Abstract
Microtubule-associated proteins (MAPs) play important roles in cellulose biosynthesis in plants. However, the molecular mechanisms mediating their interactions with cortical microtubule (MT) arrays remain to be elucidated. Here, we investigated the companion of cellulose synthase 1 (CC1), an Arabidopsis (Arabidopsis thaliana) MAP that stabilizes cellulose biosynthesis during salt stress by maintaining the integrity of the cortical MT array. The N-terminal domain of CC1 (CC1NTD) is sufficient to restore cellulose biosynthesis in Arabidopsis cc1cc2 knockout mutants. We used a combination of small-angle X-ray and neutron scattering (SAXS and SANS), single-molecule Förster resonance energy transfer, and computational modeling to determine the structural characteristics of CC1NTD and its interactions with MTs. SANS measurements combined with deuterium labeling of CC1NTD allowed the structural features of CC1NTD and MTs to be deconvoluted and analyzed separately. CC1NTD bound to the MT surface and promoted interactions between neighboring MTs to form tightly associated arrays. In addition, CC1NTD appeared to be in an extended conformation during MT interactions, which could be important for forming cross-bridges between MTs during salt stress. Overall, this study provides structural insights into the mechanisms associated with a disordered MT-binding region in an MAP and provides an explanation for CC1’s efficient organization of MTs, highlighting its importance in cellulose biosynthesis under stress conditions.
| Original language | English |
|---|---|
| Article number | kiaf392 |
| Journal | Plant Physiology |
| Volume | 199 |
| Issue number | 1 |
| DOIs | |
| State | Published - Sep 2025 |
Funding
V.G. and H.O. acknowledge funding by the Center for Lignocellulose Structure and Formation supported by the U.S. Department of Energy (DOE), Office of Science, Basic Energy Sciences under Award # DE-SC0001090. A.C.H. and J.C.S. acknowl - edge support of the project ERKPA14 funded by the DOE Office of Biological & Environmental Research (OBER) for computational modeling. W.C.L. supported SANS experiments on Bio-SANS, which is part of the Center for Structural Molecular Biology, funded by DOE OBER project ERKP291. S.C. supported SAXS experiments on the LiX beamline, which is part of the Center for BioMolecular Structure (CBMS), funded by the National Institutes of Health, National Institute of General Medical Sciences (NIGMS) through a P30 Grant (P30GM133893), and by the DOE Office of Biological and Environmental Research (KP1605010). LiX also received additional support from NIH Grant S10 OD012331. RL acknowledges support from the National Institutes of Health R35GM142946 and the start-up funds from the College of Arts and Sciences, University of Tennessee, Knoxville, for fluorescence microscopy work. Conflict of interest statement. None declared. This research used resources at the High Flux Isotope Reactor and Spallation Neutron Source, a DOE Office of Science User Facility operated by the Oak Ridge National Laboratory. This research used resources of the Compute and Data Environment for Science (CADES) at the Oak Ridge National Laboratory, which is supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC05-00OR22725. The authors thank Dr Volker Urban and Dr Sai Venkatesh Pingali for discussions about SANS data interpretation. Additionally, they thank Dr Kevin Weiss and Ms Qiu Zhang for advice and support in producing protiated and deuterated CC1NTD used in this study. V.G. and H.O. acknowledge funding by the Center for Lignocellulose Structure and Formation supported by the U.S. Department of Energy (DOE), Office of Science, Basic Energy Sciences under Award # DE-SC0001090. A.C.H. and J.C.S. acknowledge support of the project ERKPA14 funded by the DOE Office of Biological & Environmental Research (OBER) for computational modeling. W.C.L. supported SANS experiments on Bio-SANS, which is part of the Center for Structural Molecular Biology, funded by DOE OBER project ERKP291. S.C. supported SAXS experiments on the LiX beamline, which is part of the Center for BioMolecular Structure (CBMS), funded by the National Institutes of Health, National Institute of General Medical Sciences (NIGMS) through a P30 Grant (P30GM133893), and by the DOE Office of Biological and Environmental Research (KP1605010). LiX also received additional support from NIH Grant S10 OD012331. RL acknowledges support from the National Institutes of Health R35GM142946 and the start-up funds from the College of Arts and Sciences, University of Tennessee, Knoxville, for fluorescence microscopy work. 1UT/ORNL Graduate School of Genome and Science Technology, University of Tennessee, Knoxville, TN 37996, USA 2UT/ORNL Center for Molecular Biophysics, Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA 3Department of Biochemistry & Cellular and Molecular Biology, University of Tennessee, Knoxville, TN 37996, USA 4Neutron Scattering Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA 5LiX beamline, National Synchtron Light Source II, Brookhaven National Laboratory, Upton, NY 11973, USA 6Advanced Microscopy and Imaging Center, University of Tennessee, Knoxville, TN 37996, USA *Author for correspondence: [email protected] This manuscript has been authored by UT-Battelle, LLC under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy. The United States Government retains and the publisher, by accepting the article for publication, acknowledges that the United States Government retains a non-exclusive, paid-up, irrevocable, world-wide license to publish or reproduce the published form of this manuscript, or allow others to do so, for United States Government purposes. The Department of Energy will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan (http://energy.gov/downloads/ doe-public-access-plan) The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (https://academic.oup.com/plphys/pages/General-Instructions) is Hugh O’Neill ([email protected]).
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