Non-inositol 1,4,5-trisphosphate (IP3) receptor IP3-binding proteins.

Conventionally, myo-D-inositol 1, 4,5-trisphosphate (IP3) is thought to exert its second messenger effects through the gating of IP3R Ca2+ release channels, located in Ca2+-storage organelles like the endoplasmic reticulum. However, there is considerable indirect evidence to support the concept that IP3 might interact with other, non-IP3R proteins within cells. To explore this possibility further, the Protein Data Bank was searched using the term "IP3". This resulted in the retrieval of 203 protein structures, the majority of which were members of the IP3R/ryanodine receptor superfamily of channels. Only 49 of these structures were complexed with IP3. These were inspected for their ability to interact with the carbon-1 phosphate of IP3, since this is the least accessible phosphate group of its precursor, phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2). This reduced the number of structures retrieved to 35, of which 9 were IP3Rs. The remaining 26 structures represent a diverse range of proteins, including inositol-lipid metabolizing enzymes, signal transducers, PH domain containing proteins, cytoskeletal anchor proteins, the TRPV4 ion channel, a retroviral Gag protein and fibroblast growth factor 2. Such proteins may impact on IP3 signalling and its effects on cell-biology. This represents an area open for exploration in the field of IP3 signalling.

Myo-D-inositol Trisphosphate Signalling in Oomycetes.

Oomycetes are pathogens of plants and animals, which cause billions of dollars of global losses to the agriculture, aquaculture and forestry sectors each year. These organisms superficially resemble fungi, with an archetype being Phytophthora infestans, the cause of late blight of tomatoes and potatoes. Comparison of the physiology of oomycetes with that of other organisms, such as plants and animals, may provide new routes to selectively combat these pathogens. In most eukaryotes, myo-inositol 1,4,5 trisphosphate is a key second messenger that links extracellular stimuli to increases in cytoplasmic Ca2+, to regulate cellular activities. In the work presented in this study, investigation of the molecular components of myo-inositol 1,4,5 trisphosphate signaling in oomycetes has unveiled similarities and differences with that in other eukaryotes. Most striking is that several oomycete species lack detectable phosphoinositide-selective phospholipase C homologues, the enzyme family that generates this second messenger, but still possess relatives of myo-inositol 1,4,5 trisphosphate-gated Ca2+-channels.

Evolution of the cardiac dyad.

Cardiac dyads are the site of communication between the sarcoplasmic reticulum (SR) and infoldings of the sarcolemma called transverse-tubules (TT). During heart excitation-contraction coupling, Ca2+-influx through L-type Ca2+ channels in the TT is amplified by release of Ca2+-from the SR via type 2 ryanodine receptors, activating the contractile apparatus. Key proteins involved in cardiac dyad function are bridging integrator 1 (BIN1), junctophilin 2 and caveolin 3. The work presented here aims to reconstruct the evolutionary history of the cardiac dyad, by surveying the scientific literature for ultrastructural evidence of these junctions across all animal taxa; phylogenetically reconstructing the evolutionary history of BIN1; and by comparing peptide motifs involved in TT formation by this protein across metazoans. Key findings are that cardiac dyads have been identified in mammals, arthropods and molluscs, but not in other animals. Vertebrate BIN1 does not group with members of this protein family from other taxa, suggesting that invertebrate BINs are paralogues rather orthologues of this gene. Comparisons of BIN1 peptide sequences of mammals with those of other vertebrates reveals novel features that might contribute to TT and dyad formation. The analyses presented here suggest that the cardiac dyad evolved independently several times during metazoan evolution: an unexpected observation given the diversity of heart structure and function between different animal taxa. This article is part of the theme issue 'The cardiomyocyte: new revelations on the interplay between architecture and function in growth, health, and disease'.

Ryanodine receptor calcium release channels in trophoblasts and their role in cell migration.

Trophoblasts are specialized epithelial cells of the placenta that are involved in invasion, communication and the exchange of materials between the mother and fetus. Cytoplasmic Ca2+ ([Ca2+]c) plays critical roles in regulating such processes in other cell types, but relatively little is known about the mechanisms that control this second messenger in trophoblasts. In the current study, the presence of RyRs and their accessory proteins in placental tissues and in the BeWo choriocarcinoma, a model trophoblast cell-line, were examined using immunohistochemistry and Western immunoblotting. Contributions of RyRs to Ca2+ signalling and to random migration in BeWo cells were investigated using fura-2 fluorescent and brightfield videomicroscopy. The effect of RyR inhibition on reorganization of the F-actin cytoskeleton elicited by the hormone angiotensin II, was determined using phalloidin-labelling and confocal microscopy. RyR1 and RyR3 proteins were detected in trophoblasts of human first trimester and term placental villi, along with the accessory proteins triadin and calsequestrin. Similarly, RyR1, RyR3, triadin and calsequestrin were detected in BeWo cells. In this cell-line, activation of RyRs with micromolar ryanodine increased [Ca2+]c, whereas pharmacological inhibition of these channels reduced Ca2+ transients elicited by the peptide hormones angiotensin II, arginine vasopressin and endothelin 1. Angiotensin II increased the velocity, total distance and Euclidean distance of random migration by BeWo cells and these effects were significantly reduced by tetracaine and by inhibitory concentrations of ryanodine. RyRs contribute to reorganization of the F-actin cytoskeleton elicited by angiotensin II, since inhibition of these channels restores the parallelness of these structures to control levels. These findings demonstrate that trophoblasts contain a suite of proteins similar to those in other cell types possessing highly developed Ca2+ signal transduction systems, such as skeletal muscle. They also indicate that these channels regulate the migration of trophoblast cells, a process that plays a key role in development of the placenta.

Evolution of Excitation-Contraction Coupling.

In mammalian cardiomyocytes, Ca2+ influx through L-type voltage-gated Ca2+ channels (VGCCs) is amplified by release of Ca2+ via type 2 ryanodine receptors (RyR2) in the sarcoplasmic reticulum (SR): a process termed Ca2+-induced Ca2+-release (CICR). In mammalian skeletal muscles, VGCCs play a distinct role as voltage-sensors, physically interacting with RyR1 channels to initiate Ca2+ release in a mechanism termed depolarisation-induced Ca2+-release (DICR). In the current study, we surveyed the genomes of animals and their close relatives, to explore the evolutionary history of genes encoding three proteins pivotal for ECC: L-type VGCCs; RyRs; and a protein family that anchors intracellular organelles to plasma membranes, namely junctophilins (JPHs). In agreement with earlier studies, we find that non-vertebrate eukaryotes either lack VGCCs, RyRs and JPHs; or contain a single homologue of each protein. Furthermore, the molecular features of these proteins thought to be essential for DICR are only detectable within vertebrates and not in any other taxonomic group. Consistent with earlier physiological and ultrastructural observations, this suggests that CICR is the most basal form of ECC and that DICR is a vertebrate innovation. This development was accompanied by the appearance of multiple homologues of RyRs, VGCCs and junctophilins in vertebrates, thought to have arisen by 'whole genome replication' mechanisms. Subsequent gene duplications and losses have resulted in distinct assemblies of ECC components in different vertebrate clades, with striking examples being the apparent absence of RyR2 from amphibians, and additional duplication events for all three ECC proteins in teleost fish. This is consistent with teleosts possessing the most derived mode of DICR, with their Cav1.1 VGCCs completely lacking in Ca2+ channel activity.