together, these data suggest that in addition to PKA activation, another cAMP-dependent mechanism could exist and play an important role in the transduction of the cAMP signal in thyroid cells.

The visual display of regulatory information and networks

Cell regulation and signal transduction are becoming increasingly complex, with reports of new cross-signalling, feedback, and feedforward regulations between pathways and between the multiple isozymes discovered at each step of these pathways. However, this information, which requires pages of text for its description, can be summarized in very simple schemes, although there is no consensus on the drawing of such schemes. This article presents a simple set of rules that allows a lot of information to be inserted in easily understandable displays.

Search for new cyclic AMP-binding proteins

Today, there is evidence that the cAMP‐dependent kinases (PKA) are not the only intracellular receptors involved in intracellular cAMP signalling in eukaryotes. Other cAMP‐binding proteins have been recently identified, including some cyclic nucleotide‐gated channels and Epac (exchange protein directly activated by cAMP) proteins. All these proteins bind cAMP through conserved cyclic nucleotide monophosphate‐binding domains. However, all putative cAMP‐binding proteins having such domains, as revealed by computer analysis, do not necessarily bind cAMP, indicating that their presence is not a sufficient criteria to predict cAMP‐binding property for a protein.

Eosinophils affect functions of in vitro-activated human CD3-CD4+ T cells

BackgroundThe recent development of eosinophil-targeting agents has raised enthusiasm for management of patients with hypereosinophilic syndromes. Roughly half of anti-IL-5-treated patients with corticosteroid-responsive lymphocytic (L-HES) and idiopathic disease variants can be tapered off corticosteroids. Potential consequences of corticosteroid-withdrawal on clonal expansion of pre-malignant CD3-CD4+ T-cells associated with L-HES are a subject of concern. Indeed, corticosteroid treatment inhibits T-cell activation and may lower blood CD3-CD4+ cell counts. On the other hand, previous studies have shown that eosinophils support CD4 T-cell activation, suggesting that targeted eosinophil depletion may negatively regulate these cells.ObjectivesEffects of eosinophils on CD4 T-cell activation in vitro were investigated as an indirect means of exploring whether treatment-induced eosinophil depletion may affect pathogenic T-cells driving L-HES.MethodsHelper (CD4) T-cells and CD3-CD4+ cells from healthy controls and L-HES patients, respectively, were cultured in vitro in presence of anti-CD3/CD28 or dendritic cells. Effects of eosinophils on T-cell proliferation and cytokine production were investigated.ResultsEosinophils enhanced CD3-driven proliferation of CD4 T-cells from healthy subjects in vitro, while inhibiting TCR-independent proliferation and IL-5 production by CD3-CD4+ T-cells.ConclusionsWhile this study confirms previous work showing that eosinophils support activation of normal helper T-cells, our in vitro findings with CD3-CD4+ T-cells suggest that eosinophil-depletion may favor activation and expansion of this pathogenic lymphocyte subset. With the ongoing development of eosinophil-targeted therapy for various eosinophilic conditions, the indirect consequences of treatment on the underlying immune mechanisms of disease should be investigated in detail in the setting of translational research programs.

Cloning of cDNA Specifically Involved in the Thyroid cAMP Mitogenic Pathway

The activation of the cyclic AMP cascade in dog and human thyroid cells in primary culture induces the expression of differentiated gene expression, hyperfunction and proliferation. These programs are developed simultaneously in quiescent dedifferentiated cells. In this paper the strategy followed by our group to define the genes involved in the cAMP mitogenic cascade is outlined.