Modulation of G protein-coupled receptor (GPCR) signaling by local changes in intracellular calcium concentration is an established function of Calmodulin (CaM) which is known to interact with many GPCRs. Less is known about the functional role of the closely related neuronal EF-hand Ca(2+)-sensor proteins that frequently associate with CaM targets with different functional outcome. In the present study we aimed to investigate if a target of CaM-the A(2A) adenosine receptor is able to associate with two other neuronal calcium binding proteins (nCaBPs), namely NCS-1 and caldendrin. Using bioluminescence resonance energy transfer (BRET) and co-immunoprecipitation experiments we show the existence of A(2A)-NCS-1 complexes in living cells whereas caldendrin did not associate with A(2A) receptors under the conditions tested. Interestingly, NCS-1 binding modulated downstream A(2A) receptor intracellular signaling in a Ca(2+)-dependent manner. Taken together this study provides further evidence that neuronal Ca(2+)-sensor proteins play an important role in modulation of GPCR signaling.

Calneuron-1 and -2 are neuronal EF-hand type calcium sensor proteins that are prominently targeted to trans-Golgi network membranes and impose a calcium threshold at the Golgi for Phosphatidylinositol 4-OH kinase III β activation and the regulated local synthesis of phospholipids that are crucial for TGN-to-plasma membrane trafficking. In this study we show that Calneurons are type IV tail-anchored proteins that are inserted into the ER membrane via an association of a 23 amino-acid long transmembrane domain (TMD) with the TRC40/Asna1 chaperone complex. Following trafficking to the Golgi, Calneurons are probably retained in the TGN because of the length of the TMD and PI(4)P lipid binding. Both Calneurons rapidly self associate in vitro and in vivo via their TMD and EF-hand containing N-terminus. Although dimerization and potentially multimerization precludes TRC40/Asna1 binding and thereby membrane insertion, we found no evidence for a cytosolic pool of Calneurons and could demonstrate that self association of Calneurons is restricted to membrane inserted protein. The dimerization properties and the fact that they, unlike every other EF-hand Calmodulin like Ca2+-sensor, are always associated with membranes of the secretory pathway, including vesicles and plasma membrane, suggests a high degree of spatial segregation for physiological target interactions.

In recent years, substantial progress has been made towards an understanding of the physiological function of EF-hand calcium sensor proteins of the Calmodulin (CaM) superfamily in neurons. This deeper appreciation is based on the identification of novel target interactions, structural studies and the discovery of novel signalling mechanisms in protein trafficking and synaptic plasticity, in which CaM-like sensor proteins appear to play a role. However, not all interactions are of plausible physiological relevance and in many cases it is not yet clear how the CaM signaling network relates to the proposed function of other EF-hand sensors. In this review, we will summarize these findings and address some of the open questions on the functional role of EF-hand calcium binding proteins in neurons.

[This corrects the article on p. e17276 in vol. 6.].

In recent years a number of potential synapto-nuclear protein messengers have been characterized that are thought to be involved in plasticity-related gene expression, and that have the capacity of importin- mediated and activity-dependent nuclear import. However, there is a surprising paucity of data showing the nuclear import of such proteins in cellular models of learning and memory. Only recently it was found that the transcription factor cyclic AMP response element binding protein 2 (CREB2) transits to the nucleus during long-term depression (LTD), but not during long-term potentiation (LTP) of synaptic transmission in hippocampal primary neurons. Jacob is another messenger that couples NMDA-receptor-activity to nuclear gene expression. We therefore aimed to study whether Jacob accumulates in the nucleus in physiological relevant models of activity-dependent synaptic plasticity. We have analyzed the dynamics of Jacob's nuclear import following induction of NMDA-receptor dependent LTP or LTD at Schaffer collateral-CA1 synapses in rat hippocampal slices. Using time-lapse imaging of neurons expressing a Jacob-Green-Fluorescent-Protein we found that Jacob rapidly translocates from dendrites to the nucleus already during the tetanization period of LTP, but not after induction of LTD. Immunocytochemical stainings confirmed the nuclear accumulation of endogenous Jacob in comparison to apical dendrites after induction of LTP but not LTD. Complementary findings were obtained after induction of NMDA-receptor dependent chemical LTP and LTD in hippocampal primary neurons. However, in accordance with previous studies, high concentrations of NMDA and glycine as well as specific activation of extrasynaptic NMDA-receptors resembling pathological conditions induce an even more profound increase of nuclear Jacob levels. Taken together, these findings suggest that the two major forms of NMDA-receptor dependent synaptic plasticity, LTP and LTD, elicit the transition of different synapto-nuclear messengers albeit in both cases importin-mediated retrograde transport and NMDA-receptor activation is required.

