Fig. 6

Simultaneous knockdown of YIP4A/B perturbed NtPsCRP2 secretion in seedlings. a An Arabidopsis seedling showing regions analyzed in this study. b, c NtPsCRP2-GFP localized less frequently in young root hairs but more frequently in mature root hairs, both in the wild type and in the yip4a-1;yip4b double mutant. d NtPsCRP2 secretion in 4-day-old etiolated wild-type seedlings showing localization in epidermal cells of the hypocotyl. The secretion was restricted at the hypocotyl–root junction. Propidium iodide (red) marks the beginning of the root. e Close-up of NtPsCRP2-GFP in elongated epidermal cells of the hypocotyl showing accumulation in spherical, aggregated endosomal-like vesicles. f, g Conversely, in yip4a-1;yip4b seedlings, NtPsCRP2 secretion was severely distorted, showing diffuse localization and non-uniform protein aggregates (arrowheads). h The spherical marked endosomal aggregates observed in the wild type were substituted with rod-like labeled organelles in yip4a-1;yip4b hypocotyl. i NtPsCRP2 localization at the root tip of wild-type seedlings marking secretory organelles. EPI epidermis, CX cortex. j Magnified root epidermal cells from wild type showing NtPsCRP2-marked organelles constrained to the vicinity of the plasma membrane (white rectangle) and also localized in ER-body-like organelles (inset). k, l In yip4a-1;yip4b, NtPsCRP2-GFP localization in secretory organelles at the root apical meristem appeared largely disorganized, showing severe organelle aggregations resembling BFA-like compartments (arrowheads). The ER-body-like localization (arrow) was maintained, suggesting no effect on ER-body biogenesis in yip4a-1;yip4b mutants. m–o Perturbed secretion of NtPsCRP2-GFP was also recapitulated following BFA or wortmannin treatment. Arrowheads show NtPsCRP2-GFP BFA compartments, arrows point to diffusely aggregated NtPsCRP2-GFP aberrant secretion. RC lateral root cap PI propidium iodide stain, WT wild type. Scale bars = 10 μM