Recent advances in our understanding of the molecular control of secondary cell wall (SCW) formation have shed light on molecular mechanisms that underpin domestication traits related to wood formation. One such trait is the cellulose microfibril angle (MFA), an important wood quality determinant that varies along tree developmental phases and in response to gravitational stimulus. The cytoskeleton, mainly composed of microtubules and actin filaments, collectively contribute to plant growth and development by participating in several cellular processes, including cellulose deposition. Studies in Arabidopsis have significantly aided our understanding of the roles of microtubules in xylem cell development during which correct SCW deposition and patterning are essential to provide structural support and allow for water transport. In contrast, studies relating to SCW formation in xylary elements performed in woody trees remain elusive. In combination, the data reviewed here suggest that the cytoskeleton plays important roles in determining the exact sites of cellulose deposition, overall SCW patterning and more specifically, the alignment and orientation of cellulose microfibrils. By relating the reviewed evidence to the process of wood formation, we present a model of microtubule participation in determining MFA in woody trees forming reaction wood (RW).

Key message: Cortical microtubule arrays are the primary mechanism for guiding the re-orientation of cellulose microfibrils and determining MFA in secondary cell wall of wood fibre and tracheid cells in reaction wood. Microtubules are directly and indirectly involved in guiding cellulose synthase complexes (CSCs) through the plasma membrane. The angle of cellulose deposition is a critical response to environmental signals and/or stress conditions, and particularly crucial during reaction wood formation, a process in which woody plants deposit additional cell wall material to counteract gravitational forces. Tubulin genes are upregulated in response to gravitational stimulus during reaction wood formation, which can result in changes to microtubule assembly. In this study, microtubules were visualised in three woody tree species (two angiosperms: Eucalyptus globulus Labill., Populus alba L., and one gymnosperm: Pinus radiata D. Don.) using immunofluorescence to quantitatively evaluate microtubule organisation during reaction wood formation. Our results suggest that reorientation of the cortical microtubule array affects secondary cell wall deposition, even across different types of reaction wood, by ensuring context-appropriate orientation of cellulose microfibrils and determining MFA in wood cells. Pharmacological studies conducted on in vitro cultured stem segments or in vivo during reaction wood formation corroborated these important roles for microtubules during wood development. This study starts to unveil the role of tubulins during wood formation by exploring cortical microtubule array organisation in trees subjected to gravitational stimulus and it sheds light on cellular and molecular mechanisms behind cellulose deposition in tree species.

Cortical microtubules comprise heterodimeric units of α- and β-tubulin which have been shown to guide the deposition of cellulose microfibrils in plant cell walls where their arrangement is important in determining cell morphology and cell wall properties. Tubulin genes are highly expressed in woody tissues and a functional study has demonstrated a role for a β-tubulin gene family member in affecting the orientation of cellulose microfibrils in wood fibre cells, an important trait in determining the mechanical properties of wood fibres. To further understand the role of tubulins in plant cell trait determination, this study identified and investigated the expression of the α- and β-tubulin gene families in Eucalyptus and then, using transgenesis techniques, investigated the role of specific eucalypt tubulin isoforms in determining secondary cell wall traits of wood fibres in plant stems. This study found that the α- and β-tubulin gene families in Eucalyptus are relatively small compared to other species and show higher expression in woody stem tissue when compared to leaf. Functional studies revealed that cambial cells transformed with α- and β-tubulin overexpression and knockdown vectors, either on their own or in combination, lead to changes in the angle of microfibrils in the secondary cell wall of wood fibre cells with Class I- and Class I-like gene family members explicitly involved. This study demonstrates the importance of tubulins in determining the mechanical properties of wood fibres through a mechanism involving specific tubulin isoform expression during wood fibre formation.