Functional transfer cells differentiate in cultured cotyledons of Vicia faba L. seeds

Transfer-cell-like wall ingrowth deposition is induced in adaxial epidermal cells of Vicia faba L. cotyledons grown in vitro in the presence of high hexose concentrations. We have further characterised this putative transfer cell induction system by examining initiation of secondary-wall ingrowth deposition and expression of sucrose transport-related genes. Wall ingrowth deposition, as wall thickening and scattered papillate ingrowths, was visualised on the outer periclinal walls of adaxial epidermal cells of cotyledons after 1 day in culture. Over the next 2 days, wall deposition increased significantly to form a distinct band of discrete and coalescing ingrowths. Thereafter, further wall deposition was arrested. Densities of Golgi, endoplasmic reticulum, and mitochondria increased concurrently with wall ingrowth deposition. Transcripts of a H+/sucrose symporter (SUT) and a sucrose-binding protein (SBP) were detected by in situ hybridisation in differentiating transfer cells. Antibodies raised against an H+-ATPase immunolocalised evenly around the perimeter of the adaxial epidermal cells in day 1 cotyledons. Thereafter, labelling became increasingly localised to the developing wall ingrowth regions. In contrast, SEP antibodies immunolocalised exclusively to wall ingrowth regions. However, SEP exhibited a transient pattern of expression, being detected only in 2-day-cultured cotyledons. A proton gradient, sufficient to facilitate sulphorhodamine G accumulation, was established by adaxial epidermal cells after 1 day in culture. [C-14]sucrose uptake by cotyledons became sensitive to an inhibitor of carrier-mediated transport of sucrose. para-chloromercuribenzene sulfonic acid after 2 days. The initial 37% inhibition of sucrose transport by this compound declined to 5% for cotyledons cultured for 3 days. Collectively, these results suggest that differentiation of the key functional characteristics of transfer cells can be induced in vitro, providing an exciting tool for further exploration of transfer cell development.