Cell-to-cell signalling is a major mechanism controlling plant morphogenesis. Transport of signalling molecules through plasmodesmata is one way in which plants promote or restrict intercellular signalling over short distances. Plasmodesmata are membrane-lined pores between cells that regulate the intercellular flow of signalling molecules through changes in their size, creating symplasmic fields of connected cells. Here we examine the role of plasmodesmata and symplasmic communication in the establishment of plant cell totipotency, using somatic embryo induction from Arabidopsis explants as a model system. Cell-to-cell communication was evaluated using fluorescent tracers, supplemented with histological and ultrastructural analysis, and correlated with expression of a WOX2 embryo reporter. We showed that embryogenic cells are isolated symplasmically from non-embryogenic cells regardless of the explant type (immature zygotic embryos or seedlings) and inducer system (2,4-dichlorophenoxyacetic acid or the BABY BOOM (BBM) transcription factor), but that the symplasmic domains in different explants differ with respect to the maximum size of molecule capable of moving through the plasmodesmata. Callose deposition in plasmodesmata preceded WOX2 expression in future sites of somatic embryo development, but later was greatly reduced in WOX2-expressing domains. Callose deposition was also associated with a decrease DR5 auxin response in embryogenic tissue. Treatment of explants with the callose biosynthesis inhibitor 2-deoxy-D-glucose supressed somatic embryo formation in all three systems studied, and also blocked the observed decrease in DR5 expression. Together these data suggest that callose deposition at plasmodesmata is required for symplasmic isolation and establishment of cell totipotency in Arabidopsis.

Background and Aims In Utricularia nelumbifolia, the nuclei of placental nutritive tissue possess unusually shaped projections not known to occur in any other flowering plant. The main aim of the study was to document the morphology and ultrastructure of these unusual nuclei. In addition, the literature was searched to find examples of nuclear tubular projections in other plant groups, and the nuclei of closely related species of Utricularia (i.e. sects Iperua, Orchidioides, Foliosa and Utricularia) were examined. Methods To visualize the complexity of the nuclear structures, transmission electron microscopy (TEM) was used, and 3-D ultrastructural reconstructions were made using the serial block face scanning electron microscopy (SBEM) technique. The nuclei of 11 Utricularia species, i.e. U. nelumbifolia, U. reniformis, U. cornigera, U. nephrophylla (sect. Iperua), U. asplundii, U. alpina, U. quelchii (sect. Orchidioides), U. longifolia (sect. Foliosa), U. intermedia, U. minor and U. gibba (sect. Utricularia) were examined. Key Results Of the 11 Utricularia species examined, the spindle-like tubular projections (approx. 5 μm long) emanating from resident nuclei located in placental nutritive tissues were observed only in U. nelumbifolia. These tubular nuclear extensions contained chromatin distributed along hexagonally shaped tubules. The apices of the projections extended into the cell plasma membrane, and in many cases also made contact at the two opposing cellular poles, and with plasmodesmata via a short cisterna of the cortical endoplasmic reticulum. Images from the SBEM provide some evidence that the nuclear projections are making contact with those of neighbouring cells. Conclusions The term chromatubules (chromatin-filled tubules) for the nuclear projections of U. nelumbifolia placental tissue was proposed here. Due to the apparent association with the plasma membrane and plasmodesmata, it was also speculated that chromatubules are involved in nucleus–cell–cell communication. However, further experimental evidence is required before any functional hypothesis can be entertained.

We studied the organization of F-actin and the microtubular cytoskeleton in male germ-line cysts in the seminal vesicles of the earthworm Dendrobaena veneta using light, fluorescent and electron microscopy along with both chemically fixed tissue and life cell imaging. Additionally, in order to follow the functioning of the cytoskeleton, we incubated the cysts in colchicine, nocodazole, cytochalasin D and latrunculin A. The male germ-line cells of D. veneta are interconnected via stable intercellular bridges (IB), and form syncytial cysts. Each germ cell has only one IB that connects it to the anuclear central cytoplasmic mass, the cytophore. During the studies, we analyzed the cytoskeleton in spermatogonial, spermatocytic and spermatid cysts. F-actin was detected in the cortical cytoplasm and forms distinct rings in the IBs. The arrangement of the microtubules changed dynamically during spermatogenesis. The microtubules are distributed evenly in whole spermatogonial and spermatocytic cysts; however, they primarily accumulate within the IBs in spermatogonia. In early spermatids, microtubules pass through the IBs and are present in whole cysts. During spermatid elongation, the microtubules form a manchette while they are absent in the cytophore and in the IBs. Use of cytoskeletal drugs did not alter the general morphology of the cysts. Detectable effects—the occurrence of nuclei in the late spermatids and manchette fragments in the cytophore—were observed only after incubation in nocodazole. Our results suggest that the microtubules are responsible for cytoplasmic/organelle transfer between the germ cells and the cytophore during spermatogenesis and for the positioning of the spermatid nuclei.