Protein kinase D (PKD) is known to participate in various cellular functions, including secretory vesicle fission from theGolgi and plasma membrane-directed transport. Here, we report on expression and function of PKD in hippocampal neurons. Expression of an enhanced green fluorescent protein (EGFP)-tagged PKD activity reporter in mouse embryonal hippocampal neurons revealed high endogenous PKD activity at the Golgi complex and in the dendrites, whereas PKD activity was excluded from the axon in parallel with axonal maturation. Expression of fluorescently tagged wild-type PKD1 and constitutively active PKD1 S738/742E (caPKD1) in neurons revealed that both proteins were slightly enriched at the trans-Golgi network (TGN) and did not interfere with its thread-like morphology. By contrast, expression of dominant-negative kinase inactive PKD1 K612W (kdPKD1) led to the disruption of the neuronal Golgi complex, with kdPKD1 strongly localized to the TGN fragments. Similar findings were obtained from transgenic mice with inducible, neuron-specific expression of kdPKD1-EGFP. As a prominent consequence of kdPKD1 expression, the dendritic tree of transfected neurons was reduced, whereas caPKD1 increased dendritic arborization. Our results thus provide direct evidence that PKD activity is selectively involved in the maintenance of dendritic arborization and Golgi structure of hippocampal neurons. INTRODUCTIONNeurons are nondividing and extremely polarized cells, with enormous membrane surface compared with the size of the soma. These features assume a highly specialized secretory machinery, which resembles in certain parts the directed transport mechanisms described in polarized nonneuronal cells (Winckler and Mellman, 1999;Horton and Ehlers, 2004). The vast neuronal membrane surface is functionally and structurally divided into axonal and somatodendritic compartments, possessing specific lipid and protein components required for the spatially different functions, e.g., for pre-or postsynaptic activity (Dresbach et al., 2001;Sheng, 2001;Bresler et al., 2004). The constitutive and precisely directed supply of surface membrane components is indispensable for the function of neurons, especially when considering that postmitotic neurons normally serve throughout the life span of the organism. Up to now, we are far from understanding how the extreme polarization has been evolved and is maintained in neurons.Despite a likely central role in neuronal morphogenesis and membrane trafficking, little is known about the special structure and transport features of the neuronal Golgi apparatus. A thread-like and reticular structure of the Golgi apparatus has been already described in many neuronal types (Takamine et al., 2000;Fujita and Okamoto, 2005;Horton et al., 2005). Central neuronal Golgi complex is localized in the soma and often extends into the principal dendrites, but Golgi elements were also found as discontinuous structures in the distal dendrites, often near synaptic contacts or in dendritic spines (Gardiol et al., 1999;Pierc...

Sponges (Porifera) are nerve- and muscleless. Nevertheless, they react to external stimuli in a coordinated way, by body contraction, oscule closure or stopping pumping activity. The underlying mechanisms are still unknown, but evidence has been found for chemical messenger-based systems. We used the sponge Tethya wilhelma to test the effect of gamma-aminobutyric acid (GABA) and glutamate (L: -Glu) on its contraction behaviour. Minimal activating concentrations were found to be 0.5 microM (GABA) and 50 microM (L: -Glu), respectively. Taking maximum relative contraction speed and minimal relative projected body area as a measure of the sponge's response, a comparison of the dose-response curves indicated a higher sensitivity of the contractile tissue for GABA than for L: -Glu. The concentrations eliciting the same contractile response differ by about 100-fold more than the entire concentration range tested. In addition, desensitising effects and spasm-like reactions were observed. Presumably, a GABA/L: -Glu metabotropic receptor-based system is involved in the regulation of contraction in T. wilhelma. We discuss a coordination system for sponges based on hypothetical chemical messenger pathways.

