Proteins interact with the cytosolic mineralocorticoid receptor depending on the ligand

2008 ◽  
Vol 295 (1) ◽  
pp. H361-H365 ◽  
Author(s):  
Miriam Weber ◽  
Martin Wehling ◽  
Ralf Lösel
Keyword(s):  
Mineralocorticoid Receptor ◽  
Steroid Receptors ◽  
Interacting Proteins ◽  
Receptor Ligands ◽  
Affinity Isolation ◽  
Proteomics Approach ◽  
Native Electrophoresis ◽  
Monomeric Proteins ◽  
Shock Proteins ◽  
Blue Native

Steroid receptors belonging to the superfamily of nuclear receptors do not exist as single monomeric proteins but mediate their effects by the interaction with numerous other proteins, e.g., cofactors for transcription, but also other proteins involved in cellular signaling. This interaction may be ligand dependent, which explains the differential effects of receptor ligands. Whereas some receptors, e.g., the estrogen receptor, have been studied in great detail, much less is known about proteins interacting with the mineralocorticoid receptor (MR). In this study, we aimed to identify interacting proteins using a proteomics approach involving tagged receptor constructs. After affinity isolation of MR complexes, blue native electrophoresis revealed the presence of several populations of MR complexes differing in size and composition. During the identification of interacting proteins, various heat shock proteins but also several previously undescribed potential interactors were found, including 14-3-3-ε. We also demonstrate here that the cytosolic MR in the presence of detergent interacts in a ligand-selective manner with glucose-regulated protein 78 and propionyl-CoA carboxylase-β precursor, which are found in the unliganded or aldosterone-containing complex but not with spironolactone.

scholarly journals Tim18p Is a New Component of the Tim54p-Tim22p Translocon in the Mitochondrial Inner Membrane

2000 ◽  
Vol 11 (1) ◽  
pp. 103-116 ◽  
Author(s):  
Oliver Kerscher ◽  
Naresh B. Sepuri ◽  
Robert E. Jensen
Keyword(s):  
Growth Defect ◽  
Inner Membrane ◽  
New Members ◽  
Mitochondrial Inner Membrane ◽  
Native Electrophoresis ◽  
Multiple Copies ◽  
New Gene ◽  
Synthetically Lethal ◽  
Blue Native Electrophoresis ◽  
Blue Native

The mitochondrial inner membrane contains two separate translocons: one required for the translocation of matrix-targeted proteins (the Tim23p-Tim17p complex) and one for the insertion of polytopic proteins into the mitochondrial inner membrane (the Tim54p-Tim22p complex). To identify new members of the Tim54p-Tim22p complex, we screened for high-copy suppressors of the temperature-sensitivetim54-1 mutant. We identified a new gene,TIM18, that encodes an integral protein of the inner membrane. The following genetic and biochemical observations suggest that the Tim18 protein is part of the Tim54p-Tim22p complex in the inner membrane: multiple copies of TIM18 suppress thetim54-1 growth defect; thetim18::HIS3 disruption is synthetically lethal with tim54-1; Tim54p and Tim22p can be coimmune precipitated with the Tim18 protein; and Tim18p, along with Tim54p and Tim22p, is detected in an ∼300-kDa complex after blue native electrophoresis. We propose that Tim18p is a new component of the Tim54p-Tim22p machinery that facilitates insertion of polytopic proteins into the mitochondrial inner membrane.

Blue native electrophoresis as a tool for detection of PIP-containing protein complexes in the plasma membranes

2013 ◽  
Vol 60 (4) ◽  
pp. 563-570
Author(s):  
T. A. Shevyreva ◽  
M. S. Piotrovskii ◽  
B. V. Belugin ◽  
I. M. Zhestkova ◽  
M. S. Trofimova
Keyword(s):  
Plasma Membranes ◽  
Protein Complexes ◽  
Native Electrophoresis ◽  
Blue Native Electrophoresis ◽  
Blue Native

scholarly journals Mechanisms of ligand specificity of the mineralocorticoid receptor

2011 ◽  
Vol 213 (1) ◽  
pp. 15-24 ◽  
Author(s):  
Peter J Fuller ◽  
Yizou Yao ◽  
Jun Yang ◽  
Morag J Young
Keyword(s):  
Mineralocorticoid Receptor ◽  
Tissue Specificity ◽  
Steroid Receptors ◽  
Ligand Specificity ◽  
Structural Determinants ◽  
C Terminus ◽  
Agonist And Antagonist ◽  
The Central Nervous System ◽  
Antagonist Binding

The mineralocorticoid receptor (MR) differs from the other steroid receptors in that it responds to two physiological ligands, aldosterone and cortisol. In epithelial tissues, aldosterone selectivity is determined by the activity of 11β-hydroxysteroid dehydrogenase type 2, while in other tissues, including the heart and regions of the central nervous system, cortisol is the primary ligand for the MR where it may act as an antagonist. Clinical trials have demonstrated the potential of MR antagonists in the treatment of cardiovascular disease, though their use has been limited by concurrent hyperkalaemia. In order to better target the MR, an understanding of the structural determinants of tissue- and ligand-specific MR activation is needed. Interactions of the MR have been identified, which exhibit ligand discrimination and/or specificity. These interactions include those of the ligand-binding domain with ligand, with the N-terminal domain and with putative co-regulatory molecules. Agonist and antagonist binding have been characterised using chimeras between the human MR and the glucocorticoid receptor or the zebra fish MR together with molecular modelling. The interaction between the N-terminus and the C-terminus is aldosterone dependent but is unexpectedly antagonised by cortisol and deoxycorticosterone in the human MR. Nuclear receptor-mediated transactivation is critically dependent on, and modulated by, co-regulatory molecules. Proteins that interact with the MR in the presence of either aldosterone or cortisol, but not both, have been identified. The successful identification of ligand-specific interactions of the MR may provide the basis for the development of novel MR ligands with tissue specificity.

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