Search Results

You are looking at 1 - 6 of 6 items for

  • Author: Heike Biebermann x
Clear All Modify Search
Free access

Gunnar Kleinau, Anne Müller and Heike Biebermann

More than 800 different human membrane-spanning G-protein-coupled receptors (GPCRs) serve as signal transducers at biological barriers. These receptors are activated by a wide variety of ligands such as peptides, ions and hormones, and are able to activate a diverse set of intracellular signaling pathways. GPCRs are of central importance in endocrine regulation, which underpins the significance of comprehensively studying these receptors and interrelated systems. During the last decade, the capacity for multimerization of GPCRs was found to be a common and functionally relevant property. The interaction between GPCR monomers results in higher order complexes such as homomers (identical receptor subtype) or heteromers (different receptor subtypes), which may be present in a specific and dynamic monomer/oligomer equilibrium. It is widely accepted that the oligomerization of GPCRs is a mechanism for determining the fine-tuning and expansion of cellular processes by modification of ligand action, expression levels, and related signaling outcome. Accordingly, oligomerization provides exciting opportunities to optimize pharmacological treatment with respect to receptor target and tissue selectivity or for the development of diagnostic tools. On the other hand, GPCR heteromerization may be a potential reason for the undesired side effects of pharmacological interventions, faced with numerous and common mutual signaling modifications in heteromeric constellations. Finally, detailed deciphering of the physiological occurrence and relevance of specific GPCR/GPCR–ligand interactions poses a future challenge. This review will tackle the aspects of GPCR oligomerization with specific emphasis on family A GPCRs involved in endocrine regulation, whereby only a subset of these receptors will be discussed in detail.

Free access

Anita Kinne, Stephan Roth, Heike Biebermann, Josef Köhrle, Annette Grüters and Ulrich Schweizer

Mutations in the gene encoding the thyroid hormone transporter, monocarboxylate transporter 8 (MCT8), underlie severe mental retardation. We wanted to understand the functional consequences of a series of missense mutations in MCT8 in order to identify therapeutic options for affected patients. We established cell lines stably expressing 12 MCT8 variants in JEG1 and MDCK1 cells. The cell lines were characterized according to MCT8 mRNA and protein expression, tri-iodothyronine (T3) transport activity, substrate K M characteristics, surface expression, and responsiveness to T3 preincubation and chemical chaperones. Functional activities of ins235V and L568P MCT8 mutants depend on the cell type in which they are expressed. These mutants and R271H exhibited considerable transport activity when present at the cell surface as verified by surface biotinylation and kinetic analysis. Most mutants, however, were inactive in T3 transport even when present at the cell surface (e.g. S194F, A224V, ΔF230, L512P). Preincubation of G558D with T3 increased T3 uptake in MDCK1 cells to a small, but significant, extent. Chemical chaperones were ineffective. The finding that the cell type determines surface expression and T3 transport activities of missense mutants in MCT8 may be important to understand phenotypic variability among carriers of different mutations. In particular, the clinical observation that the severity of derangements of thyroid hormone levels does not correlate with mental impairments of the patients may be based on different residual activity of mutant MCT8 in different cell types.

Free access

Christer M Bäck, Stefanie Stohr, Eva A M Schäfer, Heike Biebermann, Ingrid Boekhoff, Andreas Breit, Thomas Gudermann and Thomas R H Büch

Metallothioneins (MTs) are cytoprotective proteins acting as scavengers of toxic metal ions or reactive oxygen species. MTs are upregulated in follicular thyroid carcinoma and are regarded as a marker of thyroid stress in Graves' disease. However, the mechanism of MT regulation in thyrocytes is still elusive. In other cellular systems, cAMP-, calcium-, or protein kinase C (PKC)-dependent signaling cascades have been shown to induce MT expression. Of note, all of these three pathways are activated following the stimulation of the TSH receptor (TSHR). Thus, we hypothesized that TSH represents a key regulator of MT expression in thyrocytes. In fact, TSHR stimulation induced expression of MT isoform 1X (MT1X) in human follicular carcinoma cells. In these cells, Induction of MT1X expression critically relied on intact Gq/11 signaling of the TSHR and was blocked by chelation of intracellular calcium and inhibition of PKC. TSHR-independent stimulation of cAMP formation by treating cells with forskolin also led to an upregulation of MT1X, which was completely dependent on PKA. However, inhibition of PKA did not affect the regulation of MT1X by TSH. As in follicular thyroid carcinoma cells, TSH also induced MT1 protein in primary human thyrocytes, which was PKC dependent as well. In summary, these findings indicate that TSH stimulation induces MT1X expression via Gq/11 and PKC, whereas cAMP–PKA signaling does not play a predominant role. To date, little has been known regarding cAMP-independent effects of TSHR signaling. Our findings extend the knowledge about the PKC-mediated functions of the TSHR.

