Thyrotropin modifies the synthesis of actin and other proteins during thyroid cell culture

Abstract

Primary cultures of dog thyroid cells have been used to study the effects of thyrotropin on the synthesis of proteins. The cells were cultured for 4 days in serum-free and thyrotropin-free conditions. Thyrotropin was then added for varying periods of time (6-96 h). In the absence of thyrotropin, the cells have an elongated flattened aspect. Exposure to thyrotropin for 6-24 h produces retraction and rounding up of cells whereas cells incubated with thyrotropin for longer periods of time have an epithelial cuboidal shape. After varying periods of culture the cells were labelled with [35S]methionine for 6 h and then analyzed by one-and two-dimensional gel electrophoresis, followed by autoradiography. The results were as follows.1. After exposure to thyrotropin for 32 h and 48 h, the synthesis of about 18 proteins was increased while that of about 14 others was decreased. After 6 h the labelling of three and five of these proteins was already increased or decreased, respectively.2. Some of the proteins whose synthesis is modified in the presence of thyrotropin were identified. Actin synthesis was markedly decreased with a maximum 24-48 h after the addition of thyrotropin. A modification in the ratio between CI and p tubulins was also observed together with very large changes in a group of proteins having both the relative molecular mass (30000 -40000) and the isoelectric points of tropomyosins.3. Forskolin and cholera toxin caused the same qualitative and quantitative changes as thyrotropin; this suggests that the regulation by thyrotropin of the synthesis of several thyroid cell proteins is mediated by CAMP.In conclusion, the data obtained in this work might help to explain the molecular mechanisms by which thyrotropin (and CAMP) triggers the changes in cell shape which occur during thyroid cell culture. They also indicate that one of the main effects of thyrotropin takes place at the level of several proteins which belong to the cytoskeleton and which are involved in the definition of the cytostructure of the thyroid cells.The activating effect of thyroid-stimulating hormone (thyrotropin) in vivo, on both the synthesis and secretion of thyroid hormones, is associated with marked alterations in thyroid cell morphology. Acute thyrotropin administration produces an extension of pseudopod-like process of the apical cell surface into the follicular lumen (see [I] for a review). Under chronic thyrotropin deprivation the cells are flat, whereas hyperstimulation produces hyperplasia of the thyroid gland and induces 'rounded up' cuboid cells [l]. The establishment of functional primary cultures of thyroid cells 12 -41 and of normal established cell lines [5] have permitted the study of the mechanism(s) responsible for the acute changes in cell morphology. Ultramicroscopic and histoimmunological studies have led to the conclusion that acute thyrotropin administration produces a dramatic reorganization of the microfilament system [6, 71. In contrast, little attention has been devoted to the chronic (or trophic) effects of t...

Induction of DNA synthesis in dog thyrocytes in primary culture: Synergistic effects of thyrotropin and cyclic AMP with epidermal growth factor and insulin

“…It stimulates the phosphorylation (Contor et al, unpublished results) and modifies the synthesis of a number of proteins (Passareiro et al, 1985). These effects are mimicked by forskolin or cholera toxin (above references).…”

The dog thyroid primary culture system: a model of the regulation of function, growth and differentiation expression by cAMP and other well-defined signaling cascades

“…In the case of TSH, we have postulated that this event may be associated with the stimulated macropinocytic activity (85). In response to TSH and TPA but not EGF, the synthesis of actin (89,90) and high molecular weight tropomyosin isoforms is inhibited (13). This may result from and prolong the effect of the acute disruption of actin bundles.…”

Differential protein synthesis in the induction of thyroid cell proliferation by thyrotropin, epidermal growth factor or serum

“…The effects of TSH on the labeling of proteins 2-9 had already been observed after a 48-h treatment of such cells [16]. In contrast, EGF and serum alone or in combination specifically increased the labeling of protein 1.…”

“…It is worth mentioning that protein 7 (MI x 39000; PI rn 5 ) is situated in the two-dimensional gel among a group of proteins with MI and isoelectric points of tropomyosins [16,211. It could correspond to the protein termed cyclin or PCNA (MI z 36000; PI z 5), which has been implicated in the control of growth by several workers and which migrates in a similar position in two-dimensional gel electrophoresis [13,22,231.…”