Biology subjects

Caregnato, A.

Publications and source records attributed to Caregnato, A..

3 recordsLinked to original sources

bioRxiv2026

Structure of the Arabidopsis receptor kinase SRF6 ectodomain determined from crystals obtained using the LRR crystallisation screen

Plant-specific membrane receptor kinases with structurally diverse extracellular domains regulate key processes in plant growth, development, immunity and symbiosis. Structural studies of these glycoproteins are often hampered by the limited quantities in which they can be obtained. Here, we describe the LRR crystallization screen, which has enabled the successful crystallization and structure determination of multiple receptor kinase ectodomains, including ligand-and co-receptor-bound complexes. As an example, we report the 1.5 [A] resolution crystal structure of the leucine-rich repeat (LRR) domain of STRUBBELIG-RECEPTOR FAMILY 6 (SRF6) from Arabidopsis thaliana. The SRF6 ectodomain contains seven LRRs and a disulfide-bond-stabilised N-terminal capping domain but lacks the canonical C-terminal cap and the N-glycosylation pattern typically observed in other family members. Previously reported protein-protein interactions between the SRF6 and SRF7 ectodomains and the receptor kinases BRI1, BRL1, BRL3, SERK3 and BIR1-3 could not be confirmed by quantitative isothermal titration calorimetry and grating-coupled interferometry assays, suggesting that these structurally conserved LRR receptor kinases may have signalling functions outside the brassinosteroid pathway. SynopsisA crystallisation screen that has enabled the structural analysis of various extracellular domains of plant membrane receptor kinases is described together.

plant biology↗

bioRxiv2025

A mechanistic framework for the recognition of chemically diverse brassinosteroids by BRI1-family receptor kinases

Brassinosteroids (BRs) are chemically diverse plant steroid hormones produced via a branched biosynthetic pathway. The potent BR brassinolide is sensed by the membrane receptor kinase BRIl and a SERK co-receptor, but the physiological functions of other abundant BRs remain to be characterized. Here, we present quantitative binding kinetics for four Arabidopsis thaliana BR receptors and fifteen BRs, which define the key chemical features required for high affinity receptor binding, ligand positioning, and co-receptor recognition. BRIl, BRLl, and BRL3 share overlapping ligand preferences, whereas BRL2 preferentially binds C28 BRs with moderate affinity. Structural analyses of BR-bound BRIl and BRL3 ectodomains, combined with extensive in vitro and in vivo mutagenesis studies reveal a high structural plasticity of the hormone binding pocket. Functional assays using structure-based BR agonists and antagonists uncover that BR receptor - co-receptor signaling complexes can recognize chemically diverse BRs, introducing an additional, intriguing layer of BR signaling regulation.

plant biology↗

bioRxiv2021

Disease-linked mutations trigger exposure of a protein quality control degron in the DHFR protein

Degrons are short stretches of amino acids or structural motifs that are embedded in proteins. They mediate recognition by E3 ubiquitin-protein ligases and thus confer protein degradation via the ubiquitin-proteasome system. Well-described degrons include the N-degrons, destruction boxes, and the PIP degrons, which mediate the controlled degradation of various proteins including signaling components and cell cycle regulators. In comparison, the so-called protein quality control (PQC) degrons that mediate the degradation of structurally destabilized or misfolded proteins are not well described. Here, we show that disease-linked DHFR missense variants are structurally destabilized and chaperone-dependent proteasome targets. We systematically mapped regions within DHFR to assess those that act as cytosolic PQC degrons in yeast cells. Two regions, DHFR-Deg13-36 (here Deg1) and DHFR-Deg61-84 (here Deg2), act as degrons and conferred degradation to unrelated fusion partners. The proteasomal turnover of Deg2 was dependent on the molecular chaperone Hsp70. Structural analyses by NMR and hydrogen/deuterium exchange revealed that Deg2 is buried in wild-type DHFR, but becomes transiently exposed in the disease-linked missense variants.

biochemistry↗