Publications by authors named "K Gusev"

26 Publications

Role of STIM2 and Orai proteins in regulating TRPC1 channel activity upon calcium store depletion.

Cell Calcium 2021 Jun 8;97:102432. Epub 2021 Jun 8.

Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Avenue, St. Petersburg 194064, Russia. Electronic address:

Store-operated calcium channels are the major player in calcium signaling in non-excitable cells. Store-operated calcium entry is associated with the Orai, stromal interaction molecule (STIM), and transient receptor potential canonical (TRPC) protein families. Researchers have provided conflicting data about TRPC1 channel regulation by Orai and STIM. To determine how Orai and STIM influence endogenous TRPC1 pore properties and regulation, we used single channel patch-clamp recordings. Here we showed that knockout or knockdown of Orai1 or Orai3 or overexpression of the dominant-negative mutant Orai1 E106Q did not change the conductance or selectivity of single TRPC1 channels. In addition, these TRPC1 channel properties did not depend on the amount of STIM1 and STIM2 proteins. To study STIM2-mediated regulation of TRPC1 channels, we utilized partial calcium store depletion induced by application of 10 nM thapsigargin (Tg). TRPC1 activation by endogenous STIM2 was greatly decreased in acute extracellular calcium-free experiments. STIM2 overexpression increased both the basal activity and number of silent TRPC1 channels in the plasma membrane. After calcium store depletion, overexpressed STIM2 directly activated TRPC1 in the plasma membrane even without calcium entry in acute experiments. However, this effect was abrogated by co-expression with the non-permeable Orai1 E106Q mutant protein. Taken together, our single-channel patch clamp experiments clearly demonstrated that endogenous TRPC1 forms a channel pore without involving Orai proteins. Calcium entry through Orai triggered TRPC1 channel activation in the plasma membrane, while subsequent STIM2-mediated TRPC1 activity regulation was not dependent on calcium entry.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ceca.2021.102432DOI Listing
June 2021

Electrophysiological Properties of Endogenous Single Ca Activated Cl Channels Induced by Local Ca Entry in HEK293.

Int J Mol Sci 2021 Apr 30;22(9). Epub 2021 Apr 30.

Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Avenue, 194064 St. Petersburg, Russia.

Microdomains formed by proteins of endoplasmic reticulum and plasma membrane play a key role in store-operated Ca entry (SOCE). Ca release through inositol 1,4,5-trisphosphate receptor (IPR) and subsequent Ca store depletion activate STIM (stromal interaction molecules) proteins, sensors of intraluminal Ca, which, in turn, open the Orai channels in plasma membrane. Downstream to this process could be activated TRPC (transient receptor potential-canonical) calcium permeable channels. Using single channel patch-clamp technique we found that a local Ca entry through TRPC1 channels activated endogenous Ca-activated chloride channels (CaCCs) with properties similar to Anoctamin6 (TMEM16F). Our data suggest that their outward rectification is based on the dependence from membrane potential of both the channel conductance and the channel activity: (1) The conductance of active CaCCs highly depends on the transmembrane potential (from 3 pS at negative potentials till 60 pS at positive potentials); (2) their activity (NPo) is enhanced with increasing Ca concentration and/or transmembrane potential, conversely lowering of intracellular Ca concentration reduced the open state dwell time; (3) CaCC amplitude is only slightly increased by intracellular Ca concentration. Experiments with Ca buffering by EGTA or BAPTA suggest close local arrangement of functional CaCCs and TRPC1 channels. It is supposed that Ca-activated chloride channels are involved in Ca entry microdomains.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/ijms22094767DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8124839PMC
April 2021

STIM2 Mediates Excessive Store-Operated Calcium Entry in Patient-Specific iPSC-Derived Neurons Modeling a Juvenile Form of Huntington's Disease.

Front Cell Dev Biol 2021 2;9:625231. Epub 2021 Feb 2.

Laboratory of Ionic Channels of Cell Membranes, Department of Molecular Physiology of the Cell, Institute of Cytology, Russian Academy of Sciences, St. Petersburg, Russia.

