Introduction
The thyroid gland is an essential gland found in various vertebrates, and has thyroid follicles (comprised of thyroid epithelia cells), which are supplied by a dense network of blood capillaries. The thyroid tissue’s main function is the biosynthesis and secretion of the thyroid hormone, which is an essential biological molecule existing in either of two forms, T3 and T4. The activity of the thyroid gland is controlled through the neuroendocrine system, which involves the hypothalamus and the secretory actions of the pituitary gland. TSH plays a key role in stimulating the thyroxin-binding globulin (TGB) and the uptake of blood iodine into the thyroid gland. The hormone (TSH) also stimulates peroxide production via the pentose-phosphate pathway, activating enzyme peroxidase, besides the colloidal efflux, which is mediated by the Cl-/I- and Na+ /I- co-transporters. The TSH activity also enhances synthesis of the sodium iodine symporter (NIS), which is incorporated onto the plasma membrane, playing an essential part in the thyroid regulation. Optimal regulation is important to ensure a homeostatic balance in the blood levels of the thyroid hormone, and imbalances in the hormone’s metabolism results in a variety of medical health conditions, such as thyroidism and goiter. This paper explores the regulation of thyroid hormone, discussing the various mechanisms that are involved, particularly the plasma binding proteins and the activity of MCT-8 (monocarboxylate transporter-8).
Effects of a Competitive Binding Molecule on the Plasma Binding Proteins
Plasma membrane binding transporter proteins are central to the thyroid regulation mechanism. To this end, for example, if an individual lacks the plasma binding proteins, a negative feedback is sent to the hypothalamus and pituitary gland, which respond by lowering the production and release of TSH and TRH, while a positive feedback due to high levels of the plasma proteins leads to long term effects characterized by degradation of T3/T4 and a decrease in metabolic activity. When a biomolecule other than the thyroid hormone binds to the plasma transporter proteins, it is observed that the thyroid hormone levels in the blood decrease (Gayrard et al. 2011). This phenomenon points to the essential role played by the plasma binding proteins in substrate recognition and translocation of the said hormone across the plasma membrane. The binding of the competitive molecule on the thyroid binding site displaces the hormone, which implies that it (the hormone) is not transported optimally; hence the negative changes in thyroid levels in the blood plasma. This is especially important clinically, since a foreign molecule which is capable of competing with thyroid hormone for the binding sites on the transporter proteins could impair the thyroid regulatory mechanism, leading to possible health complications.
According to a study conducted to investigate the effects of a competitive xenobiotic on the levels of the total and free plasma thyroid hormone, the xenobiotic has a dual impact on the blood thyroid hormone levels (Gayrard et al. 2011). The dual effects include an increase in total thyroid hormone plasma clearance, and an increase in free thyroid hormone in vitro. The interference with the intrinsic thyroid hormone clearance results in a significant impact as far as thyroid regulation is concerned. The study results indicate that an administration of a xenobiotic (phenylbutazone) led to a 51% decrease in total plasma T3 concentrations within three (3) hours, while free plasma T3 concentration dropped by 57. 4% (Gayrard et al. 2011). These findings underscore the role of binding transporter proteins in thyroid hormone regulation, since the binding sites are the reactive centers of the protein molecules.
The Role of PTTG-Binding Factor (PBF) in Thyroid Regulation
The PTTG-binding factor (PBF) has been shown to play a key role in the regulation of the thyroid hormone in the body. PBF is a proto-oncogene which research findings have linked to endocrine cancer, and is overexpressed in a variety of tumors, including the breast, pituitary, and thyroid (Smith et al., 2012). Moreover, the sodium iodide symporter (NIS) and pituitary tumor-transforming gene-binding factor (PBF; PTTG1IP) play a key role in regulating thyroid hormone activity in the body. The sodium iodide symporter takes iodine from the blood into the thyroid, where the biosynthesis of thyroid hormone occurs (Smith et al., 2012). Afterwards, the monocarboxylate transporter-8, whose concentration and activity are affected by the PBF, facilitates secretion of the thyroid hormone from the thyroid gland.
