Cell Wall Polysaccharides are Mislocalized to the Vacuole in echidna Mutants

Abstract

During cell wall biosynthesis, the Golgi apparatus is the platform for cell wall matrix biosynthesis and the site of packaging, of both matrix polysaccharides and proteins, into secretory vesicles with the correct targeting information. The objective of this study was to dissect the post-Golgi trafficking of cell wall polysaccharides using echidna as a vesicle traffic mutant of Arabidopsis thaliana and the pectin-secreting cells of the seed coat as a model system. ECHIDNA encodes a trans-Golgi network (TGN)-localized protein, which was previously shown to be required for proper structure and function of the secretory pathway. In echidna mutants, some cell wall matrix polysaccharides accumulate inside cells, rather than being secreted to the apoplast. In this study, live cell imaging of fluorescent protein markers as well as transmission electron microscopy (TEM)/immunoTEM of cryofixed seed coat cells were used to examine the consequences of TGN disorganization in echidna mutants under conditions of high polysaccharide production and secretion. While in wild-type seed coat cells, pectin is secreted to the apical surface, in echidna, polysaccharides accumulate in post-Golgi vesicles, the central lytic vacuole and endoplasmic reticulum-derived bodies. In contrast, proteins were partially mistargeted to internal multilamellar membranes in echidna. These results suggest that while secretion of both cell wall polysaccharides and proteins at the TGN requires ECHIDNA, different vesicle trafficking components may mediate downstream events in their secretion from the TGN.

Golgi- and Trans-Golgi Network-Mediated Vesicle Trafficking Is Required for Wax Secretion from Epidermal Cells

“…Previous studies of gnl1 and ech mutants have revealed altered ER morphology (Nakano et al, 2009;McFarlane et al, 2013), similar to the localization pattern of the ER membrane-localized CER2-GFP in gnl1-1 and ech (Supplemental Fig. S7).…”

“…Both ech and trs120-4 are defective in TGN protein complexes. ECH is a part of the ECH/Ypt/Rab-interacting protein complex, which is required specifically for the trafficking of secretory vesicles from the TGN to the plasma membrane (Gendre et al, 2011;Boutté et al, 2013;McFarlane et al, 2013). ech is a null allele (Gendre et al, 2011), while null alleles of trs120 (trs120-2 and trs120-3) are seedling lethal (Thellmann et al, 2010;Qi et al, 2011); therefore, experiments on mature plants required use of the weaker allele, trs120-4.…”

Flavonoid transport mechanisms: how to go, and with whom

“…GNL1 is required for coat protein complex I vesicle formation in retrograde trafficking from the Golgi to the ER [46]. ECHIDNA (ECH) is a part of the ECH/Rab GTPase-interacting protein complex required for vesicle trafficking from the TGN to the PM [68][69][70]. Transport particle protein subunit120-4 (TRS120-4) is a part of the large GTPase complex required for secretion to the cell wall and cell plate [71,72].…”

Ethylene Regulates Differential Growth via BIG ARF-GEF-Dependent Post-Golgi Secretory Trafficking in Arabidopsis

“…VHA-a1 has previously been connected to the secretion of BRI1 (Dettmer et al, 2006). However, despite the colocalization between ECH and VHA-a1 at the TGN as well as the mislocalization of VHA-a1 in the ech mutant, ECH-dependent AUX1 secretory trafficking in the apical hook and ECH-dependent pectin secretion in seed coat cells occur independently of VHA-a1 (Boutté et al, 2013;Gendre et al, 2013;McFarlane et al, 2013). Like ECH, BIGs (BIG3/4) localize to the TGN and colocalize with VHA-a1 (Richter et al, 2014) (Figures 6C and 6D) and ECH ( Figures 6A and 6D).…”