A Developmental Framework for Complex Plasmodesmata Formation Revealed by Large-Scale Imaging of the Arabidopsis Leaf Epidermis

Abstract

Plasmodesmata (PD) form tubular connections that function as intercellular communication channels. They are essential for transporting nutrients and for coordinating development. During cytokinesis, simple PDs are inserted into the developing cell plate, while during wall extension, more complex (branched) forms of PD are laid down. We show that complex PDs are derived from existing simple PDs in a pattern that is accelerated when leaves undergo the sink-source transition. Complex PDs are inserted initially at the three-way junctions between epidermal cells but develop most rapidly in the anisocytic complexes around stomata. For a quantitative analysis of complex PD formation, we established a high-throughput imaging platform and constructed PDQUANT, a custom algorithm that detected cell boundaries and PD numbers in different wall faces. For anticlinal walls, the number of complex PDs increased with increasing cell size, while for periclinal walls, the number of PDs decreased. Complex PD insertion was accelerated by up to threefold in response to salicylic acid treatment and challenges with mannitol. In a single 30-min run, we could derive data for up to 11k PDs from 3k epidermal cells. This facile approach opens the door to a large-scale analysis of the endogenous and exogenous factors that influence PD formation.

A Journey Through a Leaf: Phenomics Analysis of Leaf Growth inArabidopsis thaliana

“…Obtaining these data is however laborious and/or time-consuming, arguing for more automated setups to obtain high-quality cell images suitable for data extraction. High-throughput imaging using spinning disc microscopy has recently been applied in plant research (Fitzgibbon et al, 2013) to perform large-scale imaging of the leaf epidermis.…”

Salicylic Acid Regulates Plasmodesmata Closure during Innate Immune Responses in Arabidopsis

“…Rather, they undergo various types of structural modifications and degeneration/regeneration processes to meet the specific needs of cells that may set out rapid expansion, different developmental phases or differentiation, or adaption processes in response to the changes in various physiological and environmental conditions (Ehlers and Kollmann, 2001;Roberts and Oparka, 2003;Lucas and Lee, 2004;Burch-Smith et al, 2011;Burch-Smith and Zambryski, 2012). For instance, PD frequency and density change as cells grow and develop (Gunning, 1978;Seagull, 1983;Ehlers and Kollmann, 1996;Burch-Smith and Zambryski, 2010;Ehlers and van Bel, 2010) or during shifts from vegetative to reproductive phases (Ormenese et al, 2000;Ormenese et al, 2002;Ormenese et al, 2006); PDs differentiate from simple to complex forms (Faulkner et al, 2008;Fitzgibbon et al, 2013); PDs are completely disintegrated during guard cell maturation (Wille and Lucas, 1984); PD permeability undergoes temporal regulation by environmental conditions, such as daylength and temperature (Ormenese et al, 2006;Bilska and Sowinski, 2010;Rinne et al, 2011), etc. Permeability, dilation, or structure of PDs can be also altered during infection by microbial pathogens (Heinlein, 2002;BenitezAlfonso et al, 2010;Schoelz et al, 2011;Ueki and Citovsky, 2011).…”

“…It is possible that SA or SA pathways may induce a component that PDLP5 requires for its function; our data indicate that PDLP5 cannot function to close PD in the absence of this SA-dependent component, factor X (Figure 7). SA was shown to facilitate secondary PD formation in Arabidopsis seedlings grown on SA-containing medium for several days (Fitzgibbon et al, 2013). It would be interesting to determine whether there is a close link between structural modification of PD and the SA pathway during immune responses and whether SA together with PDLP5 induces PD closure through such modification.…”

Plasmodesmata dynamics are coordinated by intracellular signaling pathways

“…The cell wall needs to allow new PD to be inserted between neighboring cells in a process distinct from PD formation at the cell plate during division, but how this happens remains unclear. New techniques to rapidly quantify the number of PD connecting cells will greatly benefit attempts to identify conditions or genetic alterations that impact PD formation [54]. The increased formation of secondary PD in leaves with silenced ISE1 or ISE2 expression may present a model system for future analyses [14,16].…”