RESUMO
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.
RESUMO
Affinity for G-quadruplex (G4) structures may be a common feature of transcription-facilitating histone chaperons (HCs). This assumption is based on previous unmatched studies of HCs FACT, nucleolin (NCL), BRD3, and ATRX. We verified this assumption and considered its implications for the therapeutic applications of synthetic (exogenous) G4s and the biological significance of genomic G4s. First, we questioned whether exogenous G4s that recognize cell-surface NCL and could trap other HCs in the nucleus are usable as anticancer agents. We performed in vitro binding assays and selected leading multi-targeted G4s. They exhibited minor effects on cell viability. The presumed NCL-regulated intracellular transport of G4s was inefficient or insufficient for tumor-specific G4 delivery. Next, to clarify whether G4s in the human genome could recruit HCs, we compared available HC ChIP-seq data with G4-seq/G4-ChIP-seq data. Several G4s, including the well-known c-Myc quadruplex structure, were found to be colocalized with HC occupancy sites in cancer cell lines. As evidenced by our molecular modeling data, c-Myc G4 might interfere with the HC function of BRD3 but is unlikely to prevent the BRD3-driven assembly of the chromatin remodeling complex. The c-Myc case illustrates the intricate role of genomic G4s in chromatin remodeling, nucleosome remodeling, and transcription.
