Latest Articles

TWEAKing death

Ashwell, J. D. — 2008-07-07

Smac mimetics (inhibitor of apoptosis [IAP] antagonists) are synthetic reagents that kill susceptible tumor cells by inducing degradation of cellular IAP (cIAP) 1 and cIAP2, nuclear factor B activation, tumor necrosis factor (TNF) production, TNF receptor 1 occupancy, and caspase-8 activation. In this issue of The Journal of Cell Biology, Vince et al. (see p. ) report remarkable similarities in the events leading to tumor cell death triggered by the cytokine TWEAK (TNF-like weak inducer of apoptosis) and IAP antagonists. Although the mechanistic details differ, a common and necessary feature that is also shared by TNF receptor 2 signaling is reduction in the level of cIAP1 and, in some cases, cIAP2 and TNF receptor-associated factor 2. These findings not only extend our appreciation of how cell death pathways are kept in check in tumors, they reinforce the possible utility of induced cIDE (cIAP deficiency) in the selective elimination of neoplastic cells.

Inhibition of adipogenesis: a new job for the ER Ca2+ pool

Meldolesi, J. — 2008-07-07

Adipogenesis is the process of differentiation of adipocytes from mesenchymal multipotent cells through adipocyte precursors. In this issue, a study by the groups of Opas and Michalak (Szabo, E., Y. Qiu, S. Baksh, M. Michalak, and M. Opas. 2008. J. Cell. Biol. 182:–) demonstrates that this process is repressed by increasing intracellular Ca²⁺, which, in turn, is dependent on the expression of calreticulin, the major Ca²⁺-binding protein of the endoplasmic reticulum lumen.

Myosin-10 and actin filaments are essential for mitotic spindle function

Woolner, S., O'Brien, L. L., Wiese, C., Bement, W. M. — 2008-07-07

Mitotic spindles are microtubule-based structures responsible for chromosome partitioning during cell division. Although the roles of microtubules and microtubule-based motors in mitotic spindles are well established, whether or not actin filaments (F-actin) and F-actin–based motors (myosins) are required components of mitotic spindles has long been controversial. Based on the demonstration that myosin-10 (Myo10) is important for assembly of meiotic spindles, we assessed the role of this unconventional myosin, as well as F-actin, in mitotic spindles. We find that Myo10 localizes to mitotic spindle poles and is essential for proper spindle anchoring, normal spindle length, spindle pole integrity, and progression through metaphase. Furthermore, we show that F-actin localizes to mitotic spindles in dynamic cables that surround the spindle and extend between the spindle and the cortex. Remarkably, although proper anchoring depends on both F-actin and Myo10, the requirement for Myo10 in spindle pole integrity is F-actin independent, whereas F-actin and Myo10 actually play antagonistic roles in maintenance of spindle length.

TWEAK-FN14 signaling induces lysosomal degradation of a cIAP1-TRAF2 complex to sensitize tumor cells to TNF{alpha}

2008-07-07

Synthetic inhibitor of apoptosis (IAP) antagonists induce degradation of IAP proteins such as cellular IAP1 (cIAP1), activate nuclear factor B (NF-B) signaling, and sensitize cells to tumor necrosis factor (TNF). The physiological relevance of these discoveries to cIAP1 function remains undetermined. We show that upon ligand binding, the TNF superfamily receptor FN14 recruits a cIAP1–Tnf receptor-associated factor 2 (TRAF2) complex. Unlike IAP antagonists that cause rapid proteasomal degradation of cIAP1, signaling by FN14 promotes the lysosomal degradation of cIAP1–TRAF2 in a cIAP1-dependent manner. TNF-like weak inducer of apoptosis (TWEAK)/FN14 signaling nevertheless promotes the same noncanonical NF-B signaling elicited by IAP antagonists and, in sensitive cells, the same autocrine TNF-induced death occurs. TWEAK-induced loss of the cIAP1–TRAF2 complex sensitizes immortalized and minimally passaged tumor cells to TNF-induced death, whereas primary cells remain resistant. Conversely, cIAP1–TRAF2 complex overexpression limits FN14 signaling and protects tumor cells from TWEAK-induced TNF sensitization. Lysosomal degradation of cIAP1–TRAF2 by TWEAK/FN14 therefore critically alters the balance of life/death signals emanating from TNF-R1 in immortalized cells.

