Different Endomembrane Trafficking Pathways Establish Apical and Basal Polarities

Abstract

The endomembrane system is an interconnected network required to establish signal transduction, cell polarity, and cell shape in response to developmental or environmental stimuli. In the model plant , there are numerous markers to visualize polarly localized plasma membrane proteins utilizing endomembrane trafficking. Previous studies have shown that the large ARF-GEF GNOM plays a key role in the establishment of basal (rootward) polarity, whereas the apically (shootward) polarized membrane proteins undergo sorting via different routes. However, the mechanism that maintains apical polarity is largely unknown. Here, we used a chemical genomic approach and identified the compound endosidin 16 (ES16), which perturbed apically localized plasma membrane proteins without affecting basal polarity. We demonstrated that ES16 is an inhibitor for recycling of apical, lateral, and nonpolar plasma membrane proteins as well as biosynthetic secretion, leaving the basal proteins as the only exceptions not subject to ES16 inhibition. Further evidence from pharmaceutical and genetic data revealed that ES16 effects are mediated through the regulation of small GTPase RabA proteins and that RabA GTPases work in concert with the BIG clade ARF-GEF to modulate the nonbasal trafficking. Our results reveal that ES16 defines a distinct pathway for endomembrane sorting routes and is essential for the establishment of cell polarity.

The Plant Trans-Golgi Network: Not Just a Matter of Distinction

“…Organelle proteomics is allowing the identification of vesicle protein cargoes, and extending glycomic analysis to isolated vesicles will help us to better characterize the secretory routes of cell wall polysaccharides (Obel et al, 2009;Pattathil et al, 2010;Drakakaki et al, 2012;Parsons et al, 2013;Heard et al, 2015;Kra cun et al, 2017;Wood et al, 2017). Chemical genomics is enabling the characterization of vesicle-trafficking pathways, recently evidenced by the use of the small molecules ES7 and ES16 to discern the contributions of two RAB GTPases to cell plate formation and to demonstrate the specificity of a trafficking pathway involved in cell polarity, respectively (Davis et al, 2016b;Li et al, 2017). Spatiotemporal image correlation spectroscopy has proven useful to characterize the dynamics of vesicle trafficking to the cell plate (Hebert et al, 2005;van Oostende-Triplet et al, 2017).…”

Drug Affinity Responsive Target Stability (DARTS) to Resolve Protein–Small Molecule Interaction in Arabidopsis

“…A classical chemical genomics approach involves three major steps: (i) screening for bioactive small molecules, (ii) characterization of the small molecules, and (iii) identification of targets (Rodriguez-Furlan et al, 2014). Through simple and rapid high-throughput screenings performed in diverse plant models, it has been possible to create several collections of plant-bioactive small molecules (Drakakaki et al, 2011;Halder et al, 2015;Knoth et al, 2009;Noutoshi et al, 2012;Rodriguez-Furlan et al, 2016;Zhao et al, 2007). Additional phenotype-based assays can be used to effectively evaluate the effects of small molecules and correlate these effects with biological target pathways.…”

Plant Chemical Genetics: From Phenotype-Based Screens to Synthetic Biology

“…Recently, ES2 has been found to inhibit exocytosis in plants and human cells and to target the EXO70 subunit of the exocyst complex (Zhang et al, 2016). ES8 affects secretory pathways, exclusively toward the basal plasma membrane of the cell, thereby affecting PIN-FORMED1 trafficking and auxin distribution (Doyle et al, 2015), whereas ES16 specifically perturbs apically localized proteins through regulation of the small GTPase RabA proteins (Li et al, 2017). ES9, which was identified as an inhibitor of endocytosis in different systems (Dejonghe et al, 2016), affected endomembrane dynamics, such as Golgi compartment movements, and depleted cellular ATP.…”