Biology subjects

Belyaev, E. S.

Publications and source records attributed to Belyaev, E. S..

4 recordsLinked to original sources

bioRxiv2026

Cyanine dye conjugates of a 2'-deoxycytidine-based auto- and mitophagy activator extend Caenorhabditis elegans lifespan

BackgroundAutophagy and mitophagy are essential for cellular homeostasis and play key roles in longevity and healthy aging, whereas their age-associated decline contributes to the development of age-related diseases. The identification of small-molecule activators of these pathways therefore represents an important therapeutic objective. MethodsIn this study, we investigated a series of compounds based on a 2'-deoxycitidine-derived scaffold and systematically analyzed the impact of structural substitutions on their ability to induce autophagy and mitophagy. Chemical optimization and functional assays were combined with pathway analysis, cellular readouts of proteostasis, and in vivo lifespan assessment in Caenorhabditis elegans. ResultsThe lead compound enhanced autophagy predominantly via activation of the AMPK-ULK1 signaling pathway and induced mitophagy in a Parkin-independent manner. It promoted autophagosome formation and facilitated functional clearance of aggregation-prone mutant huntingtin. Conjugation of the lead compound with the mitochondria-targeting Cy5 dye further potentiated mitophagy induction, likely through preferential mitochondrial accumulation, while reducing cytotoxicity. Importantly, the conjugated compound significantly extended C. elegans lifespan at lower concentrations compared with the unconjugated analogue. ConclusionsTogether, these results identify a promising chemical scaffold for the development of auto-and mitophagy activators and validate mitochondria-targeted conjugation as an effective strategy to enhance their biological performance. The demonstrated in vivo efficacy supports the potential relevance of these compounds for interventions aimed at preserving proteostasis and mitochondrial quality control, with possible implications for geroprotective applications.

pharmacology and toxicology↗

bioRxiv2026

An Adenine-Based Molecular Rotor as a Universal Fluorescent Nucleobase with High Brightness

Chemically modified nucleic acids have become a powerful platform for basic research and applied technologies. Universal nucleobases are used in PCR,sequencing, and the design of nanodevices and aptamers. Fluorescent universal nucleobases have an even wider range of applications, including the development of nucleic acid-based sensors, switches, and relay logic gates. However, few such nucleobases have been proposed to date, and most of them have suboptimal optical properties. Here, we propose an adenine-based molecular rotor, 7,8-dihydro-8-oxo-6-(3-methylbenzo[d]thiazol-2(3H)-ylidene)adenine (oxo-Ade BZT), as a new, remarkably bright and potent fluorescent universal nucleobase. Its brightness in both oligodeoxyribonucleotides (ODNs) and DNA duplexes (4200 - 10000 M-1 x cm-1) originates from a high molar extinction coefficient (averaged{varepsilon} 368 37000 M-1 x cm-1), provided by the appended 3-methylbenzo[d]thiazolyl moiety, and a relatively high quantum yield (0.11 - 0.27). Melting temperature variations observed upon the incorporation of oxo-Ade BZT opposite native nucleobases in a duplex context did not exceed 10%. The basis of these universal hybridizing properties was unveiled using computational methods. According to molecular dynamics simulations, oxo-Ade BZT pushes the opposite nucleobase out of the DNA double helix and forms multiple hydrophobic contacts with the flanking base pairs. At the same time, the rotational mobility of the bonds between the oxo-Ade BZT-constituting heterobicycles decreases, and oxo-Ade BZT adopts a planar conformation in both ODNs and their duplexes, resulting in the light-up effect. These properties make oxo-Ade BZT a promising molecular tool for analytical, biophysical and biochemical studies.

biophysics↗

bioRxiv2024

Truncated and "double-headed" derivatives of TO1-B dye for Mango-based imaging systems with increased brightness and selectivity and large Stocks shift

Fluorogenic dyes that light up in complexes with genetically encodable RNA aptamers are increasingly used for intracellular RNA imaging. The growing toolbox of dyes and aptamers allows simultaneous or sequential monitoring of different RNAs. However, few orthogonal aptamer-dye pairs meet all the requirements of multiplex imaging in terms of brightness, contrast, etc., thus necessitating their optimization and further diversification. Here, we report new fluorogenic ligands for the Mango II aptamer, one of the most stable and widely used RNA tags. They are based on the cognate Mango ligand TO1-B, which is a thiazole orange (TO) derivative with a biotinylated tetraethylene glycol (TEG) residue attached via an amide linker, and its analogue with an isosteric triazolyl linker (TO1-triazolyl-TEG-biotin). Structural data suggested that the biotin residue might be dispensable for interactions with Mango, so we replaced it with a second TO residue or the alternative push-pull system, namely the dimethylaminophenyl (DMAP) group linked to the benzothiazolyl (BzT) group via the ethenyl linker. The resulting "double-headed" symmetric and asymmetric dyes showed reduced fluorescence in the free state due to intramolecular TO/TO or TO/DMAP-BzT interactions. The asymmetric dye exhibited intramolecular FRET in complex with Mango, resulting in a remarkably large (130 nm) Stocks shift, which may be advantageous for multiplex imaging. The main limitation of the "double-headed" dyes was their low brightness. To improve brightness, we further optimized the truncated (biotin-free) TO1-triazolyl-TEG dye by introducing a hydroxy or a methoxy group into the methylqunolinium (MQ) fragment of its push-pull system. The brightness of the methoxy-MQ derivative was increased by 40% compared to the reported TO1-triazolyl-TEG-biotin. Importantly, the methoxy-MQ derivative also showed increased selectivity for Mango over other noncanonical nucleic acids structures, making it a promising alternative to known TO-based dyes for high-contrast RNA imaging.

biophysics↗

bioRxiv2022

Red light-emitting short Mango-based system enables tracking a mycobacterial small noncoding RNA in infected macrophages

Progress in the studies of RNA metabolism and function relies largely on molecular imaging systems, including those comprising a fluorogenic dye and an aptamer-based fluorescence-activating tag. G4 aptamers of Mango family, typically combined with a duplex/hairpin scaffold, activate fluorescence of a green-emitting dye TO1-biotin and hold great promise for intracellular RNA tracking. Here, we report a new Mango-based imaging platform. Its key advantages are tunability of spectral properties and applicability for visualization of small RNA molecules that require minimal tag size. The former advantage is due to an expanded (green-to-red-emitting) palette of TO1-inspired fluorogenic dyes, and the latter is ensured by the truncated duplex scaffold. To illustrate applicability of the improved platform, we tagged Mycobacterium tuberculosis sncRNA with the shortened aptamer-scaffold tag and visualized it in bacteria and bacteria-infected macrophages using the new red light-emitting Mango-activated dye.

synthetic biology↗