Structure, Function and Regulation of the Plasma Membrane Calcium Pump in Health and Disease.

In this review, I summarize the present knowledge of the structural and functional properties of the mammalian plasma membrane calcium pump (PMCA). It is outlined how the cellular expression of the different spliced isoforms of the four genes are regulated under normal and pathological conditions.

The Influence of Thyroid Hormone on Ca2+ Signaling Pathways During Embryonal Development.

Thyroid hormones influence brain development through the regulation of gene expression. Ca2+-dependent gene expression is a major pathway controlled by the Ca2+/calmodulin-dependent protein kinase IV (CaMKIV), which in turn is induced by the thyroid hormone T3, as also demonstrated in a mouse embryonic stem cell line. In addition, T3 controls the expression of neurexin, synaptotagmin2 (SYT2), synaptotagmin-related gene1 (SRG1), and a number of other genes involved in neurotransmitter release in a Ca2+-dependent manner. It has been noticed that the development of dopaminergic neurons by evoking significant calcium entry occurs through TRPC calcium channels. It was also demonstrated that the T3-mediated development of an early neuronal network is characteristic for depolarizing GABAergic neurons concomitant with intracellular calcium transients. An important aspect of T3-dependent regulation of gene expression in the developing brain is its modulation by the transcription activator COUP-TF1. Regulation of alternative splicing by CaMKIV is another important aspect for embryonal neural development since it can lead to the expression of PMCA1a, the neuronal-specific isoform of the plasma membrane calcium pump. Maternal hypothyroidism or CaMKIV deficiency can have a severe influence on fetal brain development.

The Plasma Membrane Calcium Pump (PMCA): Regulation of Cytosolic Ca2+, Genetic Diversities and Its Role in Sub-plasma Membrane Microdomains.

In this chapter the four different genes of the mammalian plasma membrane calcium ATPase (PMCA) and their spliced isoforms are discussed with respect to the structural and functional properties of PMCA, the tissue distribution of the different isoforms, including their differences during development. The importance of PMCA for regulating Ca2+ signaling in microdomains under different conditions is also discussed.

Implications of the thyroid hormone on neuronal development with special emphasis on the calmodulin-kinase IV pathway.

Thyroid hormones influence brain development through regulation of gene expression. This is especially true for Ca2+-dependent regulation since a major pathway is controlled by the Ca2+/calmodulin-dependent protein kinase IV (CaMKIV) which in turn is induced by the thyroid hormone T3. In addition, CaMKIV is involved in regulation of alternative splicing of a number of protein isoforms, among them PMCA1a, the neuronal specific isoform of the plasma membrane calcium pump. On the other hand, hypothyroidism or CaMKIV deficiency can have a severe influence on brain development. This article is part of a Special Issue entitled: ECS Meeting edited by Claus Heizmann, Joachim Krebs and Jacques Haiech.

Why Calcium? How Calcium Became the Best Communicator.

Calcium carries messages to virtually all important functions of cells. Although it was already active in unicellular organisms, its role became universally important after the transition to multicellular life. In this Minireview, we explore how calcium ended up in this privileged position. Most likely its unique coordination chemistry was a decisive factor as it makes its binding by complex molecules particularly easy even in the presence of large excesses of other cations, e.g. magnesium. Its free concentration within cells can thus be maintained at the very low levels demanded by the signaling function. A large cadre of proteins has evolved to bind or transport calcium. They all contribute to buffer it within cells, but a number of them also decode its message for the benefit of the target. The most important of these "calcium sensors" are the EF-hand proteins. Calcium is an ambivalent messenger. Although essential to the correct functioning of cell processes, if not carefully controlled spatially and temporally within cells, it generates variously severe cell dysfunctions, and even cell death.

Ca(2+) homeostasis and endoplasmic reticulum (ER) stress: An integrated view of calcium signaling.

Cellular Ca(2+) homeostasis is maintained through the integrated and coordinated function of Ca(2+) transport molecules, Ca(2+) buffers and sensors. These molecules are associated with the plasma membrane and different cellular compartments, such as the cytoplasm, nucleus, mitochondria, and cellular reticular network, including the endoplasmic reticulum (ER) to control free and bound Ca(2+) levels in all parts of the cell. Loss of nutrients/energy leads to the loss of cellular homeostasis and disruption of Ca(2+) signaling in both the reticular network and cytoplasmic compartments. As an integral part of cellular physiology and pathology, this leads to activation of ER stress coping responses, such as the unfolded protein response (UPR), and mobilization of pathways to regain ER homeostasis.

The plethora of PMCA isoforms: Alternative splicing and differential expression.

In this review the four different genes of the mammalian plasma membrane calcium ATPase (PMCA) and their spliced isoforms are discussed with respect to their tissue distribution, their differences during development and their importance for regulating Ca²âº homeostasis under different conditions. This article is part of a Special Issue entitled: 13th European Symposium on Calcium.

Ca2+-signaling, alternative splicing and endoplasmic reticulum stress responses.

Ca(2+)-signaling, alternative splicing, and stress responses by the endoplasmic reticulum are three important cellular activities which can be strongly interconnected to alter the expression of protein isoforms in a tissue dependent manner or during development depending on the environmental conditions. This integrated network of signaling pathways permits a high degree of versatility and adaptation to metabolic, developmental and stress processes. Defects in its regulation may lead to cellular malfunction.

