Biology subjects

Tikhonova, I.

Publications and source records attributed to Tikhonova, I..

6 recordsLinked to original sources

bioRxiv2026

Chromosomal-Level Whole Genome Assemblies of Crassostrea Virginica

The eastern oyster, Crassostrea virginica, is an ecologically important foundation species and a valuable aquaculture organism. To support genomic studies and future breeding applications, we generated PacBio HiFi, Illumina, and Hi-C sequencing data from wild adult oysters, collected from Long Island Sound and assembled a chromosome-level reference genome. The final assembly spans 529.0 Mb and consists of 10 chromosomes with a scaffold N50 of 55.46 Mb and a contig N50 of 16.0 Mb. The resulting reference genome achieved 99.7% genome completeness and reduced duplicated universal single-copy ortholog content from 18.7% to 1.1%. Genome annotation identified 33,259 protein-coding genes and revealed that 43.2% of the genome is composed of repetitive elements. All sequencing data underwent stringent quality control, and the assembly results were validated through multiple technical evaluations. These dataset and resources of oyster genomics provided future studies such as effective breeding and sustainable farming.

genomics↗

bioRxiv2025

Structural basis and physiological significance of non-canonical Gs coupling to the prototypical Gi-coupled melatonin MT1 receptor

G protein-coupled receptors (GPCRs) transduce extracellular stimuli into intracellular signals by coupling to various heterotrimeric G proteins. However, the rules governing G protein preference remain largely elusive. MT1 and MT2 are prototypical Gi/o-coupled GPCRs responding to melatonin, a hormone secreted in a circadian manner. We show here that MT1, but not MT2, couples also to Gs proteins in vitro and activates the Gs/cAMP pathway upon long-term melatonin exposure in vivo, mimicking physiological dawn conditions. We solved the cryo-electron microscopy structure of the melatonin-MT1-Gs complex at 3.0[A] resolution, which revealed a strikingly distinct binding mode compared to the MT1-Gi complex. The third intracellular loop of MT1 emerges as a key stabilizer for Gs coupling, a feature previously unrecognized. This is the first solved receptor-Gs complex of a primary Gi-coupled GPCRs, providing new structural and functional insights into G protein selectivity and circadian switch of G protein coupling.

molecular biology↗

bioRxiv2025

Steric control of signaling bias in the immunometabolic receptor GPR84

Biased signaling in G protein-coupled receptors offers therapeutic promise, yet rational design of biased ligands remains challenging due to limited mechanistic understanding. Here, we report a molecular framework for controlling signaling bias at the immunometabolic receptor GPR84. We identified three structurally-matched ligands (OX04529, OX04954, and OX04539) with varying steric profiles that exhibit comparable Gi protein activation but dramatically different {beta}-arrestin recruitment capacities. A high- resolution cryo-EM structure of GPR84-Gi in complex with OX04529, complemented by molecular dynamics simulations and targeted mutagenesis, revealed that steric interactions between ligand substituents and Leu3366.52 and Phe1875.47 indirectly disrupt a critical polar network involving Tyr3326.48, Asn1043.36 and Asn3627.45 essential for {beta}-arrestin recruitment. Based on these insights, we developed a steric-dependent model that enabled rational design of G protein-biased agonists with predictable {beta}-arrestin recruitment profiles. This mechanistic framework provides a blueprint for designing biased agonists with customized signaling profiles at GPR84 and potentially other class A GPCRs.

pharmacology and toxicology↗

bioRxiv2023

Structural basis for the ligand recognition and signaling of free fatty acid receptors

Free fatty acid receptors 1-4 (FFA1-4) are class A G protein-coupled receptors (GPCRs). FFA1-3 share substantial sequence similarity whereas FFA4 is unrelated. Despite this FFA1 and FFA4 are activated by the same range of long chain fatty acids (LCFAs) whilst FFA2 and FFA3 are instead activated by short chain fatty acids (SCFAs) generated by the intestinal microbiota. Each of FFA1, 2 and 4 are promising targets for novel drug development in metabolic and inflammatory conditions. To gain insights into the basis of ligand interactions with, and molecular mechanisms underlying activation of, FFAs by LCFAs and SCFAs, we determined the active structures of FFA1 and FFA4 bound to the polyunsaturated LCFA docosahexaenoic acid (DHA), FFA4 bound to the synthetic agonist TUG-891, as well as SCFA butyrate-bound FFA2, each complexed with an engineered heterotrimeric Gq protein (miniGq), by cryo-electron microscopy. Together with computational simulations and mutagenesis studies, we elucidated the similarities and differences in the binding modes of fatty acid ligands with varying chain lengths to their respective GPCRs. Our findings unveil distinct mechanisms of receptor activation and G protein coupling. We anticipate that these outcomes will facilitate structure-based drug development and underpin future research to understand allosteric modulation and biased signaling of this group of GPCRs.

