The hydroxyl group in the 3-position of the phenylpropanoid compounds is introduced at the level of coumarate shikimate/quinate esters, whose synthesis implicates an acyltransferase activity. Specific antibodies raised against the recombinant tobacco (Nicotiana tabacum) acyltransferase revealed the accumulation of the enzyme in stem vascular tissues of tobacco, in accordance with a putative role in lignification. For functional analysis, the acyltransferase gene was silenced in Arabidopsis thaliana and N. benthamiana by RNA-mediated posttranscriptional gene silencing. In Arabidopsis, gene silencing resulted in a dwarf phenotype and changes in lignin composition as indicated by histochemical staining. An in-depth study of silenced N. benthamiana plants by immunological, histochemical, and chemical methods revealed the impact of acyltransferase silencing on soluble phenylpropanoids and lignin content and composition. In particular, a decrease in syringyl units and an increase in p-hydroxyphenyl units were recorded. Enzyme immunolocalization by confocal microscopy showed a correlation between enzyme accumulation levels and lignin composition in vascular cells. These results demonstrate the function of the acyltransferase in phenylpropanoid biosynthesis.

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Plasmodesmata provide the cytoplasmic conduits for cell-to-cell communication throughout plant tissues and participate in a diverse set of non–cell-autonomous functions. Despite their central role in growth and development and defence, resolving their modus operandi remains a major challenge in plant biology. Features of protein sequences and/or structure that determine protein targeting to plasmodesmata were previously unknown. We identify here a novel family of plasmodesmata-located proteins (called PDLP1) whose members have the features of type I membrane receptor-like proteins. We focus our studies on the first identified type member (namely At5g43980, or PDLP1a) and show that, following its altered expression, it is effective in modulating cell-to-cell trafficking. PDLP1a is targeted to plasmodesmata via the secretory pathway in a Brefeldin A–sensitive and COPII-dependent manner, and resides at plasmodesmata with its C-terminus in the cytoplasmic domain and its N-terminus in the apoplast. Using a deletion analysis, we show that the single transmembrane domain (TMD) of PDLP1a contains all the information necessary for intracellular targeting of this type I membrane protein to plasmodesmata, such that the TMD can be used to target heterologous proteins to this location. These studies identify a new family of plasmodesmal proteins that affect cell-to-cell communication. They exhibit a mode of intracellular trafficking and targeting novel for plant biology and provide technological opportunities for targeting different proteins to plasmodesmata to aid in plasmodesmal characterisation. In plants, cylindrical, microscopic channels called plasmodesmata provide intracellular connections between cells for communication and material transport, and are important for many aspects of plant growth and defence. We identify a novel family of plasmodesmata-located proteins (called PDLP1) with features of type I membrane receptor-like proteins. In line with the potential for this protein to regulate molecular movement from cell to cell, we show that altered expression of the protein changes the efficiency of protein diffusion from plasmodesmata. We have also analysed the manner in which PDLP1 is transported to plasmodesmata. We show that the single transmembrane domain (TMD) of the protein contains all the information necessary for targeting to plasmodesmata and that proper targeting depends upon specific interactions with other factors within the membrane. Notably, a single amino acid close to the C-terminus of the TMD is critical for determining the intracellular destination. Further, by fusing the TMD to yellow fluorescent protein, we establish that the TMD can be used to target heterologous proteins to plasmodesmata.

Brefeldin A (BFA) causes a block in the secretory system of eukaryotic cells by inhibiting vesicle formation at the Golgi apparatus. Although this toxin has been used in many studies, its effects on plant cells are still shrouded in controversy. We have reinvestigated the early responses of plant cells to BFA with novel tools, namely, tobacco Bright Yellow 2 (BY-2) suspension-cultured cells expressing an in vivo green fluorescent protein–Golgi marker, electron microscopy of high-pressure frozen/freeze-substituted cells, and antisera against Atγ-COP, a component of COPI coats, and AtArf1, the GTPase necessary for COPI coat assembly. The first effect of 10 μg/mL BFA on BY-2 cells was to induce in <5 min the complete loss of vesicle-forming Atγ-COP from Golgi cisternae. During the subsequent 15 to 20 min, this block in Golgi-based vesicle formation led to a series of sequential changes in Golgi architecture, the loss of distinct Golgi stacks, and the formation of an endoplasmic reticulum (ER)–Golgi hybrid compartment with stacked domains. These secondary effects appear to depend in part on stabilizing intercisternal filaments and include the continued maturation of cis- and medial cisternae into trans-Golgi cisternae, as predicted by the cisternal progression model, the shedding of trans-Golgi network cisternae, the fusion of individual Golgi cisternae with the ER, and the formation of large ER-Golgi hybrid stacks. Prolonged exposure of the BY-2 cells to BFA led to the transformation of the ER-Golgi hybrid compartment into a sponge-like structure that does not resemble normal ER. Thus, although the initial effects of BFA on plant cells are the same as those described for mammalian cells, the secondary and tertiary effects have drastically different morphological manifestations. These results indicate that, despite a number of similarities in the trafficking machinery with other eukaryotes, there are fundamental differences in the functional architecture and properties of the plant Golgi apparatus that are the cause for the unique responses of the plant secretory pathway to BFA.

As channels that provide cell-to-cell connectivity, plasmodesmata are central to the local and systemic spread of viruses in plants. This review discusses the current state of knowledge of the structure and function of these channels and the ways in which viruses bring about functional changes that allow macromolecular trafficking to occur. Despite the passing of two decades since the first identification of a viral movement protein that mediates these changes, our understanding of the relevant molecular mechanisms remains in its infancy. However, viral movement proteins provide valuable tools for the modification of plasmodesmata and will continue to assist in the dissection of plasmodesmal properties in relation to their core roles in cell-to-cell communication.