scholarly journals Evidence for a Homodimeric Structure of Human Monocarboxylate Transporter 8

Endocrinology ◽  
2009 ◽  
Vol 150 (11) ◽  
pp. 5163-5170 ◽  
Author(s):  
W. Edward Visser ◽  
Nancy J. Philp ◽  
Thamar B. van Dijk ◽  
Wim Klootwijk ◽  
Edith C. H. Friesema ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Psychomotor Retardation ◽  
Monocarboxylate Transporter ◽  
Membrane Association ◽  
Transport Function ◽  
Wild Type ◽  
Transfected Cells ◽  
Physiological Relevance ◽  
Dimerization Process

The human monocarboxylate transporter 8 (hMCT8) protein mediates transport of thyroid hormone across the plasma membrane. Association of hMCT8 mutations with severe psychomotor retardation and disturbed thyroid hormone levels has established its physiological relevance, but little is still known about the basic properties of hMCT8. In this study we present evidence that hMCT8 does not form heterodimers with the ancillary proteins basigin, embigin, or neuroplastin, unlike other MCTs. In contrast, it is suggested that MCT8 exists as monomer and homodimer in transiently and stably transfected cells. Apparently hMCT8 forms stable dimers because the complex is resistant to denaturing conditions and dithiothreitol. Cotransfection of wild-type hMCT8 with a mutant lacking amino acids 267–360 resulted in formation of homo-and heterodimers of the variants, indicating that transmembrane domains 4–6 are not involved in the dimerization process. Furthermore, we explored the structural and functional role of the 10 Cys residues in hMCT8. All possible Cys>Ala mutants did not behave differently from wild-type hMCT8 in protein expression, cross-linking experiments with HgCl2 and transport function. Our findings indicate that individual Cys residues are not important for the function of hMCT8 or suggest that hMCT8 has other yet-undiscovered functions in which cysteines play an essential role.

scholarly journals Minireview: Thyroid Hormone Transporters: The Knowns and the Unknowns

2011 ◽  
Vol 25 (1) ◽  
pp. 1-14 ◽  
Author(s):  
W. Edward Visser ◽  
Edith C. H. Friesema ◽  
Theo J. Visser
Keyword(s):  
Thyroid Hormone ◽  
Organic Anion ◽  
Psychomotor Retardation ◽  
Cellular Level ◽  
Monocarboxylate Transporter ◽  
Causative Role ◽  
Organic Anion Transporting Polypeptide ◽  
Neurological Abnormalities ◽  
Knockout Animals

The effects of thyroid hormone (TH) on development and metabolism are exerted at the cellular level. Metabolism and action of TH take place intracellularly, which require transport of the hormone across the plasma membrane. This process is mediated by TH transporter proteins. Many TH transporters have been identified at the molecular level, although a few are classified as specific TH transporters, including monocarboxylate transporter (MCT)8, MCT10, and organic anion-transporting polypeptide 1C1. The importance of TH transporters for physiology has been illustrated dramatically by the causative role of MCT8 mutations in males with psychomotor retardation and abnormal serum TH concentrations. Although Mct8 knockout animals have provided insight in the mechanisms underlying parts of the endocrine phenotype, they lack obvious neurological abnormalities. Thus, the pathogenesis of the neurological abnormalities in males with MCT8 mutations is not fully understood. The prospects of identifying other transporters and transporter-based syndromes promise an exciting future in the TH transporter field.

scholarly journals Importance of His192 in the Human Thyroid Hormone Transporter MCT8 for Substrate Recognition

Endocrinology ◽  
2013 ◽  
Vol 154 (7) ◽  
pp. 2525-2532 ◽  
Author(s):  
Stefan Groeneweg ◽  
Elaine C. Lima de Souza ◽  
W. Edward Visser ◽  
Robin P. Peeters ◽  
Theo J. Visser
Keyword(s):  
Thyroid Hormone ◽  
Transmembrane Domain ◽  
Substrate Recognition ◽  
Inhibitory Effect ◽  
Psychomotor Retardation ◽  
Site Directed Mutagenesis ◽  
Monocarboxylate Transporter ◽  
Human Thyroid ◽  
Extracellular Loop