An experimental study conducted to determine if PTTG-binding factor (PBF) affected the activity of MCT-8 showed that the PBF binds to MCT-8, which leads to a decrease in the amount of MCT-8 on the blood plasma membrane (Smith et al., 2012). In the said study, the researchers conducted comprehensive experiments involving co-immunoprecipitation assays, immune-fluorescent staining and cell surface biotinylation, and took an array of measurements and analyses. The experimental results and subsequent analyses showed that PBF binds to both MCT-8 and NIS, altering the subcellular localization of MCT-8 (Smith et al., 2012). The researchers also showed that MCT-8 could co-localize with PBF in vitro (see Figure 1). The research showed a significant decrease in thyroid hormones levels secreted from the thyroid after PBF binding on the MCT-8, with a mean of 2.25 compared to 5.54 in mice where no PBF had been included. As already mentioned, MCT-8 are important in mediating the secretion of the thyroid hormone from the thyroid gland. It follows, therefore, that when PBF binds the MCT-8, it decreases its expression on the plasma membrane, resulting in a reduction in the secretion of the thyroid hormone, hence the decrease in the thyroid weight that the aforementioned study reports.
Furthermore, PBF was shown to decrease the activity of TSH, after the researchers demonstrated that while the PBF-free mice showed a mean increase of 1.59 in TSH-stimulated T4 secretion, no significant increases was observed in the PBF-treated mice (Smith et al., 2012). In addition, administration of TSH resulted in a significant decrease in the amount of serum T4 in the PBF-treated mice than the PBF-free ones. Once more, these findings can be linked to the reduction in MCT-8 function and hence TH secretion, which is associated with PBF activity on the said cells (Smith et al., 2012).
Localization and Subcellular Distribution of D3 (Type 3 deiodinase) and MCT8
Closely related to the plasma binding proteins and MCT-8 are the deiodinase enzymes, which play an equally important role in thyroid hormone regulation, by facilitating transport of the thyroid hormone in conjunction with other transporters, such as the NIS (Kallo et al., 2012). To this end, the enzymes regulate the concentration of the thyroid hormone by either degrading T3 by type 3 deiodinase (D3) which results in a decrease in the hormonal levels, or conversion of T4 to T3 by type 2 deiodinase, which leads to an increase in the levels of the thyroid hormone in the blood. The study by Kolla et al. (2012) revealed an interesting distribution of T3-inducible D3 catalytic activity in the brains of rat and humans. In the study, the researchers used their findings to conclude that MCT-8 mediated transport and D3-catalyzed inactivation comprised an important regulatory pathway for thyroid hormone. The researchers used triple-labeling immunofluorescence and immune-electronmiscroscopy, bimolecular-fluorescence-complementation and double-labeling immune-fluorescent confocal microscopy techniques to detect the presence of MCT-8 on the surface of hypophysiotropic terminals and D3 in GT1-7 cells and the median eminence of the rats (Kolla et al., 2013). It is worth noting that the researcher used 20 adult, male rats weighting between 220-250 grams to do their experiment. Based on the statistical parameters of the reagents, instruments and the resultant findings, it may be seen that the study findings and conclusions are reliably useful as reported.
The localization and subcellular distribution of the type 3 deiodinase and MCT-B molecules points to the hypothalamic activity in the regulation of the thyroid hormone. As already implied, changes in T4 and T3 levels initiate a positive or negative feedback to the hypothalamus and the pituitary gland, which in turn initiate appropriate responses to restore the normal balance in the levels of the thyroid hormone. Since the type 3 deiodinase investigated by the aforementioned research plays a central role in degrading T3 in order to decrease thyroid hormone levels, it may be seen that its distribution and localization in the brain and activation/inactivation availability, as well as its functions’ synchronization with the activity of the MCT-8 comprises a significant part of the neurosecretory system of thyroid hormone regulation (Kolla et al., 2013). It is therefore right that the researchers conclude that the MCT-8 and D3 comprise the novel pathway that regulates thyroid hormone availability in rat and human hypothalamic neurosecretory neurons.