Stress induced subcellular distribution of ALG-2, RBM22 and hSlu7.

ALG-2 is a highly conserved calcium binding protein in the cytoplasm which belongs to the family of penta-EF hand proteins. Recently, we showed that ALG-2 is interacting with RBM22, a highly conserved spliceosomal nuclear protein (Montaville et al. Biochim. Biophys. Acta 1763, 1335, 2006; Krebs, Biochim. Biophys. Acta 1793, 979, 2009). In NIH 3T3 cells expressing both proteins a significant amount of ALG-2mRFP is translocated to the nucleus due to the interaction with RBM22-EGFP. hSlu7, another spliceosomal nuclear protein, known to interact with RBM22 in yeast, has been shown to translocate to the cytoplasm under cellular stress conditions. Here we provide evidence that the 2 spliceosomal proteins differ significantly in their subcellular distributions under stress conditions, and that RBM22 enhances the cytoplasmic translocation of hSlu7 under stress, especially a stress induced by thapsigargin. On the other hand, in NIH 3T3 cells expressing RBM22-EGFP and ALG-2-mRFP, ALG-2 remains translocated into the nucleus under both stress conditions, i.e. heat shock or treatment with thapsigargin. We could further demonstrate that these stress conditions had a different influence on the splicing pattern of XBP-1, a marker for the unfolded protein response indicating that ER stress may play a role in stress-induced translocation of spliceosomal proteins. The article is part of a Special Issue entitled: 11th European Symposium on Calcium.

The influence of calcium signaling on the regulation of alternative splicing.

In this review the influence of calcium signaling on the regulation of alternative splicing is discussed with respect to its influence on cell- and developmental-specific expression of different isoforms of the plasma membrane calcium pump (PMCA). In a second part the possibility is discussed that due to the interaction of the calcium-binding protein ALG-2 with a spliceosomal regulator of alternative splicing, RBM22, Ca2+-signaling may thus influence its regulatory property.

Structural characterization of Ca(2+)-ATPase-bound phospholamban in lipid bilayers by solid-state nuclear magnetic resonance (NMR) spectroscopy.

Phospholamban (PLN) regulates cardiac contractility by modulation of sarco(endo)plasmic reticulum calcium ATPase (SERCA) activity. While PLN and SERCA1a, an isoform from skeletal muscle, have been structurally characterized in great detail, direct information about the conformation of PLN in complex with SERCA has been limited. We used solid-state NMR (ssNMR) spectroscopy to deduce structural properties of both the A 36F 41A 46 mutant (AFA-PLN) and wild-type PLN (WT-PLN) when bound to SERCA1a after reconstitution in a functional lipid bilayer environment. Chemical-shift assignments in all domains of AFA-PLN provide direct evidence for the presence of two terminal alpha helices connected by a linker region of reduced structural order that differs from previous findings on free PLN. ssNMR experiments on WT-PLN show no significant difference in binding compared to AFA-PLN and do not support the coexistence of a significantly populated dynamic state of PLN after formation of the PLN/SERCA complex. A combination of our spectroscopic data with biophysical and biochemical data using flexible protein-protein docking simulations provides a structural basis for understanding the interaction between PLN and SERCA1a.

A structural model of the complex formed by phospholamban and the calcium pump of sarcoplasmic reticulum obtained by molecular mechanics.

Phospholamban (PLN) is an intrinsic membrane protein of 52 amino acids that modulates the activity of the reticular Ca(2+) ion pump. We recently solved the three-dimensional structure of chemically synthesized, unphosphorylated, monomeric PLN (C41F) by high-resolution nuclear magnetic resonance spectroscopy in chloroform/methanol. The structure is composed of two alpha-helical regions connected by a beta turn (Type III). We used this structure and the crystallographic structure of the sarcoplasmic reticulum calcium pump (SERCA) recently determined by Toyoshima and co-workers and modeled into its E(2) form by Stokes (1KJU) or by Toyoshima (1FQU). We applied restrained and unrestrained energy optimizations and used the AMBER molecular mechanics force field to model the complex formed between PLN and the pump. The results indicate that transmembrane helix 6 (M6) of the SERCA pump is energetically favored, with respect to the other transmembrane helices, as the PLN binding partner within the membrane and is the only one of these helices that also permits contact between the N-terminal residues of PLN and the critical cytosolic binding loop region of the pump. This result is in agreement with published biochemical data and with the predictions of previous mutagenesis work on the membrane sector of the pump. The model reveals that PLN does not span the entire width of the membrane, that is, its hydrophobic C-terminal end is located near the center of the transmembrane region of the SERCA pump. The model also shows that interaction with M6 is stabilized by additional contacts made by PLN to M4. The contact between the N-terminal portion of PLN and the pump is stabilized by a number of salt and hydrogen-bond bridges, which may be abolished by phosphorylation of PLN. The contacts between the cytosolic portions of PLN and the pump are only observed in the E(2) conformation of the pump. Our model of the complex also offers a plausible structural explanation for the preference of protein kinase A for phosphorylation of Ser16 of PLN.