Chemical Effectors of Plant Endocytosis and Endomembrane Trafficking

Abstract

Plant endocytosis and endomembrane trafficking relies on the coordination of a highly organized and dynamic network of intracellular organelles. Membrane trafficking and associated signal transduction pathways provide critical cellular regulation of plant development and response to environmental stimuli. However, the efficiency of studies on this complex network has been hampered due to the rapid and dynamic nature of endomembrane trafficking as well as gene redundancy and embryonic lethality in mutagenesis-based strategies. Chemical genomics emerged in recent years as a complementary approach to illuminate biological functions through the integration of organic chemistry, biology, and bioinformatics to overcome gene redundancy. The approach presents significant advantages in dosage dependence and reversibility, which offers the ideal ability to study dynamic endomembrane trafficking processes in real time. In this chapter, several successful examples of chemical screening focused on the endomembrane system is presented to illuminate the efficiency and power of chemical genomics in dissecting endomembrane trafficking and its regulation of plant development and environmental responses. Perspectives are also presented to suggest directions for future development of this field.

GABA signalling modulates plant growth by directly regulating the activity of plant-specific anion transporters

“…2c ). We examined the possibility that endocytosis of TaALMT1 contributed to the reduction in malate efflux following GABA treatment, but found no effect of the endocytosis inhibitor Brefeldin A 12 on Al 3+ -activated malate efflux or the inhibition by muscimol ( Supplementary Fig. 3 ).…”

CesA6 and PGIP2 Endocytosis Involves Different Subpopulations of TGN-Related Endosomes

“…The effect of Wm was evident and specific on transport of PGIP2 to the vacuole also at a concentration (3 µM) lower than that which affects other degrading traffic events (Di Sansebastiano et al, 2007;Ariani et al, 2018). On the contrary ES5, which specifically affects the PM recycling in plant and animal cells (Drakakaki et al, 2011;Li et al, 2012), did not influence PGIP2 vacuolar sorting. A completely different response was obtained with the same chemicals on secGFP-CesA6, which was insensitive to Wm and Sortin 2 but was clearly affected by ES5, with the formation of the typical Endosidin bodies (Drakakaki et al, 2011), thus blocking the recycling of the chimera in endosomes different from those labeled by PGIP2-GFP, and likely represented by MASCs (see Figure 2).…”

“…The possibility that TGN is the predominant compartment shared by PGIP2 and CesA6 during their endocytic route was analyzed by using ConA (Figure 2) and BFA (Figures 6, 7). The response to ConA, known to block both the trafficking from TGN to the plasma membrane and the sorting of the endocytic marker FM4-64 from the TGN/EE to the vacuole (Dettmer et al, 2006;Li et al, 2012), was not determinant, since both PGIP2-GFP and secGFP-CesA6 were retained in endosomal enlarged compartments. The effect of ConA may occur on a regulatory target common to both markers' sorting or, as suggested by the induction of a similar fluorescent patter (Figures 2C,H), on early step of sorting when the two markers share the same compartment.…”

“…Again, the change of morphology was so extreme that statistical evaluation was not necessary. On the contrary, no effect was observed on PGIP2-GFP after treatment with Endosidin 5 (ES5; 50 µM for 1 h) (Figure 2E) that should block proteins recycling from plasma membrane (Li et al, 2012). The PGIP2-GFP sensitivity to Wm and Sortin 2 demonstrated its transition through PVC/MVB/LE.…”

Remove, Recycle, Degrade: Regulating Plasma Membrane Protein Accumulation

“…Among them, the use of chemicals that interfere with specific pathways in specific tissues and at specific developmental stages has facilitated the discovery of cellular phenotypes during short controlled incubations. A growing toolbox of chemicals is available for the community, allowing the study of several specific endocytic pathways (for review, see Li et al, 2012;Hicks and Raikhel, 2014;Rodriguez-Furlan et al, 2017). With such chemicals, it is imperative to consider possible off-target effects, such as those discussed for BFA.…”