Directional cell expansion requires NIMA-related kinase 6 (NEK6)-mediated cortical microtubule destabilization

Abstract

Plant cortical microtubules align perpendicular to the growth axis to determine the direction of cell growth. However, it remains unclear how plant cells form well-organized cortical microtubule arrays in the absence of a centrosome. In this study, we investigated the functions of Arabidopsis NIMA-related kinase 6 (NEK6), which regulates microtubule organization during anisotropic cell expansion. Quantitative analysis of hypocotyl cell growth in the nek6-1 mutant demonstrated that NEK6 suppresses ectopic outgrowth and promotes cell elongation in different regions of the hypocotyl. Loss of NEK6 function led to excessive microtubule waving and distortion, implying that NEK6 suppresses the aberrant cortical microtubules. Live cell imaging showed that NEK6 localizes to the microtubule lattice and to the shrinking plus and minus ends of microtubules. In agreement with this observation, the induced overexpression of NEK6 reduced and disorganized cortical microtubules and suppressed cell elongation. Furthermore, we identified five phosphorylation sites in β-tubulin that serve as substrates for NEK6 in vitro. Alanine substitution of the phosphorylation site Thr166 promoted incorporation of mutant β-tubulin into microtubules. Taken together, these results suggest that NEK6 promotes directional cell growth through phosphorylation of β-tubulin and the resulting destabilization of cortical microtubules.

An evolutionarily conserved NIMA-related kinase directs rhizoid tip growth in the basal land plant Marchantia polymorpha

“…S11). The localization pattern of MpNEK1-Citrine is similar to that of AtNEK6-GFP, which associates with and moves along cortical microtubules (Motose et al, 2011;Takatani et al, 2017).…”

“…Consistent with this, NIMA kinase of the filamentous fungus Aspergillus nidulans, a representative member of the NEK family, has been shown to regulate directional growth of hyphae during interphase by microtubule organization (Govindaraghavan et al, 2014). The Arabidopsis genome has seven NEK members (AtNEK1-7), among which AtNEK6 plays a pivotal role in directional cell expansion (Motose et al, 2012;Takatani et al, 2015bTakatani et al, , 2017. The epidermal cells of hypocotyls and petioles in Arabidopsis nek6 mutants exhibit ectopic outgrowths (Motose et al, 2008;Sakai et al, 2008).…”

“…Because AtNEK6 has been shown to regulate microtubule organization (Motose et al, 2011;Takatani et al, 2017), we hypothesized that MpNEK1 also regulates microtubule organization. To test this possibility, we introduced CaMV35Spro: TagRFP-MpTUB2 into the wild type and Mpnek1 knockouts to visualize microtubules in rhizoids (Fig.…”

“…The epidermal cells of hypocotyls and petioles in Arabidopsis nek6 mutants exhibit ectopic outgrowths (Motose et al, 2008;Sakai et al, 2008). AtNEK6 interacts with other NEK family members and phosphorylates β-tubulin, resulting in depolymerization of cortical microtubules, during directional cell expansion (Motose et al, 2011;Takatani et al, 2017).…”

Update: Plant Cortical Microtubule Arrays

“…The functional relevance of different a/b-dimer combinations is still unknown but may soon be open for investigation with tools developed to purify specific tubulin isoforms and dimers with different posttranslational modifications from plant cells (Hotta et al, 2016). Tubulin modifications, such as C-terminal detyrosination (Duckett and Lloyd, 1994), phosphorylation, and acetylation, have long been known in plant cells and may confer functional effects in what has been dubbed a tubulin code (Cai, 2010;Xiong et al, 2013;Gadadhar et al, 2017;Takatani et al, 2017).…”