Abstract Monocarboxylate transporter 8 (MCT8) facilitates cellular uptake and efflux of thyroid hormone (TH). So far, functional domains within MCT8 are not well defined. Mutations in MCT8 result in severe psychomotor retardation due to impaired neuronal differentiation. One such mutation concerns His192 (H192R), located at the border of transmembrane domain (TMD) 1 and extracellular loop (ECL) 1, suggesting that this His residue is important for efficient TH transport. Here, we studied the role of different His residues, predicted within TMDs or ECLs of MCT8, in substrate recognition and translocation. Therefore, we analyzed the effects of the His-modifying reagent diethylpyrocarbonate (DEPC) and of site-directed mutagenesis of several His residues on TH transport by MCT8. Reaction of MCT8 with DEPC inhibited subsequent uptake of T3 and T4, whereas T3 and T4 efflux were not inhibited. The inhibitory effect of DEPC on TH uptake was prevented in the presence of T3 or T4, suggesting that TH blocks access to DEPC-sensitive residues. Three putative DEPC target His residues were replaced by Ala: H192A, H260A, and H450A. The H260A and H450A mutants showed similar TH transport and DEPC sensitivity as wild-type MCT8. However, the H192A mutant showed a significant reduction in TH uptake and was insensitive to DEPC. Taken together, these results indicate that His192 is sensitive to modification by DEPC and may be located close to a putative substrate recognition site within the MCT8 protein, important for efficient TH uptake.

scholarly journals Genotype-Phenotype Relationship in Patients with Mutations in Thyroid Hormone Transporter MCT8

Endocrinology ◽  
2008 ◽  
Vol 149 (5) ◽  
pp. 2184-2190 ◽  
Author(s):  
Jurgen Jansen ◽  
Edith C. H. Friesema ◽  
Monique H. A. Kester ◽  
Charles E. Schwartz ◽  
Theo J. Visser
Keyword(s):  
Plasma Membrane ◽  
Thyroid Hormone ◽  
Protein Expression ◽  
Elevated Serum ◽  
Monocarboxylate Transporter ◽  
Psychomotor Development ◽  
Substrate Affinity ◽  
Loss Of Function ◽  
Wild Type ◽  
Rt Pcr

Loss-of-function mutations in thyroid hormone transporter monocarboxylate transporter 8 (MCT8) lead to severe X-linked psychomotor retardation and elevated serum T3 levels. Most patients, for example those with mutations V235M, S448X, insI189, or delF230, cannot stand, walk, or speak. Patients with mutations L434W, L568P, and S194F, however, walk independently and/or develop some dysarthric speech. To study the relationship between mutation and phenotype, we transfected JEG3 and COS1 cells with wild-type or mutant MCT8. Expression and function of the transporter were studied by analyzing T3 and T4 uptake, T3 metabolism (by cotransfected type 3 deiodinase), Western blotting, affinity labeling with N-bromoacetyl-T3, immunocytochemistry, and quantitative RT-PCR. Wild-type MCT8 increased T3 uptake and metabolism about 5-fold compared with empty vector controls. Mutants V235M, S448X, insI189, and delF230 did not significantly increase transport. However, S194F, L568P, and L434W showed about 20, 23, and 37% of wild-type activity. RT-PCR did not show significant differences in mRNA expression between wild-type and mutant MCT8. Immunocytochemistry detected the nonfunctional mutants V235M, insI189, and delF230 mostly in the cytoplasm, whereas mutants with residual function were expressed at the plasma membrane. Mutants S194F and L434W showed high protein expression but low affinity for N-bromoacetyl-T3; L568P was detected in low amounts but showed relatively high affinity. Mutations in MCT8 cause loss of function through reduced protein expression, impaired trafficking to the plasma membrane, or reduced substrate affinity. Mutants L434W, L568P, and S194F showed significant residual transport capacity, which may underlie the more advanced psychomotor development observed in patients with these mutations.

scholarly journals Septic shock non-thyroidal illness syndrome causes hypothyroidism and conditions for reduced sensitivity to thyroid hormone

