The N-terminal domain of COMPANION OF CELLULOSE SYNTHASE1 promotes microtubule array formation in Arabidopsis

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.