Generation of constitutively active calcineurin by calpain contributes to delayed neuronal death following mouse brain ischemia

Abstract

Calpain, a Ca(2+)-dependent cysteine protease, in vitro converts calcineurin (CaN) to constitutively active forms of 45 kDa and 48 kDa by cleaving the autoinhibitory domain of the 60 kDa subunit. In a mouse middle cerebral artery occlusion (MCAO) model, calpain converted the CaN A subunit to the constitutively active form with 48 kDa in vivo. We also confirmed increased Ca(2+)/CaM-independent CaN activity in brain extracts. The generation of constitutively active and Ca(2+)/CaM-independent activity of CaN peaked 2 h after reperfusion in brain extracts. Increased constitutively active CaN activity was associated with dephosphorylation of dopamine-regulated phosphoprotein-32 in the brain. Generation of constitutively active CaN was accompanied by translocation of nuclear factor of activated T-cells (NFAT) into nuclei of hippocampal CA1 pyramidal neurons. In addition, a novel calmodulin antagonist, DY-9760e, blocked the generation of constitutively active CaN by calpain, thereby inhibiting NFAT nuclear translocation. Together with previous studies indicating that NFAT plays a critical role in apoptosis, we propose that calpain-induced CaN activation in part mediates delayed neuronal death in brain ischemia.

The Down Syndrome Critical Region Protein RCAN1 Regulates Long-Term Potentiation and Memory via Inhibition of Phosphatase Signaling

“…Therefore, the elevation in calcineurin activity in the hippocampus from RCAN1 mutant mice is not because of a corresponding change in the abundance of the catalytic subunit of calcineurin but more likely attributable to a loss of inhibition by RCAN1. We also noticed an increase in a 48 kDa constitutively active fragment of CnA that can be generated by calpain cleavage (Wu et al, 2004;Shioda et al, 2006) was present in the hippocampus of RCAN1 mutant mice. No difference in the abundance of the 48 kDa fragment of CnA was observed in the soluble supernatant fraction between RCAN1 mutant mice and wild-type mice.…”

“…Moreover, recent findings indicate that acute blockade of calcineurin activity improves memory and cognitive function in AD model mice (Dineley et al, 2007). Cleavage of calcineurin by calpain increases levels of the constitutively active 48 kDa CnA isoform in response to ischemic injury and in AD (Wu et al, 2004;Liu et al, 2005;Shioda et al, 2006). An increase in this cleaved form of CnA in the hippocampus of RCAN1 knock-out mice (Fig.…”

Mechanistic Role of Calpains in Postischemic Neurodegeneration

“…Many of these molecules are regulated by calmodulin, and a common effect of calpain cleavage is to remove this regulation. Excitotoxicity in hippocampal neurons (Wu et al, 2004) and focal brain ischemia in mice (Shioda et al, 2006) induces calpain cleavage of the calmodulin-dependent phosphatase calcineurin, producing a constitutively active form. Calpain also produces an active form of type IIa calmodulindependent protein kinase and degrades neuronal nitric oxide synthase after both in vitro and in vivo administration of a variety of neurotoxins (Hajimohammadreza et al, 1997).…”

The Regulator of Calcineurin 1 (RCAN1/DSCR1) Activates the cAMP Response Element-binding Protein (CREB) Pathway

“…Calcineurin has been implicated in many cellular signaling processes, such as neuronal synaptic plasticity and memory formation (20,21). Dysregulation of its activity has also been associated with brain disease and injury, such as Alzheimer's disease and ischemia (22,23).…”