BackgroundPlant microRNAs (miRNAs) are a class of small, non-coding RNAs that play an important role in development and environmental responses. Hundreds of plant miRNAs have been identified to date, mainly from the model species for which there are available genome sequences. The current challenge is to characterise miRNAs from plant species with agricultural and horticultural importance, to aid our understanding of important regulatory mechanisms in crop species and enable improvement of crops and rootstocks.ResultsBased on the knowledge that many miRNAs occur in large gene families and are highly conserved among distantly related species, we analysed expression of twenty-one miRNA sequences in different tissues of apple (Malus x domestica 'Royal Gala'). We identified eighteen sequences that are expressed in at least one of the tissues tested. Some, but not all, miRNAs expressed in apple tissues including the phloem tissue were also detected in the phloem sap sample derived from the stylets of woolly apple aphids. Most of the miRNAs detected in apple phloem sap were also abundant in the phloem sap of herbaceous species. Potential targets for apple miRNAs were identified that encode putative proteins shown to be targets of corresponding miRNAs in a number of plant species. Expression patterns of potential targets were analysed and correlated with expression of corresponding miRNAs.ConclusionsThis study validated tissue-specific expression of apple miRNAs that target genes responsible for plant growth, development, and stress response. A subset of characterised miRNAs was also present in the apple phloem translocation stream. A comparative analysis of phloem miRNAs in herbaceous species and woody perennials will aid our understanding of non-cell autonomous roles of miRNAs in plants.

According to the Münch hypothesis, solution flow through the phloem is driven by a hydrostatic pressure gradient. At the source, a high hydrostatic pressure is generated in the collection phloem by active loading of solutes, which causes a concomitant passive flow of water, generating a high turgor pressure. At the sink, solute unloading from the phloem keeps the turgor pressure low, generating a source-to-sink hydrostatic pressure gradient. Localised changes in loading and unloading of solutes along the length of the transport phloem can compensate for small, short-term changes in phloem loading at the source, and thus, maintain phloem flow to the sink tissue. We tested directly the hydrostatic pressure regulation of the sieve tube by relating changes in sieve tube hydrostatic pressure to changes in solute flow through the sieve tube. A sudden phloem blockage was induced (by localised chilling of a 1-cm length of stem tissue) while sieve-tube-sap osmotic pressure, sucrose concentration, hydrostatic pressure and flow of recent photosynthate were observed in vivo both upstream and downstream of the block. The results are discussed in relation to the Münch hypothesis of solution flow, sieve tube hydrostatic pressure regulation and the mechanism behind the cold-block phenomenon.

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Abstract. According to the Münch hypothesis, a flow of solution through the sieve tubes is driven by a hydrostatic pressure difference between the source (or collection) phloem and the sink (or release) phloem. A high hydrostatic pressure is maintained in the collection phloem by the active uptake of sugar and other solutes, with a concomitant inflow of water. A lower pressure is maintained in the release phloem through solute unloading. In this work we directly test the role of solute uptake in creating the hydrostatic pressure associated with phloem flow. Solute loading into the phloem of mature leaves of barley and sow thistle was reduced by replacing the air supply with nitrogen gas. Hydrostatic pressure in adjacent sieve elements was measured with a sieve-element pressure probe, a cell pressure probe glued to the exuding stylet of aphids that had been feeding from the phloem. Sieve element sap was sampled by aphid stylectomy; sap osmotic pressure was determined by picolitre osmometry and its sugar concentration by enzyme-linked fluorescence assays. Samples were taken with a time resolution of ∼2-3 min. In accordance with Münch's proposal a drop in osmotic and hydrostatic pressure in the source phloem following treatment of the source leaf with N 2 was observed. A decrease in sugar concentration was the major contributor to the change in osmotic pressure. By observing these variables at a time resolution of minutes we have direct observation of the predictions of Münch.