2013 ◽  
Vol 50 (2) ◽  
pp. 255-266 ◽  
Author(s):  
Isabel Castro ◽  
Leah Quisenberry ◽  
Rosa-Maria Calvo ◽  
Maria-Jesus Obregon ◽  
Joaquin Lado-Abeal
Keyword(s):  
Gene Expression ◽  
Septic Shock ◽  
Thyroid Hormone ◽  
Tissue Level ◽  
Binding Activity ◽  
Monocarboxylate Transporter ◽  
Kidney Cortex ◽  
Liver And Kidney ◽  
Response To Stress

Non-thyroidal illness syndrome (NTIS) is part of the neuroendocrine response to stress, but the significance of this syndrome remains uncertain. The aim of this study was to investigate the effect of lipopolysaccharide (LPS)-induced NTIS on thyroid hormone (TH) levels and TH molecular targets, as well as the relationship between septic shock nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) activation and TH receptor β (THRB) gene expression at a multi-tissue level in a pig model. Prepubertal domestic pigs were given i.v. saline or LPS for 48 h. Serum and tissue TH was measured by chemiluminescence and RIA. Expression of THRs and cofactors was measured by real-time PCR, and deiodinase (DIO) activity was measured by enzyme assays. Tissue NF-kB nuclear binding activity was evaluated by EMSA. LPS-treated pigs had decreased TH levels in serum and most tissues. DIO1 expression in liver and kidney and DIO1 activity in kidney decreased after LPS. No changes in DIO2 activity were observed between groups. LPS induced an increase in hypothalamus, thyroid, and liver DIO3 activity. Among the other studied genes, monocarboxylate transporter 8 and THRB were the most commonly repressed in endotoxemic pigs. LPS-induced NF-kB activation was associated with a decrease in THRB gene expression only in frontal lobe, adrenal gland, and kidney cortex. We conclude that LPS-induced NTIS in pigs is characterized by hypothyroidism and tissue-specific reduced TH sensitivity. The role of NF-kB in regulating THRB expression during endotoxemia, if any, is restricted to a limited number of tissues.

scholarly journals Identification of human CCR8 as a CCL18 receptor

2013 ◽  
Vol 210 (10) ◽  
pp. 1889-1898 ◽  
Author(s):  
Sabina A. Islam ◽  
Morris F. Ling ◽  
John Leung ◽  
Wayne G. Shreffler ◽  
Andrew D. Luster
Keyword(s):  
Chemokine Receptor ◽  
Inflammatory Diseases ◽  
Calcium Flux ◽  
Th2 Cells ◽  
Wild Type ◽  
Transfected Cells ◽  
Esophageal Tissue ◽  
Chemokine Ligand ◽  
Normal Controls

The CC chemokine ligand 18 (CCL18) is one of the most highly expressed chemokines in human chronic inflammatory diseases. An appreciation of the role of CCL18 in these diseases has been hampered by the lack of an identified chemokine receptor. We report that the human chemokine receptor CCR8 is a CCL18 receptor. CCL18 induced chemotaxis and calcium flux of human CCR8-transfected cells. CCL18 bound with high affinity to CCR8 and induced its internalization. Human CCL1, the known endogenous CCR8 ligand, and CCL18 competed for binding to CCR8-transfected cells. Further, CCL1 and CCL18 induced heterologous cross-desensitization of CCR8-transfected cells and human Th2 cells. CCL18 induced chemotaxis and calcium flux of human activated highly polarized Th2 cells through CCR8. Wild-type but not Ccr8-deficient activated mouse Th2 cells migrated in response to CCL18. CCL18 and CCR8 were coexpressed in esophageal biopsy tissue from individuals with active eosinophilic esophagitis (EoE) and were present at markedly higher levels compared with esophageal tissue isolated from EoE patients whose disease was in remission or in normal controls. Identifying CCR8 as a chemokine receptor for CCL18 will help clarify the biological role of this highly expressed chemokine in human disease.

scholarly journals Histidines in Potential Substrate Recognition Sites Affect Thyroid Hormone Transport by Monocarboxylate Transporter 8 (MCT8)