Sequential signals toward podosome formation in NIH-src cells

Oikawa, T., Itoh, T., Takenawa, T. — 2008-07-07

Podosomes (also termed invadopodia in cancer cells) are actin-rich adhesion structures with matrix degradation activity that develop in various cell types. Despite their significant physiological importance, the molecular mechanism of podosome formation is largely unknown. In this study, we investigated the molecular mechanisms of podosome formation. The expression of various phosphoinositide-binding domains revealed that the podosomes in Src-transformed NIH3T3 (NIH-src) cells are enriched with PtdIns(3,4)P2, suggesting an important role of this phosphoinositide in podosome formation. Live-cell imaging analysis revealed that Src-expression stimulated podosome formation at focal adhesions of NIH3T3 cells after PtdIns(3,4)P2 accumulation. The adaptor protein Tks5/FISH, which is essential for podosome formation, was found to form a complex with Grb2 at adhesion sites in an Src-dependent manner. Further, it was found that N-WASP bound all SH3 domains of Tks5/FISH, which facilitated circular podosome formation. These results indicate that augmentation of the N-WASP–Arp2/3 signal was accomplished on the platform of Tks5/FISH-Grb2 complex at focal adhesions, which is stabilized by PtdIns(3,4)P2.

Calreticulin inhibits commitment to adipocyte differentiation

Szabo, E., Qiu, Y., Baksh, S., Michalak, M., Opas, M. — 2008-07-07

Calreticulin, an endoplasmic reticulum (ER) resident protein, affects many critical cellular functions, including protein folding and calcium homeostasis. Using embryonic stem cells and 3T3-L1 preadipocytes, we show that calreticulin modulates adipogenesis. We find that calreticulin-deficient cells show increased potency for adipogenesis when compared with wild-type or calreticulin-overexpressing cells. In the highly adipogenic crt^–/^– cells, the ER lumenal calcium concentration was reduced. Increasing the ER lumenal calcium concentration led to a decrease in adipogenesis. In calreticulin-deficient cells, the calmodulin–Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) pathway was up-regulated, and inhibition of CaMKII reduced adipogenesis. Calreticulin inhibits adipogenesis via a negative feedback mechanism whereby the expression of calreticulin is initially up-regulated by peroxisome proliferator–activated receptor (PPAR). This abundance of calreticulin subsequently negatively regulates the expression of PPAR, lipoprotein lipase, CCAAT enhancer–binding protein , and aP2. Thus, calreticulin appears to function as a Ca²⁺-dependent molecular switch that regulates commitment to adipocyte differentiation by preventing the expression and transcriptional activation of critical proadipogenic transcription factors.

SUMOylation of the MAGUK protein CASK regulates dendritic spinogenesis

Chao, H.-W., Hong, C.-J., Huang, T.-N., Lin, Y.-L., Hsueh, Y.-P. — 2008-07-07

Membrane-associated guanylate kinase (MAGUK) proteins interact with several synaptogenesis-triggering adhesion molecules. However, direct evidence for the involvement of MAGUK proteins in synapse formation is lacking. In this study, we investigate the function of calcium/calmodulin-dependent serine protein kinase (CASK), a MAGUK protein, in dendritic spine formation by RNA interference. Knockdown of CASK in cultured hippocampal neurons reduces spine density and shrinks dendritic spines. Our analysis of the time course of RNA interference and CASK overexpression experiments further suggests that CASK stabilizes or maintains spine morphology. Experiments using only the CASK PDZ domain or a mutant lacking the protein 4.1–binding site indicate an involvement of CASK in linking transmembrane adhesion molecules and the actin cytoskeleton. We also find that CASK is SUMOylated. Conjugation of small ubiquitin-like modifier 1 (SUMO1) to CASK reduces the interaction between CASK and protein 4.1. Overexpression of a CASK–SUMO1 fusion construct, which mimicks CASK SUMOylation, impairs spine formation. Our study suggests that CASK contributes to spinogenesis and that this is controlled by SUMOylation.