Allosteric Regulation: Thyroxin-binding Globulin (TBG)
Thyroxin-binding globulins also play an important role in thyroid hormone regulation. The release of the hormone from the aforesaid binding protein is regulated by the in and out movement of the reactive center loop of the beta-sheet loop of the molecule (Qi et al. 2011). As it were, the activity of the binding sites of the proteins and their receptors on the plasma membranes is mediated by the reactive moieties within those centers. As such, any modifications that occur alter the binding activity, effectively affecting the sites’ biological activity. What is more, release of the thyroid hormone from the thyroid gland, for example, is dependent on molecular modifications, which involve movement of the reactive center loops within the molecule (Qi et al. 2011). These molecular changes are then transferred to the hormone binding site within the transporter protein. These modifications and movements work to facilitate the recognition and translocation of the thyroid hormone across the plasma membrane.
In the case of the aforementioned binding protein and according to Qi et al. (2011), cleavage of the reactive loop results in its complete insertion into the beta sheet A and a substantial but incomplete decrease in binding affinity in TBG” (p. 16163). The research by Qi et al. (2011) to study allosteric modulation of hormone release from thyroxine and corticosteroid-binding globulins reported that the coordinated movements of the binding protein’s reactive loop and the hormonal binding site facilitate the allosteric regulation of the release of the hormone. Changes in temperature and hormonal concentrations can induce such movements and modifications. Figure 2 shows the allosteric mechanism of hormone binding and release (See Appendix).
The said study involved protein expression, purification, cleavage, crystallization and test of binding affinities. This coupled with the statistical analysis support the conclusions. The hormone release mechanism based on binding proteins movement and molecular modifications is important in thyroid hormone translocation in the body. Once the plasma binding protein such as TGB and NIS and MCT-8 have bound to the plasma membrane and moved across it, the said modifications must take place in order to release the hormone into the blood system.
The Role of His192 in Thyroid Hormone Regulation
As aforesaid, MCT-8 is among the most important protein transporters that bind to the plasma membrane to facilitate trafficking of thyroid hormones across the plasma membrane. Research has demonstrated that in addition to the charged lysine, glutamate, aspartate and arginine residues, histidine residues are involved in the binding proteins’ activity in substrate recognition and translocation. For these residues to exert their effect, they interact with the proteins (in this case MCT-8) within its functional domains. To this end, a study by Stefan et al. (2013), which sought to study the role of His192 in the human thyroid hormone transporter MCT-8, revealed that the residue bolstered the binding protein’s substrate recognition and translocation, implying that it played a key role in the thyroid regulatory mechanism. In their study, the researchers found that His192, located in the border of the extracellular loop (ECL) 1 and trans-membrane domain (TMD) 1, was important for efficient thyroid hormone transport and concluded as such.
The study’s conclusion, which is almost certainly central to understanding the functional domains within the MCT-8, was arrived at following a structured experiment in which site-directed mutagenesis of several His residues including H192A, H260A and H450A and analysis of the impacts of the His-modifying reagent diethylpyrocarbonate (DPEC) were performed. The results indicated that His 192 is sensitive to DEPC modifications and is possibly located near a putative recognition site within the MCT-8 protein (Stefan et al., 2013). Whereas H192A mutations decreases T3 uptake by 20% to 30%, H260A and H450A mutations did not inhibit thyroid hormone uptake (Stefan et al. 2013). The experiment showed no significant decrease in MCT-8-mediated hormone efflux even after incubation with DPEC. These findings led to the conclusion that T3 AND T4 protected the His192 from being modified by DPEC. The assertion that His residues within MCT-8 are involved in thyroid hormone transport is important, especially because the activity of the binding protein is affected by its definition of functional domains, which would be affected by residues such as His among others (Stefan et al. 2013). The effect underscores the role of binding proteins in the regulation of thyroid hormone in the blood, as it is shown to affect T3 and T4 uptake and efflux.
