The copines are a family of C2‐ and von Willebrand factor A‐domain‐containing proteins that have been proposed to respond to increases in intracellular calcium by translocating to the plasma membrane. The copines have been reported to interact with a range of cell signalling and cytoskeletal proteins, which may therefore be targeted to the membrane following increases in cellular calcium. However, neither the function of the copines, nor their actual movement to the plasma membrane, has been fully established in mammalian cells. Here, we show that copines‐1, ‐2, ‐3, ‐6 and ‐7 respond differently to a methacholine‐evoked intracellular increase in calcium in human embryonic kidney cell line‐293 cells, and that their membrane association requires different levels of intracellular calcium. We demonstrate that two of these copines associate with different intracellular vesicles following calcium entry into cells, and identify a novel conserved amino acid sequence that is required for their membrane translocation in living cells. Our data show that the von Willebrand factor A‐domain of the copines modulates their calcium sensitivity and intracellular targeting. Together, these findings suggest a different set of roles for the members of this protein family in mediating calcium‐dependent processes in mammalian cells.
Structured digital abstract
http://mint.bio.uniroma2.it/mint/search/interaction.do?interactionAc=MINT-8049236: Copine‐6 (uniprotkb:http://www.uniprot.org/uniprot/Q9Z140) and transferrin (uniprotkb:http://www.uniprot.org/uniprot/P02787) colocalize (http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0403) by fluorescence microscopy (http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0416)
http://mint.bio.uniroma2.it/mint/search/interaction.do?interactionAc=MINT-8049176: CD2 (uniprotkb:http://www.uniprot.org/uniprot/P06729) and Copine‐2 (uniprotkb:http://www.uniprot.org/uniprot/P59108) colocalize (http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0403) by fluorescence microscopy (http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0416)
In the present study, the effects of intraperitoneal injections of GABAA receptor agonist and antagonist on memory formation and morphine state-dependent learning were investigated in rats. Pre-training administration of morphine (1–15 mg/kg) in a step-down passive avoidance task induced state-dependent learning with impaired memory retrieval on the test day. The impairment of memory was restored after the pre-test administration of the same dose of morphine. The pre-test administration of the GABAA receptor agonist, muscimol (0.01, 0.05 and 0.1 mg/kg), significantly decreased state-dependent retrieval induced by pre-test morphine (5 mg/kg). The state-dependency effect of morphine (1 mg/kg) was significantly potentiated by the pre-test administration of the GABAA receptor antagonist, bicuculline (0.125, 0.25 and 0.5 mg/kg). Furthermore, the pre-training injection of muscimol (0.01 mg/kg) impaired memory retrieval which was restored by pre-test morphine (1, 3 and 5 mg/kg) administration. However, the pre-training administration of bicuculline did not affect retention by itself. In addition, amnesia induced by pre-training morphine (5 mg/kg) was significantly reversed in rats which had received pre-test injections of muscimol (0.01, 0.05 and 0.1 mg/kg). Pre-test injections of bicuculline (0.125, 0.25 and 0.5 mg/kg) significantly decreased morphine-induced amnesia. It is concluded that the GABAA receptor mechanisms may be involved in the memory formation and it is postulated that these receptors may play an important role in morphine state-dependent learning.
Alzheimer’s disease (AD) is a neurodegenerative disease with cognitive impairment. Oxidative stress in neurons is considered as a reason for development of AD. Antioxidant agents such as quercetin slow down AD progression, but the usage of this flavonoid has limitations because of its low bioavailability. We hypothesized that quercetin-conjugated superparamagnetic iron oxide nanoparticles (QT-SPIONs) have a better neuroprotective effect on AD than free quercetin and regulates the antioxidant, apoptotic, and APP gene, and miRNA-101. In this study, male Wistar rats were subjected to AlCl3, AlCl3 + QT, AlCl3 + SPION, and AlCl3 + QT-SPION for 42 consecutive days. Behavioral tests and qPCR were used to evaluate the efficiency of treatments. Results of behavioral tests revealed that the intensity of cognitive impairment was decelerated at both the middle and end of the treatment period. The effect of QT-SPIONs on learning and memory deficits were closely similar to the control group. The increase in expression levels of APP gene and the decrease in mir101 led to the development of AD symptoms in rats treated with AlCl3 while these results were reversed in the AlCl3 + QT-SPIONs group. This group showed similar results with the control group. QT-SPION also decreased the expression levels of antioxidant enzymes along with increases in expression levels of anti-apoptotic genes. Accordingly, the antioxidant effect of QT-SPION inhibited progression of cognitive impairment via sustaining the balance of antioxidant enzymes in the hippocampus of AD model rats.
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