Endocrinology ◽  
2013 ◽  
Vol 154 (7) ◽  
pp. 2553-2561 ◽  
Author(s):  
Doreen Braun ◽  
Iva Lelios ◽  
Gerd Krause ◽  
Ulrich Schweizer
Keyword(s):  
Thyroid Hormone ◽  
Ring Structure ◽  
Psychomotor Retardation ◽  
Major Facilitator Superfamily ◽  
Site Directed Mutagenesis ◽  
Monocarboxylate Transporter ◽  
Transport Channel ◽  
Hormone Transport ◽  
Bonding Properties ◽  
Major Facilitator

Abstract Mutations in monocarboxylate transporter 8 (MCT8; SLC16A2) cause the Allan-Herndon-Dudley syndrome, a severe X-linked psychomotor retardation syndrome. MCT8 belongs to the major facilitator superfamily of 12 transmembrane-spanning proteins and transports thyroid hormones across the blood-brain barrier and into neurons. How MCT8 distinguishes thyroid hormone substrates from structurally closely related compounds is not known. The goal of this study was to identify critical amino acids along the transport channel cavity, which participate in thyroid hormone recognition. The fact that T3 is bound between a His-Arg clamp in the crystal structure of the T3 receptor/T3 complex prompted us to investigate whether such a motif might potentially be relevant for T3 recognition in MCT8. We therefore replaced candidate histidines and arginines by site-directed mutagenesis and performed activity assays in MDCK-1 cells and Xenopus oocytes. Histidines were replaced by alanine, phenylalanine, and glutamine to probe for molecular properties like aromatic ring structure and H-bonding properties. It was found that some mutations in His192 and His415 significantly changed substrate transport kinetics. Arg301 at the intracellular end of the substrate channel is at an ideal distance to His415 to participate in a His-Arg clamp and mutation to alanine-abrogated hormone transport. Molecular modeling demonstrates a perfect fit of T3 poised into the substrate channel between His415 and Arg301 and observing the same geometry as in the T3 receptor.

scholarly journals Potential role of Rab4 in the regulation of subcellular localization of Glut4 in adipocytes.

1996 ◽  
Vol 16 (12) ◽  
pp. 6879-6886 ◽  
Author(s):  
M Cormont ◽  
M N Bortoluzzi ◽  
N Gautier ◽  
M Mari ◽  
E van Obberghen ◽  
...  
Keyword(s):  
Cell Surface ◽  
Subcellular Distribution ◽  
Potential Role ◽  
Glucose Transporter ◽  
Small Gtpase ◽  
Wild Type ◽  
Transfected Cells ◽  
Isolated Adipocytes ◽  
Glucose Transporter Glut4

A role for Rab4 in the translocation of the glucose transporter Glut4 induced by insulin has been recently proposed. To study more directly the role of this small GTPase, freshly isolated adipocytes were transiently transfected with the cDNAs of both an epitope-tagged Glut4-myc and Rab4, a system which allows direct measurement of the concentration of Glut4 molecules at the cell surface. When cells were cotransfected with Glut4-myc and Rab4, the concentration of Glut4-myc at the cell surface decreased in parallel with the increased expression of Rab4, suggesting that Rab4 participates in the intracellular retention of Glut4. In parallel, the amount of Rab4 associated with the Glut4-containing vesicles increased. When Rab4 was moderately overexpressed, the number of Glut4-myc molecules recruited to the cell surface in response to insulin was similar to that observed in mock-transfected cells, and thus the insulin efficiency was increased. When Rab4 was expressed at a higher level, the amount of Glut4-myc present at the cell surface in response to insulin decreased. Since the overexpressed protein was predominantly cytosolic, this suggests that the cytosolic Rab4 might complex some factor(s) necessary for insulin action. This hypothesis was strengthened by the fact that Rab4 deltaCT, a Rab4 mutant lacking the geranylgeranylation sites, inhibited insulin-induced recruitement of Glut4-myc to the cell surface, even when moderately overexpressed. Rab3D was without effect on Glut4-myc subcellular distribution in basal or insulin-stimulated conditions. While two mutated proteins unable to bind GTP did not decrease the number of Glut4-myc molecules in basal or insulin-stimulated conditions at the plasma membrane, the behavior of a mutated Rab4 protein without GTPase activity was similar to that of the wild-type Rab4 protein, indicating that GTP binding but not its hydrolysis was required for the observed effects. Altogether, our results suggest that Rab4, but not Rab3D, participates in the molecular mechanism involved in the subcellular distribution of the Glut4 molecules both in basal and in insulin-stimulated conditions in adipocytes.