Conclusion
In conclusion, optimal regulation of thyroid hormone is important, since imbalances in the hormone’s metabolism is associated with various medical conditions. The thyroid regulatory mechanism includes the plasma membrane binding transporter proteins that bind, cross the membrane and release the hormone into the bloodstream, the hypothalamic systems which also involves the pituitary gland, and the MCT-8 molecules. Multiple research findings have shown that the functionality of MCT-8 and the plasma proteins is affected by such factors as competitive binding molecules and the pituitary tumor-transforming gene-binding factor. Deiodinases and thyroxin-binding globulins also play a central role in modulating the thyroid hormone to ensure that its blood levels remain within the homeostatic standards. More research is imperative to bolster the current understanding of the functional and regulatory mechanisms of thyroid hormone.
Works Cited
Gayrard, Veronique, Picard-Hagen Nicole, Viguie Catherine, Toutain Pierre-Louis. Competitive
Binding to plasma thyroid hormone transport proteins and thyroid disruption by phenylbutazone used as a probe. General and Comparative Endocrinology, 174 (2011): 225-231.
Kallo, Imre, et al. A Novel Pathway Regulates Thyroid Hormone Availability in Rat and Human
Hypothalamic. PLoS ONE, 7.6 (2012): e37860. doi:10.1371/journal.pone.0037860.
Neurosecretory Neurons
Qi, Xiaoqiang, et al. Allosteric Modulation of Hormone Release from Thyroxine and
Corticosteroid-binding Globulins. Journal of Biological Chemistry, 286. 18 (2011): 16163-16173.
Smith, V. E., et al. PTTG-Binding Factor (PBF) Is a Novel Regulator of the Thyroid Hormone
Transporter MCT8. Endocrinology, 153.7 (2012): 3526-3536.
Stefan, Groenweg, Elaine Lima C.S., Edward Visser W., Robin Peeters P., and Theo Visser, J.
Importance of His192 in the Human Thyroid Hormone Transporter MCT8 for Substrate Recognition. Endocrinology, 154.7 (2013): 2525-2532.
Appendix
Figure 1: The allosteric mechanism of hormone binding and release. Native CBG (A) has a fully exposed reactive center loop (RCL) with a connecting loop on top of helix D (green, arrow) in a helical conformation. This loop is unwound following partial insertion of the reactive loop as seen in the native TBG structure (B) or full insertion into the central _-sheet A in the reactive loop-cleaved CBG structure (C). D, the changes in flexibility of hD resulting from the dynamic flip-flop movement of the reactive loop are transmitted to the hormone binding pocket.
Source: Qi et al. (2011). Journal of Biological Chemistry. 286. 18: 16164
Figure 2: MCT8 and PBF localization in COS-7 cells. A, Confocal images of MCT8-HA after co-transfection with a VO control into COS-7 cells, and endogenous PBF, both detected by immune-fluorescent analysis. MCT8-HA (green) was located predominantly within the plasma membrane, whereas endogenous PBF (red) was found in the nucleus and within intracellular vesicles. B, Representative fluorescent immunocytochemistry experiments examining staining of PBF (red) and MCT8-HA (green) after transient cotransfection into COS-7 cells using both epifluorescent (i) and confocal (ii) microscopy. PBF was predominantly expressed within cytoplasmic vesicles. MCT8-HA demonstrated increased staining within intracellular vesicles where it strongly colocalized with PBF, as seen in yellow in the merged images.
Source: Smith et al., (2012). Endocrinology, 153: 3530.
