The Type III TGF-{beta} receptor regulates filopodia formation via a Cdc42-mediated IRSP53: NWASP interaction in epithelial cells
Cell adhesion and migration are tightly controlled by regulated changes in the actin cytoskeleton. Previously we reported that the TGF-β superfamily coreceptor, the type III TGF-β receptor (TβRIII/betaglycan), regulates cell adhesion, migration and invasion and suppresses cancer progression in part, through activation of the small GTPase, Cdc42, and Cdc42-dependent alterations to the actin cytoskeleton. Here we demonstrate that TβRIII specifically promotes filopodial formation and extension in MCF10A and HMEC mammary epithelial cells. Mechanistically, cell surface TβRIII and Cdc42 colocal...
Source: BJ Cell - June 10, 2013 Category: Biochemistry Authors: S Young Oh, E H Knelson, G C Blobe, K Mythreye Tags: BJ Cell Source Type: research
The CPVT-associated RyR2 Mutation G230C Enhances Store Overload-Induced Ca2{+} Release and Destabilizes the NH2-terminal domains
Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited, life-threatening arrhythmogenic disorder. CPVT is caused by delayed afterdepolarizations (DADs) that are induced by spontaneous Ca2+ release during sarcoplasmic reticulum (SR) Ca2+ overload, a process also known as store overload induced Ca2+ release (SOICR). A number of mutations in the cardiac ryanodine receptor (RyR2) are linked to CPVT. Many of these CPVT-associated RyR2 mutations enhance the propensity for SOICR and DADs by sensitizing RyR2 to luminal or luminal/cytosolic Ca2+ activation. Recently, a novel CPVT Ry...
Source: BJ Cell - June 7, 2013 Category: Biochemistry Authors: Y Liu, L Kimlicka, F Hiess, X Tian, R Wang, L Zhang, P P. Jones, F van Petegem, S Chen Tags: BJ Disease Source Type: research
Hsp104 as a key modulator of prion-mediated oxidative stress in Saccharomyces cerevisiae
Maintenance of cellular redox homeostasis forms an important part of the cellular defence mechanism and continued cell viability. Despite extensive studies, the role of the chaperone Hsp104 in propagation of misfolded protein aggregates in the cell and generation of oxidative stress remains poorly understood. Expression of RNQ1-RFP in Saccharomyces cerevisiae cells led to the generation of the prion form of the protein and increased oxidative stress. In this work, we show that disruption of Hsp104 in an isogenic yeast strain led to solubilisation of RNQ1-RFP. This reduced the oxidative stress generated in the cell. The hig...
Source: BJ Cell - June 7, 2013 Category: Biochemistry Authors: K Singh, A A Saleh, A K Bhadra, I Roy Tags: BJ Disease Source Type: research
Spinocerebellar ataxia-13 Kv3.3 potassium channels: Arginine-to-Histidine mutations affect both functional and protein expression on the cell surface
Voltage-gated potassium channel Kv3.3 is the causative gene of spinocerebellar ataxia type 13 (SCA13), an autosomal dominant neurological disorder. The four dominant mutations identified to date cause Kv3.3 channels to be nonfunctional or have altered gating properties in Xenopus oocytes. In the present study, we report that SCA13 mutations affect functional as well as protein expression of Kv3.3 channels in a mammalian cell line. The reduced protein level of SCA13 mutants is caused by a shorter protein half-life, and blocking the ubiquitin-proteasome pathway increases the total protein of SCA13 mutants more than wild type...
Source: BJ Cell - June 5, 2013 Category: Biochemistry Authors: J Zhao, J Zhu, W B Thornhill Tags: BJ Cell Source Type: research
Cell Cycle Regulation of Purine Synthesis by Phosphoribosyl- Pyrophosphate and Inorganic Phosphate
Cells must increase synthesis of purine nucleotides/deoxynucleotides before or during S phase. We found that rates of purine synthesis via the de novo and salvage pathways increased 5.0- and 3.3-fold, respectively, as cells progressed from mid G1 to early S phase. The increased purine synthesis could be attributed to a 3.2-fold increase in intracellular 5-phosphoribosyl-α-1-pyrophosphate (PRPP), a rate-limiting substrate for de novo and salvage purine synthesis. PRPP can be produced by the oxidative and non-oxidative pentose phosphate pathways, and we found a 3.1-fold increase in flow through the non-oxidative pathw...
