Arabidopsis Myosin XIK Interacts with the Exocyst Complex to Facilitate Vesicle Tethering during Exocytosis

Abstract

Myosin motors are essential players in secretory vesicle trafficking and exocytosis in yeast and mammalian cells; however, similar roles in plants remain a matter for debate, at least for diffusely-growing cells. Here, we demonstrate that Arabidopsis (Arabidopsis thaliana) myosin XIK, via its globular tail domain (GTD), participates in the vesicle tethering step of exocytosis through direct interactions with the exocyst complex. Specifically, myosin XIK GTD bound directly to the SEC5B subunit of exocyst in vitro and functional fluorescently-tagged XIK colocalized with multiple exocyst subunits at plasma membrane (PM)-associated stationary foci. Moreover, genetic and pharmacological inhibition of myosin XI activity reduced the frequency and lifetime of stationary exocyst complexes at the PM. By tracking single exocytosis events of cellulose synthase (CESA) complexes (CSCs) with high spatiotemporal resolution imaging and pair-wise colocalization analysis of myosin XIK, exocyst subunits and CESA6, we demonstrated that XIK associates with secretory vesicles earlier than exocyst and is required for the recruitment of exocyst to the PM tethering site. This study reveals an important functional role for myosin XI in secretion and provides new insights about the dynamic regulation of exocytosis in plants.

Autoregulation and Dual Stepping Mode of MYA2, an Arabidopsis Myosin XI Responsible for Cytoplasmic Streaming

“…The same intra-molecular regulation in MYA2 suggests that MYA2 is not merely responsible for driving continuous cytoplasmic streaming, but also for more complex transport equipped with a recycling system. It has recently been reported that MYA2 is involved in several intracellular transports and functions, such as processing-body movement, pathogen responses, auxin responses, cell death, cell division, regulation of exocytosis, lipid body transport 18, [30][31][32] .…”

Revising the Role of Cortical Cytoskeleton During Secretion: Actin and Myosin XI Function in Vesicle Tethering

“…For example, putative CSC delivery vesicles arrive in the cortical cytoplasm near the PM, display erratic local movement for several seconds, become spatially restricted in their movement and maintain a static position for 60-90s, and then undergo steady and linear movement as catalytically-active CESA complexes (see refs. [17,23,25] and Figure 1A). The pause phase is hypothesized to encompass tethering, docking, and fusion of CSC vesicles, possibly followed by the activation of CESAs before they synthesize cellulose and show linear motility.…”

“…Recent advances in high spatiotemporal resolution live-cell imaging and utilization of functional, fluorescent protein-tagged CESAs have allowed CSC trafficking to emerge as a powerful model for studying exocytosis in plant cells [17,[23][24][25]. In particular, single-particle tracking has illuminated specific steps in the exocytosis process based on the size, unique spatial localization and motility patterns of CSCs on different endomembrane compartments [17,23,24].…”

“…Through genetic studies as well as fluorescent tagging and colocalization analyses, several proteins that transiently associate with CSC particles during the pause phase and may play a role in the efficient delivery of CSCs to the PM have been identified (Figure 1B). Some are exocytosis-related machinery proteins, including the exocyst tethering complex which coappears with CSCs at the beginning of the pause phase for ~12 s, suggesting that the vesicle tethering stage immediately follows the arrival of CSC compartments at the PM [25,26]. The plant-specific protein PATROL1 (PTL1), a homolog of Munc13 proteins in animals with a role in SNARE complex assembly, colocalizes with CSCs for 1-2 s at the pause phase and often appears 1-2 s after the appearance of exocyst subunits [26].…”

“…KEULE, a plant homolog of Sec1/Munc18 proteins implicated in regulating SNARE function, also arrives at the beginning of the pause phase with an average lifetime of 3.7 s and is likely to play a role in docking and/or fusion of CSC vesicles [27]. The actin-based motor, Myosin XIK, coappears with cortical CSC compartments and remains present at the beginning of the pause phase for 3-9 s before disappearing [23,25]. Finally, a new plant-specific protein, TRANVIA (TVA), arrives at the PM with CESA delivery compartments and remains associated with CSCs during the pause phase for an average of 6.3 s, and likely plays a role in CSC delivery [24].…”