li, S. | bioRxiv Science

bioRxiv2026

The effect of melittin intervention on murine cervical cancer cells: An in-depth proteomics investigation

Melittin, the principal bioactive peptide of bee venom, exhibits promising antitumor activity, whereas its molecular mechanisms in cervical cancer remain incompletely understood. In this study, the biological effects and molecular responses of melittin in U14 cervical cancer cells were investigated using Astral data-independent acquisition (Astral-DIA)-based quantitative proteomics combined with molecular validation. The effects of melittin on cell migration, invasion, and cell death were evaluated by Transwell assays and PI/Hoechst staining. Differentially expressed proteins (DEPs) were screened and subjected to Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and protein-protein interaction (PPI) analyses. Representative oxidative stress-related genes and proteins were further validated by RT-qPCR and Western blotting. Melittin significantly inhibited the migration and invasion of U14 cervical cancer cells and increased cell death. Quantitative proteomics identified 9,782 protein groups and 187 DEPs, including 71 up- and 116 down-regulated proteins. KEGG pathway enrichment analysis revealed oxidative phosphorylation (OXPHOS) as the most significantly enriched pathway, together with glutathione metabolism, ferroptosis-related pathways, reactive oxygen species signaling, and mitophagy. GO term enrichment analysis indicated that DEGs were mainly engaged in mitochondrial function, electron transport, oxidoreductase activity, and energy metabolism. RT-qPCR assay demonstrated altered expression of Duox1, Gpx4, Gsx2, Nfe2l2, and Gstp2. Additionally, PPI analysis identified Gstp2 and ODC1 as representative hub proteins involved in redox regulation and metabolic adaptation. Furthermore, Western blotting confirmed increased GSTP2 expression following melittin treatment. Overall, these findings provide a comprehensive proteomic landscape of melittin-treated U14 cervical cancer cells and suggest that mitochondrial OXPHOS remodeling and redox-associated pathways may contribute to the antitumor activity of melittin.

cell biology↗

bioRxiv2020

Crystal and cryo-EM structures of the cytosolic G protein alpha chaperone and guanine nucleotide exchange factor Ric-8A bound to Gαi1

Ric-8A is a cytosolic Guanine Nucleotide exchange Factor (GEF) that activates heterotrimeric G protein alpha subunits (G)1. Ric-8A is essential to life in multicellular eukaryotes by virtue of its chaperone activity that is required for G biogenesis and membrane localization2, 3. Ric-8A adopts an armadillo (ARM)/HEAT repeat domain architecture and is structurally unrelated to G Protein-Coupled Receptors (GPCR)4. Both GEF and chaperone activities are stimulated by Casein Kinase II phosphorylation5. The mechanisms by which Ric-8A catalyzes GDP release and GTP binding to G, or exerts chaperone activity are unknown. Here, we report the structure of the nanobody-stabilized complex of nucleotide-free Gi1 (isoform 1 of G family i) and phosphorylated Ric-8A at near atomic resolution by cryo-electron microscopy and X-ray crystallography. We find that Ric-8A envelops the GTPase domain of G, disrupting all three switch regions that convey G nucleotide-binding and signaling activity, and displaces the C-terminal helix and helical domain of G. These cooperative interactions dismantle the GDP binding site and promote GDP release, while protecting structural elements of G that are dynamic in the nucleotide-free state. The structures also show how in vivo phosphorylation stabilizes G-binding elements of Ric-8A, thereby enhancing its GEF and chaperone activities.

biochemistry↗