Sumoylation regulates lamin A function and is lost in lamin A mutants associated with familial cardiomyopathies

Zhang, Y.-Q., Sarge, K. D. — 2008-07-07

Lamin A mutations cause many diseases, including cardiomyopathies and Progeria Syndrome. The covalent attachment of small ubiquitin-like modifier (SUMO) polypeptides regulates the function of many proteins. Until now, no examples of human disease-causing mutations that occur within a sumoylation consensus sequence and alter sumoylation were known. We show that lamin A is sumoylated at lysine 201 and that two lamin A mutants associated with familial dilated cardiomyopathy, E203G and E203K, exhibit decreased sumoylation. E203 occupies the conserved +2 position in the sumoylation consensus KXE. Lamin A mutants E203G, E203K, and K201R all exhibit a similar aberrant subcellular localization and are associated with increased cell death. Fibroblasts from an individual with the E203K lamin A mutation also exhibit decreased lamin A sumoylation and increased cell death. These results suggest that SUMO modification is important for normal lamin A function and implicate an involvement for altered sumoylation in the E203G/E203K lamin A cardiomyopathies.

CLASP regulates mitochondrial distribution in Schizosaccharomyces pombe

Chiron, S., Bobkova, A., Zhou, H., Yaffe, M. P. — 2008-07-07

Movement of mitochondria in Schizosaccharomyces pombe depends on their association with the dynamic, or plus ends, of microtubules, yet the molecular basis for this interaction is poorly understood. We identified mmd4 in a screen of temperature-sensitive S. pombe strains for aberrant mitochondrial morphology and distribution. Cells with the mmd4 mutation display mitochondrial aggregation near the cell ends at elevated temperatures, a phenotype similar to mitochondrial defects observed in wild-type cells after microtubule depolymerization. However, microtubule morphology and function appear normal in the mmd4 mutant. The mmd4 lesion maps to peg1⁺, which encodes a microtubule-associated protein with homology to cytoplasmic linker protein–associated proteins (mammalian microtubule plus end–binding proteins). Peg1p localizes to the plus end of microtubules and to mitochondria and is recovered with mitochondria during subcellular fractionation. This mitochondrial-associated fraction of Peg1p displays properties of a peripherally associated protein. Peg1p is the first identified microtubule plus end–binding protein required for mitochondrial distribution and likely functions as a molecular link between mitochondria and microtubules.

Shc coordinates signals from intercellular junctions and integrins to regulate flow-induced inflammation

Liu, Y., Sweet, D. T., Irani-Tehrani, M., Maeda, N., Tzima, E. — 2008-07-07

Atherosclerotic plaques develop in regions of the vasculature associated with chronic inflammation due to disturbed flow patterns. Endothelial phenotype modulation by flow requires the integration of numerous mechanotransduction pathways, but how this is achieved is not well understood. We show here that, in response to flow, the adaptor protein Shc is activated and associates with cell–cell and cell–matrix adhesions. Shc activation requires the tyrosine kinases vascular endothelial growth factor receptor 2 and Src. Shc activation and its vascular endothelial cadherin (VE-cadherin) association are matrix independent. In contrast, Shc binding to integrins requires VE-cadherin but occurs only on specific matrices. Silencing Shc results in reduction in both matrix-independent and matrix-dependent signals. Furthermore, Shc regulates flow-induced inflammatory signaling by activating nuclear factor B–dependent signals that lead to atherogenesis. In vivo, Shc is activated in atherosclerosis-prone regions of arteries, and its activation correlates with areas of atherosclerosis. Our results support a model in which Shc orchestrates signals from cell–cell and cell–matrix adhesions to elicit flow-induced inflammatory signaling.

Actin and myosin in the mitotic spindle

2008-07-07

Pegging mitochondrial position

2008-07-07

SUMO keeps lamin A in place

2008-07-07

Atherosclerosis needs Shc

2008-07-07