The sarcolemmal calcium pump inhibits the calcineurin/nuclear factor of activated T-cell pathway via interaction with the calcineurin A catalytic subunit

The calcineurin/nuclear factor of activated T-cell (NFAT) pathway represents a crucial transducer of cellular function. There is increasing evidence placing the sarcolemmal calcium pump, or plasma membrane calcium/calmodulin ATPase pump (PMCA), as a potential modulator of signal transduction pathways. We demonstrate a novel interaction between PMCA and the calcium/calmodulin-dependent phosphatase, calcineurin, in mammalian cells. The interaction domains were located to the catalytic domain of PMCA4b and the catalytic domain of the calcineurin A subunit. Endogenous calcineurin activity, assessed by measuring the transcriptional activity of its best characterized substrate, NFAT, was significantly inhibited by 60% in the presence of ectopic PMCA4b. This inhibition was notably reversed by the co-expression of the PMCA4b interaction domain, demonstrating the functional significance of this interaction. PMCA4b was, however, unable to confer its inhibitory effect in the presence of a calcium/calmodulin-independent constitutively active mutant calcineurin A suggesting a calcium/calmodulin-dependent mechanism. The modulatory function of PMCA4b is further supported by the observation that endogenous calcineurin moves from the cytoplasm to the plasma membrane when PMCA4b is overexpressed. We suggest recruitment by PMCA4b of calcineurin to a low calcium environment as a possible explanation for these findings. In summary, our results offer strong evidence for a novel functional interaction between PMCA and calcineurin, suggesting a role for PMCA as a negative modulator of calcineurin-mediated signaling pathways in mammalian cells. This study reinforces the emerging role of PMCA as a molecular organizer and regulator of signaling transduction pathways.

Novel functional interaction between the plasma membrane Ca2+ pump 4b and the proapoptotic tumor suppressor Ras-associated factor 1 (RASSF1)

Plasma membrane calmodulin-dependent calcium ATPases (PMCAs) are enzymatic systems implicated in the extrusion of calcium from the cell. We and others have previously identified molecular interactions between the cytoplasmic COOH-terminal end of PMCA and PDZ domain-containing proteins. These interactions suggested a new role for PMCA as a modulator of signal transduction pathways. The existence of other intracellular regions in the PMCA molecule prompted us to investigate the possible participation of other domains in interactions with different partner proteins. A two-hybrid screen of a human fetal heart cDNA library, using the region 652-840 of human PMCA4b (located in the catalytic, second intracellular loop) as bait, revealed a novel interaction between PMCA4b and the tumor suppressor RASSF1, a Ras effector protein involved in H-Ras-mediated apoptosis. Immunofluorescence co-localization, immunoprecipitation, and glutathione S-transferase pull-down experiments performed in mammalian cells provided further confirmation of the physical interaction between the two proteins. The interaction domain has been narrowed down to region 74-123 of RASSF1C (144-193 in RASSF1A) and 652-748 of human PMCA4b. The functionality of this interaction was demonstrated by the inhibition of the epidermal growth factor-dependent activation of the Erk pathway when PMCA4b and RASSF1 were co-expressed. This inhibition was abolished by blocking PMCA/RASSSF1 association with an excess of a green fluorescent protein fusion protein containing the region 50-123 of RASSF1C. This work describes a novel protein-protein interaction involving a domain of PMCA other than the COOH terminus. It suggests a function for PMCA4b as an organizer of macromolecular protein complexes, where PMCA4b could recruit diverse proteins through interaction with different domains. Furthermore, the functional association with RASSF1 indicates a role for PMCA4b in the modulation of Ras-mediated signaling.

Parkinson’s Is Time on Your Side? Evidence for Difficulties with Sensorimotor Synchronization

There is lack of consistent evidence as to how well PD patients are able to accurately time their movements across space with an external acoustic signal. For years, research based on the finger-tapping paradigm, the most popular paradigm for exploring the brain's ability to time movement, has provided strong evidence that patients are not able to accurately reproduce an isochronous interval [i.e., Ref. (1)]. This was undermined by Spencer and Ivry (2) who suggested a specific deficit in temporal control linked to emergent, rhythmical movement not event-based actions, which primarily involve the cerebellum. In this study, we investigated motor timing of seven idiopathic PD participants in event-based sensorimotor synchronization task. Participants were asked to move their finger horizontally between two predefined target zones to synchronize with the occurrence of two sound events at two time intervals (1.5 and 2.5 s). The width of the targets and the distance between them were manipulated to investigate impact of accuracy demands and movement amplitude on timing performance. The results showed that participants with PD demonstrated specific difficulties when trying to accurately synchronize their movements to a beat. The extent to which their ability to synchronize movement was compromised was found to be related to the severity of PD, but independent of the spatial constraints of the task.

Power Spectral Density Side-Channel Attack Overlapping Window Method

Cryptographic algorithms have been designed to be computationally secure, however it has been shown that when they are implemented in hardware, that these devices leak side channel information that can be used to mount an attack that recovers the secret encryption key. In this paper an overlapping window power spectral density (PSD) side channel attack, targeting an FPGA device running the Advanced Encryption Standard is proposed. This improves upon previous research into PSD attacks by reducing the amount of pre-processing (effort) required. It is shown that the proposed overlapping window method requires less processing effort than that of using a sliding window approach, whilst overcoming the issues of sampling boundaries. The method is shown to be effective for both aligned and misaligned data sets and is therefore recommended as an improved approach in comparison with existing time domain based correlation attacks.