Background: Sponges (Porifera) are nerve-and muscleless metazoa, but display coordinated motor reactions. Therefore, they represent a valuable phylum to investigate coordination systems, which evolved in a hypothetical Urmetazoon prior to the central nervous system (CNS) of later metazoa. We have chosen the contractile and locomotive species Tethya wilhelma (Demospongiae, Hadromerida) as a model system for our research, using quantitative analysis based on digital time lapse imaging. In order to evaluate candidate coordination pathways, we extracorporeally tested a number of chemical messengers, agonists and antagonists known from chemical signalling pathways in animals with CNS.

The receptor interacting serine/threonine kinase 2 (RIPK2) is essential for linking activation of the pattern recognition receptors NOD1 and NOD2 to cellular signaling events. Recently, it was shown that RIPK2 can form higher order molecular structures in vitro. Here, we demonstrate that RIPK2 forms detergent insoluble complexes in the cytosol of host cells upon infection with invasive enteropathogenic bacteria. Formation of these structures occurred after NF-κB activation and depended on the caspase activation and recruitment domain of NOD1 or NOD2. Complex formation upon activation required RIPK2 autophosphorylation at Y474 and was influenced by phosphorylation at S176. We found that the E3 ligase X-linked inhibitor of apoptosis (XIAP) counteracts complex formation of RIPK2, accordingly mutation of the XIAP ubiquitylation sites in RIPK2 enhanced complex formation. Taken together, our work reveals novel roles of XIAP in the regulation of RIPK2 and expands our knowledge on the function of RIPK2 posttranslational modifications in NOD1/2 signaling.

Mammalian NLR proteins contribute to the regulation and induction of innate and adaptive immunity in mammals although the function of about half of the currently identified NLR proteins remains poorly characterized. Here we analysed the function of the primate-specific NLRP11 gene product. We show that NLRP11 is highly expressed in immune cells, including myeloid cells, B cells and some B cell lymphoma lines. Overexpression of NLRP11 in human cells did not trigger key innate immune signalling pathways including NF-κB and type I interferon responses. NLRP11 harbours a pyrin domain (PYD), which is responsible for inflammasome formation in related NLR proteins. However, NLRP11 did neither interact with the inflammasome adaptor protein ASC nor did it trigger caspase-1 activation. By contrast, expression of NLRP11 specifically repressed NF-κB and type I interferon responses, two key innate immune pathways involved in inflammation. This effect was independent of the PYD domain and ATPase activity of NLRP11. SiRNA-mediated knock-down of NLRP11 in human myeloid THP1 cells validated these findings and revealed enhanced lipopolysaccharide (LPS) and Sendai Virus (SeV)-induced cytokine and interferon responses, respectively in cells with reduced NLRP11 expression. In summary, our work identifies a novel role of NLRP11 in the regulation of inflammatory responses in human cells.An inflammatory response is triggered in response to cell damage and to defend against invading pathogens. In mammals, this is mediated by the activation of cellular pathways cumulating into the release of cytokines, chemokines and interferons. Downstream, this orchestrates recruitment of effector cells, alerts a systemic response and restores tissue homeostasis (1). Activation of pattern-recognition receptors (PRRs 2 ), expressed in and on host cells, which respond towards pathogen-derived substances, cell damage and stress, initiate this response. On the cellular level, inflammation is mediated by the activation of proinflammatory signaling cascades such as the activation of caspase-1, NF-κB, IRFs, amongst others. Chronic In humans, the TLR, NLR and C-type lectin family members are the most relevant PRRs. Many mammalian NLR proteins have been associated with multiple functions in inflammation and innate and adaptive immune responses (4) and dysfunctions in NLRs are associated with a range of diseases including Crohn's disease, periodic fever syndromes and Blau Syndrom (5), highlighting their physiologic relevance NLR proteins have a typical tripartite domain architecture and can be classified functionally according to their N-terminal domain which is, in most cases a CARD or PYD domain (6). Most but not all PYD-containing NLRs can interact with the adaptor molecule ASC to form high molecular weight complexes in cells, referred to as inflammasomes, which function as a platform for the activation of caspase-1 and subsequent IL-1β and IL-18 processing. The NLR family-member pyrin domain-containing protein 11 (NLRP11, NALP11, NOD17, PYPAF6, PAN...