Mitochondrial fission mediates ceramide-induced metabolic disruption in skeletal muscle
Ceramide is a sphingolipid that serves as an important second messenger in an increasing number of stress-induced pathways. Ceramide has long been known to affect the mitochondria, resulting in altered mitochondrial physiology. We sought to assess the impact of ceramide on skeletal muscle mitochondrial structure and function. A primary observation was the dramatic division of mitochondria in ceramide-treated cells. This effect is likely a result of increased Drp1 action, as ceramide increased Drp1 expression and Drp1 inhibition prevented mitochondrial fission despite ceramide. Further, we found that ceramide treatment redu...
Source: BJ Cell - September 30, 2013 Category: Biochemistry Authors: M E. Smith, T S. Tippetts, E S. Brassfield, B J. Tucker, A Ockey, A C. Swensen, T S. Anthonymuthu, T D. Washburn, D A. Kane, J T. Prince, B T. Bikman Tags: BJ Metabolism Source Type: research
A TFG-TEC nuclear localization mutant forms complexes with the wild-type TFG-TEC oncoprotein and suppresses its activity
Human extraskeletal myxoid chondrosarcomas (EMCs) are soft tissue tumors characterized by specific chromosomal abnormalities. Recently, a proportion of EMCs were found to harbor a characteristic translocation, t(3;9)(q11-12;q22), involving the TFG (TRK-fused gene) at 3q11-12 and the TEC (Translocated in Extraskeletal Chondrosarcoma) gene at 9q22. The present study used both in vitro and in vivo systems to show that the TFG-TEC protein self-associates, and that this is dependent upon the coiled-coil (CC) domain (aa 97–124), the AF1 domain (aa 275–562), and the DNA-binding domain (DBD) (aa 563–655). The ...
Source: BJ Cell - September 30, 2013 Category: Biochemistry Authors: B Lim, A Kim, H Jun, J Kim Tags: BJ Gene Source Type: research
Complex contributions of fibronectin to initiation and maturation of microfibrils
Fibrillins constitute the backbone of extracellular multi-functional assemblies present in elastic and non-elastic matrices, termed microfibrils. Assembly of fibrillins into microfibrils and their homeostasis is poorly understood and is often compromised in connective tissue disorders such as Marfan syndrome and other fibrillinopathies. Using interaction mapping studies, we demonstrate that fibrillins require the complete gelatin-binding region of fibronectin for interaction, which is comprised of domains FNI6-FNI9. However, the interaction of fibrillin-1 with the gelatin-binding domain of fibronectin is not involved in fi...
Source: BJ Cell - September 27, 2013 Category: Biochemistry Authors: L Sabatier, J Djokic, C Fagotto-Kaufmann, M Chen, D S Annis, D F Mosher, D P Reinhardt Tags: BJ Cell Source Type: research
Defining the interaction of perforin with calcium and the phospholipid membrane
Following its secretion from cytotoxic lymphocytes into the immune synapse, perforin binds to target cell membranes through its Ca2+-dependent C2 domain. Membrane-bound perforin then forms pores that allow passage of pro-apoptopic granzymes into the target cell. Here, structural and biochemical studies reveal that Ca2+ binding triggers a conformational change in the C2 domain that permits four key hydrophobic residues to interact with the plasma membrane. However, contrary to previous suggestions, these movements and membrane binding do not trigger irreversible conformational changes in the pore-forming MACPF...
Source: BJ Cell - September 27, 2013 Category: Biochemistry Authors: D A.K. Traore, A J. Brennan, R H.P. Law, C Dogovski, M A. Perugini, N Lukoyanova, E W.W. Leung, R S. Norton, J A. Lopez, K A. Browne, H Yagita, G J. Lloyd, A Ciccone, S Verschoor, J A. Trapani, J C. Whisstock, I Voskoboinik Tags: BJ Cell Source Type: research
Dual lipidation of the brain-specific Cdc42 isoform regulates its functional properties
Cdc42 is a member of the Rho GTPase family which regulates a variety of cellular activities by controlling actin cytoskeleton and gene expression. Cdc42 is expressed in form of two splice variants. The canonical Cdc42 isoform is prenylated (Cdc42-prenyl), whereas the brain-specific isoform can be palmitoylated (Cdc42-palm). In the present study we have demonstrated palmitoylation of endogenous Cdc42 in rodent and human brains and identified Cys188 and Cys189 as acylation sites of Cdc42-palm. Moreover, we have shown that Cys188 can also be prenylated. Analysis of acylation-deficient mutants revealed that lipidation of Cys18...