Source: BJ Cell - June 4, 2013 Category: Biochemistry Authors: A Fridman, A Saha, A Chan, D E Casteel, R B. Pilz, G R Boss Tags: BJ Metabolism Source Type: research
Trafficking defects in PAS domain mutant Kv11.1 channels: roles of reduced domain stability and altered domain-domain interactions
Loss of Kv11.1 potassium channel function is the underlying cause of pathology in long QT syndrome type 2, one of the commonest causes of sudden cardiac death in the young. Previous studies have identified the cytosolic Per-Arnt-Sim (PAS) domain as a hotspot for mutations that cause Kv11.1 trafficking defects. To investigate the underlying basis of this observation we have quantified the effect of mutants on domain folding as well as interactions between the PAS domain and remainder of the channel. Apart from R56Q, all mutants impaired the thermostability of the isolated PAS domain. Six mutants, located in the vicinity of ...
Source: BJ Cell - May 31, 2013 Category: Biochemistry Authors: Y Ke, C Ann Ng, M J Hunter, S A Mann, J Heide, A P Hill, J I Vandenberg Tags: BJ Cell Source Type: research
Pharmacological characteristics of Kv1.1- and 1.2-containing channels are influenced by the stoichiometry and positioning of their {alpha} subunits
Voltage-sensitive neuronal Kv1 channels, composed of 4 a subunits and 4 associated auxiliary β subunits, control neuronal excitability and neurotransmission. Limited information exists on the combinations of a subunit isoforms (i.e. Kv1.1-1.6) or their positions in the oligomers, and how these affect sensitivity to blockers. It is known that tetraethylammonium (TEA) inhibits Kv1.1 channel largely due to binding a critical tyrosine (Y379) in the pore, whereas V381 at the equivalent location in Kv1.2 makes it insensitive. With the eventual aim of developing blockers for therapeutic purposes, Kv1.1 and 1.2 a subunit ge...
Source: BJ Cell - May 31, 2013 Category: Biochemistry Authors: A Al-Sabi, S Kumar Kaza, J Dolly, J Wang Tags: BJ Cell Source Type: research
The poly-serine domain of the lysyl-5 hydroxylase Jmjd6 mediates subnuclear localization
Jmjd6 is an Fe(II) and 2-oxoglutarate (2OG) dependent oxygenase that catalyses hydroxylation of lysine residues in proteins involved in pre-mRNA splicing. Jmjd6 plays an essential role in vertebrate embryonic development and has been shown to modulate alternative splicing in response to hypoxic stress. Here we show that an alternatively spliced version of Jmjd6 lacking the polyS domain localises to the nucleolus, predominantly in the fibrillar centre. PolyS domain deleted Jmjd6 also interacts with nucleolar proteins. Furthermore, co-immunoprecipitation experiments and fluorescent 2-hybrid (F2H) assays demonstrate that Jmjd...
Source: BJ Cell - May 21, 2013 Category: Biochemistry Authors: A Wolf, M Mantri, A Heim, U Müller, E Fichter, M M Mackeen, L Schermelleh, G Dadie, H Leonhardt, C Vénien-Bryan, B M Kessler, C J Schofield, A Böttger Tags: BJ Cell Source Type: research
ATP-dependent regulation of actin monomer-filament equilibrium by cyclase-associated protein and ADF/cofilin
Cyclase-associated protein (CAP) is a conserved regulator of actin filament dynamics. In the nematode Caenorhabditis elegans, CAS-1 is an isoform of CAP that is expressed in striated muscle and regulates sarcomeric actin assembly. Here, we report that CAS-2, a second CAP isoform in C. elegans, attenuates the actin-monomer-sequestering effect of actin depolymerizing factor (ADF)/cofilin to increase steady-state levels of actin filaments in an ATP-dependent manner. CAS-2 binds to actin monomers without a strong preference to either ATP- or ADP-actin. CAS-2 strongly enhances exchange of actin-bound nucleotides even in the pre...
Source: BJ Cell - May 14, 2013 Category: Biochemistry Authors: K Nomura, S Ono Tags: BJ Cell Source Type: research
Spermidine promotes adipogenesis of 3T3-L1 cells by preventing interaction of ANP32 with HUR and PP2A
We previously showed that the polyamine spermidine is indispensable for differentiation of 3T3-L1 preadipocytes. Here, we studied the mechanism of spermidine function by using polyamine biosynthesis inhibitor α-difluoromethylornithine in combination with the metabolically stable polyamine analogs γ-methylspermidine or (R,R)-α,ω-bismethylspermine. At the early phase of differentiation, spermidine-depleted 3T3-L1 cells showed decreased translation of transcription factor CCAAT/enhancer binding protein β (C/EBPβ), decreased protein phosphatase 2A (PP2A) activity and increased cytoplas...
