[Truncated abstract] The phloem long distance translocation system is not only involved in the transport of nutrients and photo-assimilates to different organs of the plant, but it also appears to be important for the transport of information molecules including growth-regulators, proteins and RNA. Translocation of signals appears to be involved in the coordination of developmental processes and also in the response of the plant to environmental cues. Much of the information about macromolecules in phloem comes from analyses of exudates collected from the stylets of sap sucking insects or from incisions made to the vasculature. Among the legumes, members of the genus Lupinus exude phloem 'freely' from incisions made to the vasculature at most organs of the plant. This feature was exploited in this study to document some of the macromolecules present in exudate of L. albus and which might represent potential mobile signals. Phloem exudate was collected mainly from the sutures of developing pods and from inflorescence racemes. Two-dimensional polyacrylamide gel electrophoresis and tandem mass spectrometry were used to identify 83 proteins in exudate. Analysis of a cDNA library constructed from exudate identified 609 unique transcripts. Both proteins and mRNA were classified into functional groups. The largest group was related to general and energy metabolism, suggesting some metabolic activity probably to support the sieve element (SE). Other significant functional groups were represented by proteins and transcripts involved in protein synthesis, turnover and sorting, and in redox homeostasis. Proteins in these categories could play a role in maintaining the functions and stability of proteins in SE. Macromolecules involved in signalling such as transcripts encoding proteins mediating calcium levels and the Flowering locus T (FT) protein were also identified in phloem exudate of L. albus. FT protein has been recently identified as a mobile signal that induces flowering. ... The hen1 mutant accumulates low, sometimes even undetectable levels of miRNA due to the lack of methylation. No translocation of the five miRNA assayed under nutrient replete (non stress) conditions was observed. Translocation of miR395 in response to sulphur (S) deficiency was also investigated, and while conclusive evidence of translocation was not obtained, the data suggested some movement from roots to shoots (possibly in xylem) of a signal in response to S-deficiency. Future work is required to provide greater insight into the translocation path and identity of this S-deficiency signal. This study suggests that not all miRNA identified in phloem exudates are mobile, which raises the question about their biological relevance in SE and how they reached this location (e.g. through the action of a non-selective transport mechanism). However, there is also the possibility that miRNA are translocated only in response to specific internal or external cues not tested in this study. This is the first study that provides information on macromolecules present in the phloem exudate of a member of the Fabaceae. The information obtained from this work, provides a basis for future studies in the identification of potential mobile signals that may play a role in a communication network that traffics information around the plant, regulating its various developmental processes and responding to environmental cues.
|Qualification||Doctor of Philosophy|
|Publication status||Unpublished - 2009|