Biology subjects

Usak, D.

Publications and source records attributed to Usak, D..

2 recordsLinked to original sources

bioRxiv2026

MILDEW RESISTANCE LOCUS O (MLO) proteins function as trimeric inward calcium channels

Calcium signalling and structural roles are fundamental for plant growth, development, and environmental adaptation. Recent studies have identified MILDEW RESISTANCE LOCUS O (MLO) proteins as novel calcium-permeable channels with roles in root growth, cell wall development, pollen tube growth, and perception. However, the molecular mechanisms underlying MLO function remain unknown. Here, we demonstrate that multimerisation is essential for MLO activity. Chemical crosslinking, split-ubiquitin interaction assays, and single-molecule photobleaching revealed that MLO proteins form stable dimeric and trimeric assemblies at the plasma membrane. Structural modelling uncovered a molecular architecture of the MLO trimer with a central ion-conducting pore, which was further examined by molecular dynamics simulations in a lipid membrane environment. Computational electrophysiology showed preferential inward Ca2+ transport, confirming that MLO proteins function as calcium influx transporters, and identified a conserved set of pore-lining residues that coordinate ion translocation. Functional and structural analyses indicated that the mechanism of calcium permeation is evolutionarily conserved. Our findings provide mechanistic insight into MLO-mediated calcium influx across the plasma membrane and establish multimerisation as a critical determinant of this calcium channels activity.

plant biology↗

bioRxiv2026

The molecular architecture of the Arabidopsis callose synthase complex

ABSTRACTCallose synthase is responsible for the targeted deposition of the {beta}-1,3-glucan polymer, callose which underlines essential plant developmental processes, including cell division, pathogen defense or cell-cell communication. The architecture of the callose synthase complex (CALSC) as well as the molecular mechanisms of callose synthesis remain unknown. Here we report an integrative characterisation of the Arabidopsis CALS complex, with the most enriched subunits, CALS1, CALS2 and CALS3, forming its core. Structurally, CALSC assembles into a trimer, requiring the plant-specific Bag domain to mediate inter-subunit associations. The biological importance of CALSC assembly is highlighted by the simultaneous loss of CALS1 and CALS3, which abolishes plasmodesmal callose deposition and affects symplastic transport. Site-directed mutagenesis and molecular dynamics simulations depict the topology of the CALS1 active site in detail, including the components of the enzymatic reaction. We pinpoint the translocating tunnel through which the nascent glucan is delivered and mechanistically confirm the role of transmembrane helix 8 in regulating glucan export. Our work provides unprecedented insight into the molecular architecture of the CALSC and the distinct changes from maturation to activity at the plasma membrane, while showcasing the mechanisms involved in callose synthesis at the molecular level.

plant biology↗