Source: BJ Cell - May 14, 2013 Category: Biochemistry Authors: M T Hyvönen, T Koponen, J Weisell, M Pietilä, A R Khomutov, J Vepsäläinen, L Alhonen, T A Keinänen Tags: BJ Cell Source Type: research
skNAC depletion stimulates myoblast migration and perturbs sarcomerogenesis by enhancing calpain 1 and 3 activity
Skeletal and heart muscle specific variant of nascent polypeptide associated complex (skNAC) is a skeletal and heart muscle-specific protein known to be involved in the regulation of sarcomerogenesis. The respective mechanism, however, is largely unknown. Here, we demonstrate that skNAC regulates calpain activity. Specifically, we show that inhibition of skNAC expression leads to enhanced, and overexpression of the skNAC gene to repressed, activity of calpain 1 and – to a lesser extent – calpain 3 in myoblasts. In skNAC siRNA-treated cells, enhanced calpain activity is associated with increased migration rate...
Source: BJ Cell - May 13, 2013 Category: Biochemistry Authors: J Berkholz, A Zakrzewicz, B Munz Tags: BJ Cell Source Type: research
Receptor for activated C-kinase 1 (RACK1) interacts with F-box and WD-repeat domain containing-2 (FBW2) to upregulate glial cell missing-1 (GCM1) stability and placental cell migration and invasion
In this study, we perform tandem affinity purification coupled with mass spectrometry analysis identifying RACK1 as an FBW2-interacting protein. RACK1 is a multifaceted scaffold protein containing seven WD repeats. We demonstrate that the WD repeats in both RACK1 and FBW2 are required for the interaction of RACK1 and FBW2. Furthermore, RACK1 competes with GCM1 for FBW2 and thereby prevents GCM1 ubiquitination, which is also supported by the observation that GCM1 is destabilized in RACK1-knockdown BeWo placental cells. Importantly, RACK1 knockdown leads to decreased expression of the GCM1 target gene high-temperature requir...
Source: BJ Cell - May 7, 2013 Category: Biochemistry Authors: C Wang, H Lo, S Lin, H Chen Tags: BJ Signal Source Type: research
The N-terminal region of two-pore channel 1 regulates trafficking and activation by NAADP
Two-pore channels (TPCs) are NAADP-sensitive Ca2+-permeable ion channels expressed on acidic organelles. Here we show that deletion of the N-terminal region redirects TPC1 to the ER. Introduction of fluorophores at the N-terminus of TPC1 did not affect sub-cellular location but reversibly abolishes NAADP sensitivity. Our data reveal a dual role for the N-terminus in localization and function of TPC1 (Source: BJ Cell)
Source: BJ Cell - May 2, 2013 Category: Biochemistry Authors: D Churamani, R Hooper, T Rahman, E Brailoiu, S Patel Tags: BJ Signal Source Type: research
The proteasome cap RPT5/Rpt5p subunit prevents aggregation of unfolded ricin A chain
The plant cytotoxin ricin enters mammalian cells by receptor-mediated endocytosis, undergoing retrograde transport to the endoplasmic reticulum (ER) where its catalytic A chain (RTA) is reductively separated from the holotoxin to enter the cytosol and inactivate ribosomes. The currently accepted model is that the bulk of ER-dislocated RTA is degraded by proteasomes. We show here that the proteasome has a more complex role in ricin intoxication than previously recognised, that the previously reported increase in sensitivity of mammalian cells to ricin in the presence of proteasome inhibitors simply reflects toxicity of the ...
Source: BJ Cell - April 26, 2013 Category: Biochemistry Authors: P Pietroni, N Vasisht, J P Cook, D M Roberts, J Lord, R Hartmann-Petersen, L M Roberts, R A Spooner Tags: BJ Cell Source Type: research
Phosphorylation of the guanine-nucleotide exchange factor CalDAG-GEFI by protein kinase A regulates Ca2{+}-dependent activation of platelet Rap1b GTPase
In blood platelets, the small GTPase Rap1b is activated by cytosolic Ca2+, and promotes integrin aIIbb3 inside-out activation and platelet aggregation. cAMP is the major inhibitor of platelet function and antagonizes Rap1b stimulation through a mechanism that remains unclear. In this work we demonstrate that the Ca2+-dependent exchange factor for Rap1b, CalDAG-GEFI, is a novel substrate for the cAMP-activated protein kinase A (PKA). CalDAG-GEFI phosphorylation occurred in intact platelets treated with the cAMP-increasing agent forskolin, and was inhibited by the PKA inhibitor H89. Purified recombinant CalDAG-...
Source: BJ Cell - April 19, 2013 Category: Biochemistry Authors: G Francesco Guidetti, D Manganaro, A Consonni, I Canobbio, C Balduini, M Torti Tags: BJ Signal Source Type: research