scholarly journals Molecular aspects of thyroid hormone transporters, including MCT8, MCT10, and OATPs, and the effects of genetic variation in these transporters

2009 ◽  
Vol 44 (1) ◽  
pp. 1-11 ◽  
Author(s):  
Wendy M van der Deure ◽  
Robin P Peeters ◽  
Theo J Visser
Keyword(s):  
Genetic Variation ◽  
Thyroid Hormone ◽  
Organic Anion ◽  
Elevated Serum ◽  
Tissue Level ◽  
Psychomotor Retardation ◽  
Monocarboxylate Transporter ◽  
Organic Anion Transporting Polypeptides ◽  
The Central Nervous System ◽  
Molecular Aspects

Thyroid hormone is a pleiotropic hormone with widespread biological actions. For instance, adequate levels of thyroid hormone are critical for the development of different tissues such as the central nervous system, but are also essential for the regulation of metabolic processes throughout life. The biological activity of thyroid hormone depends not only on serum thyroid hormone levels, but is also regulated at the tissue level by the expression and activity of deiodinases, which activate thyroid hormone or mediate its degradation. In addition, thyroid hormone transporters are necessary for the uptake of thyroid hormone into target tissues. With the discovery of monocarboxylate transporter 8 (MCT8) as a specific thyroid hormone transporter and the finding that mutations in this transporter lead to a syndrome of severe psychomotor retardation and elevated serum 3,3′,5-tri-iodothyronine levels known as the Allan–Herndon–Dudley syndrome, the interest in this area of research has greatly increased. In this review, we will focus on the molecular aspects of thyroid hormone transporters, including MCT8, MCT10, organic anion transporting polypeptides, and the effects of genetic variation in these transporters.

scholarly journals Consequences of Monocarboxylate Transporter 8 Deficiency for Renal Transport and Metabolism of Thyroid Hormones in Mice

Endocrinology ◽  
2010 ◽  
Vol 151 (2) ◽  
pp. 802-809 ◽  
Author(s):  
Marija Trajkovic-Arsic ◽  
Theo J. Visser ◽  
Veerle M. Darras ◽  
Edith C. H. Friesema ◽  
Bernhard Schlott ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Thyroid Hormones ◽  
Psychomotor Retardation ◽  
High Serum ◽  
Monocarboxylate Transporter ◽  
Iodothyronine Deiodinase ◽  
Serum T4 ◽  
Mct8 Deficiency ◽  
Low Serum

Patients carrying inactivating mutations in the gene encoding the thyroid hormone transporting monocarboxylate transporter (MCT)-8 suffer from a severe form of psychomotor retardation and exhibit abnormal serum thyroid hormone levels. The thyroidal phenotype characterized by high-serum T3 and low-serum T4 levels is also found in mice mutants deficient in MCT8 although the cause of these abnormalities is still unknown. Here we describe the consequences of MCT8 deficiency for renal thyroid hormone transport, metabolism, and function by studying MCT8 null mice and wild-type littermates. Whereas serum and urinary parameters do not indicate a strongly altered renal function, a pronounced induction of iodothyronine deiodinase type 1 expression together with increased renal T3 and T4 content point to a general hyperthyroid state of the kidneys in the absence of MCT8. Surprisingly, accumulation of peripherally injected T4 and T3 into the kidneys was found to be enhanced in the absence of MCT8, indicating that MCT8 deficiency either directly interferes with the renal efflux of thyroid hormones or activates indirectly other renal thyroid hormone transporters that preferentially mediate the renal uptake of thyroid hormones. Our findings indicate that the enhanced uptake and accumulation of T4 in the kidneys of MCT8 null mice together with the increased renal conversion of T4 into T3 by increased renal deiodinase type 1 activities contributes to the generation of the low-serum T4 and the increase in circulating T3 levels, a hallmark of MCT8 deficiency.

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