pharmacology and toxicology↗

bioRxiv2023

β-arrestin-dependent and -independent endosomal G protein activation by the vasopressin type 2 receptor

The vasopressin type 2 receptor (V2R) is an essential GPCR in renal regulation of water homeostasis. Upon stimulation, the V2R activates Gs and Gq/11, which is followed by robust recruitment of {beta}-arrestins and receptor internalization into endosomes. Unlike canonical GPCR signaling, the {beta}-arrestin association with the V2R does not terminate Gs activation, and thus, Gs-mediated signaling is sustained while the receptor is internalized. Here, we demonstrate that this V2R ability to co-interact with G protein/{beta}-arrestin and promote endosomal G protein signaling is not restricted to Gs, but also involves Gq/11. Furthermore, our data implies that {beta}-arrestins potentiate Gs/Gq/11 activation at endosomes rather than terminating their signaling. Surprisingly, we found that the V2R internalizes and promote endosomal G protein activation independent of {beta}-arrestins to a minor degree. These new observations challenge the current model of endosomal GPCR signaling and suggest that this event can occur in both {beta}-arrestin-dependent and -independent manners. IMPACT STATEMENTThe vasopressin type 2 receptor promotes dual Gs and Gq/11 signaling at early endosomes in {beta}-arrestin-dependent and -independent manners.

cell biology↗

bioRxiv2021

The immunophilin protein FKBPL and its peptide derivatives are novel regulators of vascular integrity and inflammation via NF-kappaB signaling

A breakdown in vascular integrity and excessive inflammation are hallmarks of serious pathological conditions including sepsis, acute respiratory distress syndrome (ARDs) and most recently, severe COVID-19. FK506 - binding protein like (FKBPL) is a member of the immunophilin protein superfamily with potent anti-tumor activity through inhibition of angiogenesis and cancer stemness. An FKBPL-based 23mer peptide, ALM201, displayed a good safety and pharmacokinetic profile in a Phase 1a oncology clinical trial and was subsequently designated orphan drug status by the FDA in ovarian cancer. Here we describe a novel role for FKBPL and its peptides in regulating vascular integrity and cytokine production though modulating NF-{kappa}B signaling. FKBPL knockdown promoted endothelial cell barrier permeability, which was further exacerbated upon stimulation with lipopolysaccharide (LPS) and accompanied by increased expression of TNF mRNA and phosphorylation of p65(RelA). Whilst treatment with the FKBPL based pre-clinical peptide, AD-01, increased VE-cadherin endothelial tight junctions following LPS stimulation. Bone marrow derived macrophages (BMDM) from FKBPL haploinsufficient mice (Fkbpl+/-) also demonstrated increased phosphorylation of p65(RelA) in response to LPS stimulation compared to wild-type mice. Furthermore, treatment with AD-01 inhibited p65(RelA) phosphorylation following LPS stimulation resulting in reduced NF-{kappa}B target gene expression and proinflammatory cytokine production. In an in vivo LPS survival model, Fkbpl+/- mice have reduced survival compared to wild-type mice. Moreover, treatment of wild-type mice with the clinical FKBPL-based peptide, ALM201, following LPS injection resulted in a 100% survival rate in mice at experimental endpoint, as well as an abrogation of production of pro-inflammatory cytokines, TNF and IL-6, in peritoneal lavage washings. Analysis of human genetic biobanks found an association between common genetic variants associated with FKBPL and traits associated with inflammatory disorders such as psoriasis, rheumatoid arthritis and high lymphocyte count. In summary, for the first time, we describe a novel role for FKBPL as a regulator of inflammation and vascular integrity through modulating NF-{kappa}B signaling and FKBPL based therapies demonstrate potent anti-inflammatory activity.

immunology↗