Tesis Doctorado
Identificatión of proteins involved in cell wall synthesis by integratión of high-throughput technologies
Autor
Dupree, Paul
University of Cambridge
Institución
Resumen
The wall is a fundamental component of plant cells that is made of a large diversity of cellulosic and non-cellulosic polysaccharides. Beyond the maintenance of the structure of plants, the cell wallhas an arguably relevant role in human activities, given the extensive use of plant wall material in the form of wood, fibres and more recently, as a promising source of biofuels.Faster advance in the research of the molecular mechanisms underpinning the synthesis of cell wall has been hampered by the difficulties of traditional methods in biochemistry and genetics to deal with the genetic complexity of plants. The high diversity of genes and the lack of evidentphenotypes have torced the use of more laborious screening procedures that result in higher cost and lower productivity of scientific research. The use of high-throughput proteomic and microarraytechniques could accelerate the discovery the discovery of genes involved in synthesis of cell wall.Based on the occurrence of the wall synthetic machinery within the Golgi apparatus, proteomic techniques have been applied already to identify resident proteins in this organelle in an unprecedented scale in Arabidopsis. However the generally poor characterisation of proteins fromplants makes it difficult to ascribe functions to many of the proteins identified in these experiments.Therefore other aspects of cellular organisation such as the formation of complexes and also gene co-expression can be used to approach the interpretation of large sets of proteins from variousperspectives. In this thesis the main goal is to maximise the use of information from high throughput techniques in arder to identify new proteins involved in the synthesis of cell wall. To this end, thedata from proteomics and microarrays was analysed in two stages: firstly a thorough assessment was conducted about basic aspects of individual techniques such as reproducibility of replicates andpredictive capability of the data. Through this assessment a salid base for further interpretation could be generated. Then depending on the particular datasets either, Golgi candidates, potentialcomplexes or co-expressed genes were identified through relevant literature and other bioinformatic resources. As a result, many new candidates for synthesis of the cell wall were identified, with a few of them also identified as likely members of large protein complexes. The datasets from individual techniques were also combined to generate more confident predictions. Forexample, the refinement of potential complexes through the integration with organelle localisation data. Lastly, the exercise of combining and analysing large datasets led to key points and strategiesto improve the efficiency of individual techniques and the downstream analysis of the data.