Sensory Neurons Arouse C. elegans Locomotion via Both Glutamate and Neuropeptide Release

C. elegans undergoes periods of behavioral quiescence during larval molts (termed lethargus) and as adults. Little is known about the circuit mechanisms that establish these quiescent states. Lethargus and adult locomotion quiescence is dramatically reduced in mutants lacking the neuropeptide receptor NPR-1. Here, we show that the aroused locomotion of npr-1 mutants results from the exaggerated activity in multiple classes of sensory neurons, including nociceptive (ASH), touch sensitive (ALM and PLM), and stretch sensing (DVA) neurons. These sensory neurons accelerate locomotion via both neuropeptide and glutamate release. The relative contribution of these sensory neurons to arousal differs between larval molts and adults. Our results suggest that a broad network of sensory neurons dictates transitions between aroused and quiescent behavioral states.     

Apremilast,a novel PDE4 inhibitor,inhibits spontaneous production of tumour necrosis factor-alpha from human rheumatoid synovial cells and ameliorates experimental arthritis

Introduction  

Type 4 phosphodiesterases (PDE4) play an important role in immune cells through the hydrolysis of the second messenger, cAMP. Inhibition of PDE4 has previously been shown to suppress immune and inflammatory responses, demonstrating PDE4 to be a valid therapeutic target for immune-mediated pathologies. We assessed the anti-inflammatory effects of a novel PDE4 inhibitor, apremilast, in human synovial cells from rheumatoid arthritis (RA) patients, as well as two murine models of arthritis.     

Cordon-bleu: a new taste in actin nucleation

Polymerization of actin filaments powers many dynamic cellular events and is directed by a small group of actin nucleators. In this issue, Ahuja et al. (2007) identify and characterize a new actin nucleator called cordon-bleu (Cobl) that may drive morphogenesis of the central nervous system during development.     

Dendritic cells infected with vpr-positive human immunodeficiency virus type 1 induce CD8+ T-cell apoptosis via upregulation of tumor necrosis factor alpha

Human immunodeficiency virus type 1 (HIV-1) viral protein R (Vpr) plays a crucial role in viral replication and pathogenesis by inducing cell cycle arrest, apoptosis, translocation of preintegration complex, potentiation of glucocorticoid action, impairment of dendritic cell (DC) maturation, and T-cell activation. Recent studies involving the direct effects of Vpr on DCs and T cells indicated that HIV-1 containing Vpr selectively impairs phenotypic maturation, cytokine network, and antigen presentation in DCs and dysregulates costimulatory molecules and cytokine production in T cells. Here, we have further investigated the indirect effect of HIV-1 Vpr(+) virus-infected DCs on the bystander CD8(+) T-cell population. Our results indicate that HIV-1 Vpr(+) virus-infected DCs dysregulate CD8(+) T-cell proliferation and induce apoptosis. Vpr-containing virus-infected DC-mediated CD8(+) T-cell killing occurred in part through enhanced tumor necrosis factor alpha production by infected DCs and subsequent induction of death receptor signaling and activation of the caspase 8-dependent pathway in CD8(+) T cells. Collectively, these results provide evidence that Vpr could be one of the important contributors to the host immune escape by HIV-1 through its ability to dysregulate both directly and indirectly the DC biology and T-cell functions.     

Regulation of nicotinic receptor trafficking by the transmembrane Golgi protein UNC-50

Nicotinic acetylcholine receptors (AChRs) are pentameric ligand-gated ion channels that mediate fast synaptic transmission at the neuromuscular junction (NMJ). After assembly in the endoplasmic reticulum (ER), AChRs must be transported to the plasma membrane through the secretory apparatus. Little is known about specific molecules that mediate this transport. Here we identify a gene that is required for subtype-specific trafficking of assembled nicotinic AChRs in Caenorhabditis elegans. unc-50 encodes an evolutionarily conserved integral membrane protein that localizes to the Golgi apparatus. In the absence of UNC-50, a subset of AChRs present in body-wall muscle are sorted to the lysosomal system and degraded. However, the trafficking of a second AChR type and of GABA ionotropic receptors expressed in the same muscle cells is not affected in unc-50 mutants. These results suggest that, in addition to ER quality control, assembled AChRs are sorted within the Golgi system by a mechanism that controls the amount of cell-surface AChRs in a subtype-specific way.     