Source: BJ Cell - September 23, 2013 Category: Biochemistry Authors: A Wirth, C Chen-Wacker, Y Wu, N Gorinski, M A. Filippov, G Pandey, E Ponimaskin Tags: BJ Cell Source Type: research
In vivo characterisation of the properties of SUMO1-specific monobodies
Monobodies are small recombinant proteins designed to bind with high affinity to target proteins. Monobodies have been generated to mimic the SUMO-interacting motif (SIM) present in many SUMO target proteins but their properties have not been determined in cells. Here, we characterise the properties of two SUMO1 monobodies (hS1MB4 and hS1MB5) in HEK293 and HeLa cells and examine their ability to purify SUMO substrates from cell lines and rat brain. Both hS1MB4 and hS1MB5 compared favourably with commercially available antibodies and were highly selective for binding to SUMO1 over SUMO2/3 in pull-down assays against endogen...
Source: BJ Cell - September 17, 2013 Category: Biochemistry Authors: A Berndt, K A. Wilkinson, M J. Heimann, P Bishop, J M. Henley Tags: BJ Cell Source Type: research
The ubiquitin proteasome system regulates the stability and activity of the glucose sensor glucokinase in pancreatic beta cells
The ubiquitin proteasome system is important to maintain pancreatic beta cell function. Inhibition of the proteasome significantly reduced glucose-induced insulin secretion. Key regulators of the stimulus-secretion cascade seem to be affected by protein misfolding, if the proteasome is down-regulated as recently reported in humans with type 2 diabetes. It remains unknown, however, whether the glucose sensor enzyme glucokinase is involved in this process. A direct interaction between glucokinase and ubiquitin could be shown in vivo by fluorescence resonance energy transfer suggesting regulation of glucokinase by the proteas...
Source: BJ Cell - September 12, 2013 Category: Biochemistry Authors: A Hofmeister-Brix, S Lenzen, S Baltrusch Tags: BJ Cell Source Type: research
Endoplasmic reticulum-associated ubiquitin conjugating enzyme- Ube2j1 is a novel substrate of MAPKAP kinase-2 involved in MK2-mediated TNF{alpha} production
The p38 MAPK/MK2 signalling pathway is a major regulator of stress- and cytokine-induced gene expression at the transcriptional and post-transcriptional level. By phospho-proteomics we identified the ER-associated ubiquitin-conjugating enzyme Ube2j1 as a potential substrate of MK2. We demonstrate that Ube2j1 is phosphorylated in a cytokine-, cytosolic stress- and LPS-induced manner. The cytosolic stress-induced phosphorylation of Ube2j1 proceeds at serine 184, a site previously described to be phosphorylated in response to ER-stress, which is located in a perfect MK2 consensus motif. The cytosolic stress-induced phosphoryl...
Source: BJ Cell - September 11, 2013 Category: Biochemistry Authors: M B Menon, C Tiedje, J Lafera, N Ronkina, T Konen, A Kotlyarov, M Gaestel Tags: BJ Signal Source Type: research
FGT-1 is the major glucose transporter in C. elegans and is central to aging pathways
Caenorhabditis elegans is widely used as a model for investigation of the relationships between aging, nutrient restriction and signalling via the DAF-2 receptor for insulin-like peptides and AGE-1 (PI 3-kinase) but the identity of the glucose transporters that may link these processes is unknown. We unexpectedly find that of the eight putative GLUT-like genes only the two splice variants of one gene have a glucose transport function in an oocyte expression system. We have named this gene (fgt-1 (Facilitated Glucose Transporter, isoform 1). We show that knockdown of fgt-1 RNA leads to loss of glucose transport and reduced ...
Source: BJ Cell - September 11, 2013 Category: Biochemistry Authors: Y Feng, B G Williams, F Koumanov, A J Wolstenholme, G D Holman Tags: BJ Metabolism Source Type: research
{Soft hyphen}{Soft hyphen}Reorientation of the first signal-anchor sequence during potassium channel biogene{Soft hyphen}{Soft hyphen}sis at the Sec61 complex
The majority of the polytopic proteins that are synthesised at the endoplasmic reticulum are integrated co-translationally via the Sec61 translocon, which provides lateral access for their hydrophobic transmembrane regions (TMs) to the phospholipid bilayer. A prolonged association between TMs of the potassium channel subunit, TASK-1, and the Sec61 complex suggests that the ER translocon coordinates the folding/assembly of the TMs present in the nascent chain. The N-terminus of both TASK-1 and Kcv, another potassium channel subunit of viral origin, has access to the N-glycosylation machinery located in the ER lumen, indicat...
