Downstream Regulatory Element Antagonist Modulator Regulates Ca²⁺Homeostasis and Viability in Cerebellar Neurons

Abstract

The Na+/Ca2+ exchangers NCX1, NCX2, and NCX3 are vital for the control of cellular Ca2+ homeostasis. Here, we show that a doublet of downstream regulatory element sites in the promoter of the NCX3 gene mediates transcriptional repression of NCX3 by the Ca2+-modulated transcriptional repressor downstream regulatory element antagonist modulator (DREAM). Overexpression of a DREAM EF-hand mutant insensitive to Ca2+ (EFmDREAM) in hippocampus and cerebellum of transgenic mice significantly reduced NCX3 mRNA and protein levels without modifying NCX1 and NCX2 expression. Cerebellar granules from EFmDREAM transgenic mice showed increased levels of cytosolic Ca2+ and were more vulnerable to increased Ca2+ influx after partial opening of voltage-gated plasma membrane Ca2+ channels induced by increasing K+ in the culture medium but survived better in the conditions of reduced Ca2+ influx prevailing in low extracellular K+. Overexpression of NCX3 in EFmDREAM transgenic granules using a lentiviral vector restored the normal survival response to high K+ observed in wild-type granules. Thus, the downregulation of the regulator of Ca2+ homeostasis NCX3 by Ca2+-regulated DREAM is a striking example of the autoregulatory property of the Ca2+ signal in neurons.

Kv channel-interacting proteins as neuronal and non-neuronal calcium sensors

“…Especially KChIP3, but also KChIP1, KChIP2 and KChIP4, may fulfill the function of a transcriptional repressor, either directly, by binding to downstream regulatory element (DRE) sites of genes [19,55–61], or indirectly, by binding to other transcription factors [62–64]. Here the focus is put on direct KChIP-mediated control of gene transcription (Figure 1C), which appears to be commonly linked to excitability.…”

“…Thus, in nociceptive spinal cord neurons the prodynorphin gene is switched on during massive action potential firing with elevated cytoplasmic Ca 2+ levels, to modulate pain perception via κ-opioid receptors [19,41,65]. In an analogous manner the Na + /Ca 2+ exchanger (NCX) 3 expression in cerebellar neurons is under the control of the transcriptional repressor KChIP3 (DREAM), which may occupy a doublet of DRE sites in the NCX3 gene when Ca 2+ is low [55]. Dissociation of KChIP3 (DREAM) from these DRE sites in an EF-hand-dependent manner and activation of the NCX3 gene occurs if cytoplasmic Ca 2+ is elevated, which has been interpreted as a negative feed-back control loop of Ca 2+ homeostasis [55].…”

“…In an analogous manner the Na + /Ca 2+ exchanger (NCX) 3 expression in cerebellar neurons is under the control of the transcriptional repressor KChIP3 (DREAM), which may occupy a doublet of DRE sites in the NCX3 gene when Ca 2+ is low [55]. Dissociation of KChIP3 (DREAM) from these DRE sites in an EF-hand-dependent manner and activation of the NCX3 gene occurs if cytoplasmic Ca 2+ is elevated, which has been interpreted as a negative feed-back control loop of Ca 2+ homeostasis [55]. KChIP3 seems to also exist as a cytosolic protein which is translocated to the nucleus if Ca 2+ signaling is out of control [66].…”

Changes in calcium dynamics following the reversal of the sodium-calcium exchanger have a key role in AMPA receptor-mediated neurodegeneration via calpain activation in hippocampal neurons

“…Furthermore, NCX3 was shown to be more relevant for cell survival than NCX1 or NCX2, namely in cultured cerebellar granule neurons. 14, 15 We recently reported that neurotoxicity induced by activation of AMPA receptors is characterized by calpain activation, lack of caspase activation, nuclear condensation/fragmentation, release of cytochrome c from mitochondria, decreased intracellular ATP levels, production of nitric oxide, moderate superoxide production and increased levels of peroxynitrite. [16][17][18] In this in vitro model of excitotoxicity, the cell death mechanisms are mostly dependent on calpains, 11 providing a good model for studying calpain-dependent phenomena.…”

The DREAM Protein Negatively Regulates the NMDA Receptor through Interaction with the NR1 Subunit

“…Downstream regulatory element antagonist modulator (DREAM), which was first identified as a Ca 2ϩ -regulated transcriptional repressor, contains four Ca 2ϩ -binding EF hand domains and belongs to the neuronal calcium sensor (NCS) family (Carrió n et al, 1999;Burgoyne, 2007). DREAM was named for its ability to block gene expression in its Ca 2ϩ -free form via direct binding with the downstream regulatory element sequence in target genes such as preprodynorphin, c-fos, Hrk, and Na ϩ and Ca 2ϩ exchanger NCX3 (Carrió n et al, 1999;Sanz et al, 2001;Gomez-Villafuertes et al, 2005). DREAM was also named calsenilin or potassium channel-interacting protein 3 (KChIP3) (Buxbaum et al, 1998;An et al, 2000), suggesting that it has multifunctional properties.…”