Dopamine mediates context-dependent modulation of sensory plasticity in C. elegans

Dopamine has been implicated in the modulation of diverse forms of behavioral plasticity, including appetitive learning and addiction. An important challenge is to understand how dopamine's effects at the cellular level alter the properties of neural circuits to modify behavior. In the nematode C. elegans, dopamine modulates habituation of an escape reflex triggered by body touch. In the absence of food, animals habituate more rapidly than in the presence of food; this contextual information about food availability is provided by dopaminergic mechanosensory neurons that sense the presence of bacteria. We find that dopamine alters habituation kinetics by selectively modulating the touch responses of the anterior-body mechanoreceptors; this modulation involves a D1-like dopamine receptor, a Gq/PLC-beta signaling pathway, and calcium release within the touch neurons. Interestingly, the body touch mechanoreceptors can themselves excite the dopamine neurons, forming a positive feedback loop capable of integrating context and experience to modulate mechanosensory attention.     

Anoikis evasion in inflammatory breast cancer cells is mediated by Bim-EL sequestration

Inflammatory breast cancer (IBC) is a rare and highly invasive type of breast cancer, and patients diagnosed with IBC often face a very poor prognosis. IBC is characterized by the lack of primary tumor formation and the rapid accumulation of cancerous epithelial cells in the dermal lymphatic vessels. Given that normal epithelial cells require attachment to the extracellular matrix (ECM) for survival, a comprehensive examination of the molecular mechanisms underlying IBC cell survival in the lymphatic vessels is of paramount importance to our understanding of IBC pathogenesis. Here we demonstrate that, in contrast to normal mammary epithelial cells, IBC cells evade ECM-detachment-induced apoptosis (anoikis). ErbB2 and EGFR knockdown in KPL-4 and SUM149 cells, respectively, causes decreased colony growth in soft agar and increased caspase activation following ECM detachment. ERK/MAPK signaling was found to operate downstream of ErbB2 and EGFR to protect cells from anoikis by facilitating the formation of a protein complex containing Bim-EL, LC8, and Beclin-1. This complex forms as a result of Bim-EL phosphorylation on serine 59, and thus Bim-EL cannot localize to the mitochondria and cause anoikis. These results reveal a novel mechanism that could be targeted with innovative therapeutics to induce anoikis in IBC cells.Inflammatory breast cancer (IBC) is a rare and highly invasive type of breast cancer, and patients diagnosed with IBC often face a very poor prognosis. The 5-year survival rate for patients with IBC is <40%, while the 5-year survival rate of all other breast cancers combined is approximately 90%.^{1, 2, 3, 4} This poor prognosis can be attributed to a number of factors, including the propensity for misdiagnosis of the disease due to its unique clinical presentation.^{5, 6, 7} In contrast to most breast cancers, IBC is characterized by the lack of discernible primary tumor formation and the accumulation of cancerous epithelial cells in the dermal lymphatic vessels.⁸ This lodging of IBC cells in the dermal lymphatics manifests as what appears to be inflammation, oftentimes causing clinicians to incorrectly diagnose the malady. Given that IBC cells are inherently aggressive, misdiagnosis is particularly problematic as a correct diagnosis or appropriate treatment is prolonged until more advanced disease is discovered. Thus it is imperative to gain a better understanding of the unique molecular mechanisms underlying IBC pathogenesis so that improved therapies can be designed to specifically eliminate IBC cells in a manner that improves patient outcome.Unfortunately, few treatment options exist that are specifically designed to combat IBC. A review of nearly 400 IBC patients treated at The University of Texas MD Anderson Cancer Center between 1974 and 2005 demonstrated that there has been no significant improvement in prognosis for patients with IBC over the past 30 years.¹ Many recent studies have focused on assessing the efficacy of chemotherapeutic regimens in IBC cells/patients where success had previously been observed only in the treatment of non-IBCs.^{9, 10} Some progress has been made in understanding the mechanisms underlying the invasive nature of IBC. For instance, Akt1 has been identified as a possible chemotherapeutic target that appears to be involved in the aggressive behavior of IBC cells.¹¹ Other studies have identified RhoC, which is overexpressed in 90% of IBC tissue samples, as a potent oncogene contributing to IBC pathogenesis.^{11, 12, 13, 14, 15} More recently, evidence implicating the membrane protein TIG1 and the receptor tyrosine kinase Axl in the oncogenic behavior of IBC cells has been uncovered.¹⁶ However, despite these advances, knowledge of the biological mechanisms underlying IBC pathogenesis remains fairly rudimentary, and additional research dedicated to understanding the unique molecular pathways involved in IBC progression remains essential.Given that IBC cells do not form a palpable primary tumor and instead flourish in suspension in the lymph of the dermal lymphatic vessels, we hypothesized that IBC cells must have an inherent ability to survive in the absence of attachment to the extracellular matrix (ECM). Normal mammary epithelial cells require attachment to the ECM to inhibit anoikis, which is defined as caspase-dependent cell death caused by ECM detachment.¹⁷ It has become clear that tumor progression and metastasis require cancer cells to inhibit anoikis, oftentimes through alterations in intracellular signaling pathways.^{18, 19, 20} Interestingly, previous studies have shown that ErbB2 and EGFR, which are hyperactivated in a substantial percentage of IBC patients,²¹ can effectively antagonize the anoikis program to facilitate anchorage-independent growth.^{22, 23, 24, 25, 26, 27, 28} However, a detailed examination of the molecular mechanisms underlying anoikis inhibition in IBC cells has yet to be completed. In this study, we demonstrate that signaling from EGFR and ErbB2 through ERK/MAPK has a major role in the ability of IBC cells to survive in the absence of ECM attachment. Surprisingly, we have discovered that ERK-mediated anoikis suppression in IBC cells is not due to targeting of the pro-apoptotic protein Bim-EL for degradation that has previously been observed in mammary epithelial cells.^{23, 27} Rather, ERK activation in IBC cells promotes the formation of a protein complex containing Bim-EL, Beclin-1, and LC8, which functions to sequester Bim-EL from the mitochondria and thereby block anoikis. In support of the importance of this signaling pathway in IBC patients, five of the seven IBC patient samples assayed showed discernible Bim-EL staining. Collectively, these data identify a novel mechanism utilized by IBC cells to survive during ECM detachment and reveal a potential target for the development of anoikis-inducing chemotherapeutics targeting IBC cells.     

