Biology subjects

Fernandez-Fernandez, J.

Publications and source records attributed to Fernandez-Fernandez, J..

3 recordsLinked to original sources

bioRxiv2026

Ribosomal protein eL22 contributes to the assembly of 60S ribosomal subunits in Saccharomyces cerevisiae

Ribosome biogenesis is a highly coordinated pathway that involves the assembly of ribosomal RNAs (rRNAs) with ribosomal proteins (r-proteins) to generate functional ribosomal subunits (r-subunits). The Saccharomyces cerevisiae (yeast) large 60S r-subunit consists of three rRNA molecules and 46 r-proteins. The contributions of nearly all r-proteins of the yeast large r-subunit have been characterized; however, a few non-essential proteins remain poorly understood. Although non-essential, human eL22 has been identified as a key player in p53 regulation during ribosomal stress and as a highly mutated target in cancers. Despite this function, the role of eL22 in ribosome maturation is still ill-defined. In this study, we characterized yeast eL22 r-protein. Our results show that eL22 assembles into intermediate nucleolar pre-60S ribosomal particles. Loss of eL22 impairs cell growth and reduces 60S r-subunit accumulation, phenotypes that are exacerbated at low temperatures. Analysis of pre-rRNA processing by pulse-chase labeling, northern blot hybridization, and primer extension reveals a defect in 27S pre-rRNA maturation, specifically at the level of 27SB pre-rRNA processing. Consequently, nuclear export of eL22-deficient pre-60S particles is mildly impaired. Furthermore, we identify genetic interactions between eL22 and neighboring r-proteins, eL38 and eL31. We conclude that eL22 assembly is required for optimal pre-60S maturation during middle nucleolar stages, particularly at low temperatures, a function likely supported by the cooperative action of other r-proteins associated with common elements of 25S rRNA. HighlightsO_LIWe have studied the role of r-protein eL22 in yeast ribosome assembly. C_LIO_LIeL22 is required for 60S ribosomal subunit production. C_LIO_LIThe absence of eL22 is critical at low temperatures. C_LIO_LIeL22 is important for 27SB pre-rRNA processing and nuclear export of pre-ribosomes. C_LIO_LIeL22 functionally interacts with r-proteins eL38 and eL31 in domain III of 25S rRNA. C_LI

genetics↗

bioRxiv2026

Otenabant is a Selective Antagonist of Human PIEZO1

Background and purposePIEZO1 mechanosensitive cation channels translate mechanical cues into intracellular Ca2+ and Na+ elevations, enabling cells to respond to physical alterations in their environment. PIEZO1 contributes to red blood cells (RBC) volume homeostasis and gain-of-function PIEZO1 mutations cause hereditary xerocytosis (HX), a rare mostly compensated hemolytic anemia, and aberrant channel activation exacerbates sickling and vascular dysfunction in sickle cell disease. Despite strong genetic and physiological evidence supporting PIEZO1 as a therapeutic target, potent and selective inhibitors are limited, and existing compounds show modest specificity or poorly explored mechanisms. Improved pharmacological tools are needed. Experimental approachWe conducted a high-throughput screen of FDA-approved drugs to identify PIEZO1 inhibitors. Compounds were tested at concentrations of 10 {micro}M in a monocytic cell line, using intracellular Ca2+ elevations evoked by the PIEZO1 agonist Yoda1 as read-out. The inhibitory activity of the best hit was validated and compared to existing PIEZO1 inhibitors using electrophysiological analysis, orthogonal PIEZO1-dependent assays across cell lines and human RBCs. As functional proof, we investigated the impact of three PIEZO1 inhibitors on RBC deformability by ektacytometry, after Yoda1 pre-stimulation. Key resultsThis screen identified Otenabant, a selective Cannabinoid Receptor Type 1 (CB1) antagonist, as a potent PIEZO1 inhibitor. Otenabant dose-dependently inhibited Ca2+ elevations mediated by endogenous or exogenously expressed human PIEZO1, but was ineffective against mouse Piezo1, revealing species-specific channel differences. Otenabant inhibited mechanosensitive currents elicited by shear stress in fibroblasts and by repeated poking in PIEZO1-expressing HEK-293 cells, altering the currents activation and inactivation kinetics, and prevented Yoda1-induced hyperpolarization in RBCs. Otenabant was able to reverse the negative impact of Yoda1 on RBC deformability. Conclusions and implicationsThese findings demonstrate the utility of Yoda-based screening for discovering PIEZO1 antagonists and identify Otenabant as a promising chemical scaffold for developing selective PIEZO1 inhibitors with therapeutic potential.

pharmacology and toxicology↗

bioRxiv2025

Topology-Encoded Polarity in Oppositely Charged Binary IDPP Condensates: Multiphase Organization from Non-Coacervating Partners as a Minimal Model of Complex Coacervation

Synthetic condensates provide a way to engineer compartmentalized microenvironments that mimic the properties and functions of natural ones, yet the principles that govern their phase behavior and internal organization remain incompletely defined. Introducing charged residues into intrinsically disordered protein polymers (IDPPs) with LCST phase behavior typically suppresses phase separation under physiological conditions. Here we show that pairing two such oppositely charged IDPPs restores and programs LCST-driven liquid-liquid phase separation (LLPS), enabling a minimalist two-component platform for constructing synthetic condensates whose formation, size, and internal organization are encoded directly in sequence. LLPS emerges from an asymmetric, entropy-driven interplay between hydrophobic collapse, solvent reorganization, and salt-bridge topology. The balance between inter- and intrachain ionic pairing leads to distinct dense-phase microenvironments with tunable residual charge and micropolarity, thereby controlling condensate formation, and miscibility and the emergence of single-phase or multiphase protein condensates. The condensate interior further alters the ionization thermodynamics of charged residues shifting their apparent pKa and enabling tunable pH responses. Systems dominated by interchain salt bridges form low-polarity condensates that mix uniformly with hydrophobic partners, whereas molecular architectures favoring intrachain pairing retain residual charge and, in the presence of hydrophobic partners, undergo spontaneous internal demixing into multiphase assemblies. These findings establish a mechanistic, sequence-level framework for encoding phase behavior, micropolarity, and mesoscale organization in synthetic condensates, and demonstrate how minimalistic LCST-IDPP pairs can be engineered to create programmable microenvironments, opening avenues toward engineered condensates with higher-order organization and adaptive capabilities.

biophysics↗