Source: BJ Cell - September 10, 2013 Category: Biochemistry Authors: H R Watson, L Wunderley, T Andreou, J Warwicker, S High Tags: BJ Cell Source Type: research
Regulatory T cells and cAMP suppress effector T cells independently of PKA-CREM/ICER: a potential role for Epac
cAMP signaling is both a major pathway as well as a key therapeutic target for inducing immune tolerance and is involved in regulatory T cell (Treg cell) function. To achieve potent immunoregulation, cAMP can act through several downstream effectors. One proposed mechanism is that cAMP-mediated suppression, including immunosuppression by Treg cells, results from activation of PKA leading to the induction of the transcription factor inducible cAMP early repressor (ICER). Here, we examined CD4+CD25- effector T cell (Teff cell) and CD4+CD25+ Treg cell immune responses in cAMP responsive element modulator ...
Source: BJ Cell - September 5, 2013 Category: Biochemistry Authors: A G. Vang, W Housley, H Dong, C Basole, S Z. Ben-Sasson, B E. Kream, P M. Epstein, R B. Clark, S Brocke Tags: BJ Signal Source Type: research
Human junctophilin-2 undergoes a structural rearrangement upon binding phosphatidylinositol (3,4,5) triphosphate {-} the S101R mutation identified in hypertrophic cardiomyopathy obviates this response
In conclusion this study provides new insights into the structural and lipid binding properties of JP2 and how the S101R mutation may impact upon the stability of the dyad organization with the potential to alter JP2-protein interactions regulating Ca2+ cycling. (Source: BJ Cell)
Source: BJ Cell - September 4, 2013 Category: Biochemistry Authors: H J. Bennett, J Bernard Davenport, R F. Collins, A W. Trafford, C Pinali, A Kitmitto Tags: BJ Cell Source Type: research
The synaptotagmin 1 linker may function as an electrostatic zipper that opens for docking but closes for fusion pore opening
Synaptotagmin 1 (Syt1), a major Ca2+ sensor for fast neurotransmitter release, contains tandem Ca2+-binding C2 domains (C2AB), a single transmembrane α-helix, and a highly charged 60-residue-long linker in between. Using the single vesicle docking and content mixing assay we found that the linker region of Syt1 is essential for its two signature functions: Ca2+-independent vesicle docking and Ca2+-dependent fusion pore opening. The linker contains the basic amino acid-rich N-terminal region and the acidic amino acid-rich C-terminal region. When the charge segregation was disrupted, fusion...
Source: BJ Cell - September 4, 2013 Category: Biochemistry Authors: Y Lai, X Lou, Y Jho, T Yoon, Y Shin Tags: BJ Cell Source Type: research
Synthetic Retinoid Am80 Upregulates Apelin Expression by Promoting Interaction of RAR{alpha} with KLF5 and Sp1 in Vascular Smooth Muscle Cells
In this study, we investigated the molecular mechanism of apelin expression regulation by synthetic retinoid Am80 in VSMCs. The results showed that Am80 markedly upregulated apelin mRNA and protein levels in VSMCs. Furthermore, KLF5 and Sp1 cooperatively mediated Am80-induced apelin expression through their direct binding to the TGF-β control element (TCE) on the apelin promoter. Interestingly, upon Am80 stimulation, the retinoic acid receptor alpha (RARα) was recruited to the apelin promoter by interacting with KLF5 and Sp1 prebound to the TCE site of the apelin promoter to form a transcriptional activation ...
Source: BJ Cell - August 30, 2013 Category: Biochemistry Authors: X Lv, B Zheng, S Li, A Han, C Wang, J Shi, X Zhang, Y Liu, Y Li, J Wen Tags: BJ Cell Source Type: research
PKC{delta} as a regulator for TGF-{beta} stimulated connective tissue growth factor production in human hepatocarcinoma (HepG2) cells
In this study, the role of PKCδ in TGF-β1-mediated CTGF expression was investigated using HepG2 cells. TGF-β1 treatment specifically elevated PKCδ activation and CTGF expression. In contrast, blockade of PKCδ by the selective inhibitor, Rottlerin, or by siRNA knockdown significantly reduced TGF-β1-induced CTGF production. The regulatory mechanism was further demonstrated in HepG2 cells whereby TGF-β1-induced PKCδ activation negatively regulated nuclear levels of PPM1A through the RhoA/ROCK pathway. Moreover, we showed that both Smad signaling and the PKCδ pathw...
Source: BJ Cell - August 29, 2013 Category: Biochemistry Authors: S Lee, J Kang, S Choi, O Kwon Tags: BJ Cell Source Type: research