How actin crosslinking and bundling proteins cooperate to generate an enhanced cell mechanical response

Actin-crosslinking proteins organize actin filaments into dynamic and complex subcellular scaffolds that orchestrate important mechanical functions, including cell motility and adhesion. Recent mutation studies have shown that individual crosslinking proteins often play seemingly non-essential roles, leading to the hypothesis that they have considerable redundancy in function. We report live-cell, in vitro, and theoretical studies testing the mechanical role of the two ubiquitous actin-crosslinking proteins, alpha-actinin and fascin, which co-localize to stress fibers and the basis of filopodia. Using live-cell particle tracking microrheology, we show that the addition of alpha-actinin and fascin elicits a cell mechanical response that is significantly greater than that originated by alpha-actinin or fascin alone. These live-cell measurements are supported by quantitative rheological measurements with reconstituted actin filament networks containing pure proteins that show that alpha-actinin and fascin can work in concert to generate enhanced cell stiffness. Computational simulations using finite element modeling qualitatively reproduce and explain the functional synergy of alpha-actinin and fascin. These findings highlight the cooperative activity of fascin and alpha-actinin and provide a strong rationale that an evolutionary advantage might be conferred by the cooperative action of multiple actin-crosslinking proteins with overlapping but non-identical biochemical properties. Thus the combination of structural proteins with similar function can provide the cell with unique properties that are required for biologically optimal responses.     

Molecular and functional characterization of a novel splice variant of ANKHD1 that lacks the KH domain and its role in cell survival and apoptosis

Multiple ankyrin repeat motif-containing proteins play an important role in protein-protein interactions. ANKHD1 proteins are known to possess multiple ankyrin repeat domains and a single KH domain with no known function. Using yeast two-hybrid system analysis, we identified a novel splice variant of ANKHD1. This splice variant of ANKHD1, which we designated as HIV-1 Vpr-binding ankyrin repeat protein (VBARP), does not contain the signature KH domain, and codes for only a single ankyrin repeat motif. We characterized VBARP by molecular and functional analysis, revealing that VBARP is ubiquitously expressed in different tissues as well as cell lines of different lineage. In addition, blast searches indicated that orthologs and homologs to VBARP exist in different phyla, suggesting that VBARP might be evolutionarily conserved, and thus may be involved in basic cellular function(s). Furthermore, biochemical analysis revealed the presence of two VBARP isoforms coding for 69 and 49 kDa polypeptides, respectively, that are primarily localized in the cytoplasm. Functional analysis using short interfering RNA approaches indicate that this gene product is essential for cell survival through its regulation of caspases. Taken together, these results indicate that VBARP is a novel splice variant of ANKHD1 and may play a role in cellular apoptosis (antiapoptotic) and cell survival pathway(s).