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N-methyl-D-aspartate receptors (NMDAR) are glutamate-gated calcium channels named after their artificial agonist. NMDAR are implicated in cell proliferation under normal and pathophysiological conditions. However, the role of NMDAR during mitosis has not yet been explored in individual cells. We found that neurotransmitter-evoked calcium entry via endogenous NMDAR in cortical astrocytes was transient during mitosis. The same occurred in HEK293 cells transfected with the NR1/NR2A subunits of NMDAR. This transient calcium entry during mitosis was due to phosphorylation of the first intracellular loop of NMDAR (S584 of NR1 and S580 of NR2A) by cyclin B/CDK1. Expression of phosphomimetic mutants resulted in transient calcium influx and enhanced NMDAR inactivation independent of the cell cycle phase. Phosphomimetic mutants increased entry of calcium in interphase and generated several alterations during mitosis: increased mitotic index, increased number of cells with lagging chromosomes and fragmentation of pericentriolar material. In summary, by controlling cytosolic calcium, NMDAR modulate mitosis and probably cell differentiation/proliferation. Our results suggest that phosphorylation of NMDAR by cyclin B/CDK1 during mitosis is required to preserve mitotic fidelity. Altering the modulation of the NMDAR by cyclin B/CDK1 may conduct to aneuploidy and cancer.

Our results from quantitative RT-PCR, Western blotting, immunohistochemistry, and the tissue microarray of medullary thyroid cancer (MTC) cell lines and patient specimens confirm that VGSC subtype NaV1.7 is uniquely expressed in aggressive MTC and not expressed in normal thyroid cells and tissues. We establish the druggability of NaV1.7 in MTC by identifying a novel inhibitor (SV188) and investigate its mode of binding and ability to inhibit INa current in NaV1.7. The whole-cell patch-clamp studies of the SV188 in the NaV1.7 channels expressed in HEK-293 cells show that SV188 inhibited the INa current in NaV1.7 with an IC50 value of 3.6 µM by a voltage- and use-dependent blockade mechanism, and the maximum inhibitory effect is observed when the channel is open. SV188 inhibited the viability of MTC cell lines, MZ-CRC-1 and TT, with IC50 values of 8.47 μM and 9.32 μM, respectively, and significantly inhibited the invasion of MZ-CRC-1 cells by 35% and 52% at 3 μM and 6 μM, respectively. In contrast, SV188 had no effect on the invasion of TT cells derived from primary tumor, which have lower basal expression of NaV1.7. In addition, SV188 at 3 μM significantly inhibited the migration of MZ-CRC-1 and TT cells by 27% and 57%, respectively.

ObjectiveLow voltage-activated (LVA) calcium channels are crucial for regulating oscillatory behavior in several types of neurons and other excitable cells. LVA channels dysfunction has been implicated in epilepsy, neuropathic pain, cancer, among other diseases. Unlike for High Voltage-Activated (HVA) channels, voltage-dependence and kinetics of currents carried by recombinant LVA, i.e., CaV3 channels, are quite similar to those observed in native currents. Therefore, whether these channels are regulated by HVA auxiliary subunits, remain controversial. Here, we used the α1-subunits of CaV3.1, CaV3.2, and CaV3.3 channels, together with HVA auxiliary β-subunits to perform electrophysiological, confocal microscopy and immunoprecipitation experiments, in order to further explore this possibility.ResultsFunctional expression of CaV3 channels is up-regulated by all four β-subunits, although most consistent effects were observed with the β1b-subunit. The biophysical properties of CaV3 channels were not modified by any β-subunit. Furthermore, although β1b-subunits increased colocalization of GFP-tagged CaV3 channels and the plasma membrane of HEK-293 cells, western blots analysis revealed the absence of physical interaction between CaV3.3 and β1b-subunits as no co-immunoprecipitation was observed. These results provide solid evidence that the up-regulation of LVA channels in the presence of HVA-β1b subunit is not mediated by a high affinity interaction between both proteins.Electronic supplementary materialThe online version of this article (10.1186/s13104-018-3917-1) contains supplementary material, which is available to authorized users.

Polyhedrins are viral proteins present in a large family of baculoviruses that form occlusion bodies (polyhedra). These structures protect the virus particles from the outside environment until they are ingested by susceptible insects. Occluded viruses can sustain inclement weather for long periods of time. Therefore, the polyhedra is a natural preservative that keeps the viral structure intact at ambient temperature for years. In a previous study we identified the first 110 amino acids from polyhedrin (PH(1-110)) as a good candidate to carry antigens of interest. As a proof of concept, we produced a fusion protein with PH(1-110) and the green fluorescent protein (PH(1-110)GFP). The fusion protein associates spontaneously during its synthesis resulting in the formation of nanoparticles. Nasal immunization with these nanoparticles and in the absence of any adjuvant, results in a robust immune response with the production of IgG immunoglobulins that remained elevated for months and that selectively recognize the GFP but not PH(1-110). These results indicate that PH(1-110) is poorly immunogenic but capable of enhancing the immune response to GFP.