Free access

Carolin L Piechowski, Anne Rediger, Christina Lagemann, Jessica Mühlhaus, Anne Müller, Juliane Pratzka, Patrick Tarnow, Annette Grüters, Heiko Krude, Gunnar Kleinau and Heike Biebermann

Obesity is one of the most challenging global health problems. One key player in energy homeostasis is the melanocortin-4 receptor (MC4R), which is a family A G-protein-coupled receptor (GPCR). It has recently been shown that MC4R has the capacity to form homo- or heterodimers. Dimerization of GPCRs is of great importance for signaling regulation, with major pharmacological implications. Unfortunately, not enough is yet known about the detailed structural properties of MC4R dimers or the functional consequences of receptor dimerization. Our goal, therefore, was to explore specific properties related to MC4R dimerization. First, we aimed to induce the dissociation of dimers to monomers and to compare the functional parameters of wild-type and MC4R variants. To inhibit homodimerization, we designed MC4R chimeras with the cannabinoid-1 receptor, a receptor that does not interact with MC4R. Indeed, we identified several substitutions in the intracellular loop 2 (ICL2) and adjacent regions of transmembrane helix 3 (TMH3) and TMH4 that lead to partial dimer dissociation. Interestingly, the capacity for signaling activity was generally increased in these MC4R variants, although receptor expression remained unchanged. This increase in activity for dissociated receptors might indicate a link between receptor dimerization and signaling capacity. Moreover, dimer dissociation was also observed in a naturally occurring activating MC4R mutation in ICL2. Taken together, this study provides new information on the structural prerequisites for MC4R dimerization and identifies an approach to induce the dissociation of MC4R dimers. This might be useful for further investigation of pharmacological properties.

Free access

Jana Fischer, Gunnar Kleinau, Anne Müller, Peter Kühnen, Denise Zwanziger, Anita Kinne, Maren Rehders, Lars C Moeller, Dagmar Führer, Annette Grüters, Heiko Krude, Klaudia Brix and Heike Biebermann

The monocarboxylate transporter 8 (MCT8) is a member of the major facilitator superfamily (MFS). These membrane-spanning proteins facilitate translocation of a variety of substrates, MCT8 specifically transports iodothyronines. Mutations in MCT8 are the underlying cause of severe X-linked psychomotor retardation. At the molecular level, such mutations led to deficiencies in substrate translocation due to reduced cell-surface expression, impaired substrate binding, or decreased substrate translocation capabilities. However, the causal relationships between genotypes, molecular features of mutated MCT8, and patient characteristics have not yet been comprehensively deciphered. We investigated the relationship between pathogenic mutants of MCT8 and their capacity to form dimers (presumably oligomeric structures) as a potential regulatory parameter of the transport function of MCT8. Fourteen pathogenic variants of MCT8 were investigated in vitro with respect to their capacity to form oligomers. Particular mutations close to the substrate translocation channel (S194F, A224T, L434W, and R445C) were found to inhibit dimerization of MCT8. This finding is in contrast to those for other transporters or transmembrane proteins, in which substitutions predominantly at the outer-surface inhibit oligomerization. Moreover, specific mutations of MCT8 located in transmembrane helix 2 (del230F, V235M, and ins236V) increased the capacity of MCT8 variants to dimerize. We analyzed the localization of MCT8 dimers in a cellular context, demonstrating differences in MCT8 dimer formation and distribution. In summary, our results add a new link between the functions (substrate transport) and protein organization (dimerization) of MCT8, and might be of relevance for other members of the MFS. Finally, the findings are discussed in relationship to functional data combined with structural–mechanistical insights into MCT8.

Free access

Juliane Dinter, Jessica Mühlhaus, Simon Friedrich Jacobi, Carolin Leonie Wienchol, Maxi Cöster, Jaroslawna Meister, Carolin Stephanie Hoefig, Anne Müller, Josef Köhrle, Annette Grüters, Heiko Krude, Jens Mittag, Torsten Schöneberg, Gunnar Kleinau and Heike Biebermann

Most in vivo effects of 3-iodothyronamine (3-T1AM) have been thus far thought to be mediated by binding at the trace amine-associated receptor 1 (TAAR1). Inconsistently, the 3-T1AM-induced hypothermic effect still persists in Taar1 knockout mice, which suggests additional receptor targets. In support of this general assumption, it has previously been reported that 3-T1AM also binds to the α-2A-adrenergic receptor (ADRA2A), which modulates insulin secretion. However, the mechanism of this effect remains unclear. We tested two different scenarios that may explain the effect: the sole action of 3-T1AM at ADRA2A and a combined action of 3-T1AM at ADRA2A and TAAR1, which is also expressed in pancreatic islets. We first investigated a potential general signaling modification using the label-free EPIC technology and then specified changes in signaling by cAMP inhibition and MAPKs (ERK1/2) determination. We found that 3-T1AM induced Gi/o activation at ADRA2A and reduced the norepinephrine (NorEpi)-induced MAPK activation. Interestingly, in ADRA2A/TAAR1 hetero-oligomers, application of NorEpi resulted in uncoupling of the Gi/o signaling pathway, but it did not affect MAPK activation. However, 3-T1AM application in mice over a period of 6 days at a daily dose of 5 mg/kg had no significant effects on glucose homeostasis. In summary, we report an agonistic effect of 3-T1AM on the ADRA2A-mediated Gi/o pathway but an antagonistic effect on MAPK induced by NorEpi. Moreover, in ADRA2A/TAAR1 hetero-oligomers, the capacity of NorEpi to stimulate Gi/o signaling is reduced by co-stimulation with 3-T1AM. The present study therefore points to a complex spectrum of signaling modification mediated by 3-T1AM at different G protein-coupled receptors.