Some of the most exciting advances in molecular-functional imaging of cancer are occurring at the interface between chemistry and imaging. Several of these advances have occurred through the development of novel imaging probes that report on molecular pathways, the tumor micro-environment and the response of tumors to treatment; as well as through novel image-guided platforms such as nanoparticles and nanovesicles that deliver therapeutic agents against specific targets and pathways. Cancer cells have a remarkable ability to evade destruction despite the armamentarium of drugs currently available. While these drugs can destroy cancer cells, normal tissue toxicity is a major limiting factor, a problem further compounded by poor drug delivery. One major challenge for chemistry continues to be to eliminate cancer cells without damaging normal tissues. Here we have selected examples of MRI and optical imaging, to demonstrate how integrating imaging with novel probes can facilitate the successful treatment of this multifaceted disease.

Caldendrin and recoverin are Ca(2+)-sensor proteins operating in neuronal systems. In a search for novel binding partners of recoverin we employed an affinity column and identified caldendrin as a possible interaction partner. Caldendrin and recoverin colocalized in the retina in a subset of bipolar cells and in the pineal gland as revealed by immunofluorescence studies. The binding process was controlled by Ca(2+) as revealed by pull-down assays, changes in tryptophane fluorescence during binding and surface plasmon resonance studies. Importantly, caldendrin existed as a Ca(2+)-independent homodimer whereas a complex of recoverin and caldendrin formed with low to moderate affinity in the presence of Ca(2+). Co-transfection of COS-7 cells with plasmids harboring the gene for fluorescently labelled recoverin and caldendrin was used to study the cellular distribution by time-lapse fluorescence microscopy. Apparently, the increase of intracellular Ca(2+) facilitates the translocation of caldendrin to intracellular membranes, which is under control of complex formation with recoverin.

Several studies indicate that NMDA receptor signaling is involved in Abeta oligomer-mediated impairment of neuronal function and morphology. Utilizing primary neuronal cell culture and hippocampal slices from rat and mouse, we found that Abeta oligomer administration readily impairs long-term potentiation, reduces baseline synaptic transmission, decreases neuronal spontaneous network activity and induces retraction of synaptic contacts long before major cytotoxic effects are visible. Interestingly, all these effects can be blocked with the NR2B-containing NMDA-receptor antagonist ifenprodil or Ro 25-6981 suggesting that activation of downstream effectors of these receptors is involved in early detrimental actions of Abeta oligomers. In line we found that Jacob, a messenger that can couple extrasynaptic NMDA-receptor activity to CREB dephosphorylation, accumulates in the nucleus after Abeta oligomer administration and that the nuclear accumulation of Jacob can be blocked by a simultaneous application of ifenprodil. We conclude that Abeta oligomers induce early neuronal dysfunction mainly by activation of NR2B-containing NMDA-receptors.

The regulated local synthesis of PtdIns4P and PtdIns(4,5)P(2) is crucial for TGN (trans-Golgi network)-plasma membrane trafficking. The activity of PI4Kbeta (phosphoinositide 4-kinase IIIbeta) at the Golgi membrane is a first mandatory step in this process. In addition to PI4Kbeta activity, elevated Ca(2+) levels are also needed for the exit of vesicles from the TGN. The reason for this Ca(2+) requirement is at present unclear. In the present review, we discuss the role of neuronal Ca(2+)-sensor proteins in the regulation of PI4Kbeta and suggest that this regulation might impose a need for elevated Ca(2+) levels for a late step of vesicle assembly.

The SH3-HOOK-GUK domains of the postsynaptic scaffolding proteins SAP90/PSD-95 and SAP97 are established targets of synaptic plasticity processes in the brain. A crucial molecular mechanism involved is the transition of this domain to different conformational states. We purified the SH3-HOOK-GUK domain of both proteins to examine variations in protein conformation and stability. As monitored by circular dichroism and differential scanning calorimetry, SAP97 (Tm = 64(0)C) is significantly more thermal stable than SAP90/PSD-95 (Tm = 52(0)C) and follows a bimodal phase transition. GdmCl-induced equilibrium unfolding of both proteins follows the two-state transitions and thus does not involve the accumulation of stable intermediate state(s). Equilibrium unfolding of SAP97 is highly cooperative from a native state to an unfolded state. In contrast, SAP90/PSD-95 follows a non-cooperative transition from native to unfolded states. A highly cooperative unfolding reaction in case of SAP97 indicates that the protein existed initially as a compact, well-folded structure, while the gradual, non-cooperative melting reaction in case of SAP90/PSD-95 indicates that the protein is in comparison more flexible.