Huntington's disease (HD) is a severe autosomal-dominant neurodegenerative disorder caused by a mutation within a gene, encoding huntingtin protein. Here we have used the induced pluripotent stem cell technology to produce patient-specific terminally differentiated GABA-ergic medium spiny neurons modeling a juvenile form of HD (HD76). We have shown that calcium signaling is dramatically disturbed in HD76 neurons, specifically demonstrating higher levels of store-operated and voltage-gated calcium uptakes. However, comparing the HD76 neurons with the previously described low-repeat HD models, we have demonstrated that the severity of calcium signaling alterations does not depend on the length of the polyglutamine tract of the mutant huntingtin. Here we have also observed greater expression of huntingtin and an activator of store-operated calcium channels STIM2 in HD76 neurons. Since shRNA-mediated suppression of STIM2 decreased store-operated calcium uptake, we have speculated that high expression of STIM2 underlies the excessive entry through store-operated calcium channels in HD pathology. Moreover, a previously described potential anti-HD drug EVP4593 has been found to attenuate high levels of both huntingtin and STIM2 that may contribute to its neuroprotective effect. Our results are fully supportive in favor of the crucial role of calcium signaling deregulation in the HD pathogenesis and indicate that the cornerstone of excessive calcium uptake in HD-specific neurons is a calcium sensor and store-operated calcium channels activator STIM2, which should become a molecular target for medical treatment and novel neuroprotective drug development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fcell.2021.625231DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7884642PMC
February 2021

Final Results of GERDA on the Search for Neutrinoless Double-β Decay.

Phys Rev Lett 2020 Dec;125(25):252502

Max-Planck-Institut für Physik, 80805 Munich, Germany.

The GERmanium Detector Array (GERDA) experiment searched for the lepton-number-violating neutrinoless double-β (0νββ) decay of ^{76}Ge, whose discovery would have far-reaching implications in cosmology and particle physics. By operating bare germanium diodes, enriched in ^{76}Ge, in an active liquid argon shield, GERDA achieved an unprecedently low background index of 5.2×10^{-4} counts/(keV kg yr) in the signal region and met the design goal to collect an exposure of 100 kg yr in a background-free regime. When combined with the result of Phase I, no signal is observed after 127.2 kg yr of total exposure. A limit on the half-life of 0νββ decay in ^{76}Ge is set at T_{1/2}>1.8×10^{26}  yr at 90% C.L., which coincides with the sensitivity assuming no signal.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1103/PhysRevLett.125.252502DOI Listing
December 2020

Impact of the DSP-H1684R Genetic Variant on Ion Channels Activity in iPSC-Derived Cardiomyocytes.

Cell Physiol Biochem 2020 Jul;54(4):696-706

Almazov National Medical Research Centre of the Ministry of Health of the Russian Federation, St. Petersburg, Russia.

Background/aims: Mutations of desmosomal genes are known to cause arrhythmogenic cardiomyopathy characterized by arrhythmias and sudden cardiac death. Previously, we described a novel genetic variant H1684R in desmoplakin gene (DSP), associated with a progressive cardiac conduction disease (PCCD). In the present study, we aimed to investigate an effect of the DSP-H1684R genetic variant on the activity of ion channels.

Methods: We used cardiomyocytes derived from induced pluripotent stem cells (iPSC cardiomyocytes) from a patient with DSP-H1684R genetic variant and from two healthy donors. Immunofluorescent staining and western blot analyses were used to characterize patient-specific cardiomyocytes. By the whole-cell voltage-clamp technique we estimated the activity of voltage-gated sodium, calcium, and potassium channels that are responsible for action potential generation and its shape. Action potentials' parameters were measured using whole-cell current-clamp technique.

Results: In patient-specific cardiomyocytes we observed both lower amplitudes of currents through sodium Na1.5 channels and L-type calcium channels, but higher amplitude of current through transient-outward potassium channels in comparison to donor cardiomyocytes. Current-clamp measurements revealed shortening of action-potential in DSP-H1684R-carrying iPSC cardiomyocytes. Therefore, observed alterations in the channels activity might have a great impact on the properties of action potential and development of PCCD.

Conclusion: Our results show that desmoplakin genetic variants, besides conduction slowing caused by structural heart remodeling, could affect multiple ion channel activity aggravating arrhythmia manifestation in PCCD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.33594/000000249DOI Listing
July 2020
-->