Work

Biophysical characterization and ion transport with cell‐based and proteoliposome reconstitution assays of invertebrate K + ‐Cl − cotransporters

Year: 2025

Type: article

Abstract: The cation‐chloride cotransporter (CCC) family includes ion symporters that cotransport monovalent cations and Cl − , playing a crucial role in controlling cytoplasmic ion content. K + ‐Cl − cotransporters (KCCs) facilitate the symport of ions across the plasma membrane. The CCCs participate in various physiological processes, such as transepithelial ion transport and regulation of cell volume. Among KCCs, KCC2 has unique and essential functions in the central nervous system. KCC from Drosophila melanogaster ( Dm KCC) is an ortholog of mammalian KCCs. Its critical role in neuronal transmission has been demonstrated. Also, the cnidarian Hydra vulgaris has a functional KCC ( Hv KCC). Comparative analyses of these transporters with vertebrate counterparts can provide insights into the mechanism of KCC ion transport, regulation, and evolution. Thus, here we purified Dm KCC and Hv KCC and characterized their biophysical properties using differential scanning fluorimetry and light scattering. We evaluated their functionality in cells and developed a method to study ion transport with flame photometry. Further, a fluorescence‐based assay for Dm KCC reconstituted into proteoliposomes was developed. The activity of Dm KCC was found to be dependent on Ca 2+ , which is reminiscent of some other chloride transport protein families and potentially important for the KCC protein family overall.

Source: FEBS Open Bio

DOI: 10.1002/2211-5463.70063

Authors: Satoshi Fudo, Marina Verkhovskaya, Coralie Di Scala, Claudio Rivera, Tommi Kajander

Institutions: University of Helsinki, HiLIFE – Elämäntieteiden Instituutti, Inserm, Aix-Marseille Université, Institut de Neurobiologie de la Méditerranée

Language: English

Topic: Ion Transport and Channel Regulation

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: gold

Estimated APC paid: $2,210