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Functional Characterisation of the Plant Nuclear Pore Complex

Final Report Summary - PLANTNPC (Functional Characterisation of the Plant Nuclear Pore Complex)

The Nuclear Pore Complex (NPC) is a large protein grouping that controls movement of molecules between the nucleus and the cytoplasm in all eukaryotic cells. It is made up of many individual nucleoporin (NUP) proteins that are arranged into different sub-complexes and repeated in eight-fold symmetry around a central channel. Much of our knowledge about the NPC comes from studies in yeast and animal cells yet we know very little about how this structure functions in plants. Therefore the aim of this project was to characterise the Plant NPC in the model organism Arabidopsis thaliana using genetic and cell biological techniques.
During the course of the project we have isolated many nucleoporin mutants that show a variety of phenotypes. These include A, those mutants that are inviable, showing that these NUP proteins are essential for cell growth, B, those mutants that are early flowering and small in stature, showing the NUP proteins are likely involved in many signaling pathways and C, those mutants that have a similar phenotype to wildtype plants, making it difficult for us to assess their specific function.
At the cellular level we showed that a number of nup mutants have altered nuclear shape and show a defect in nuclear mRNA export. These are novel findings and for the first time we have quantified the latter response using a low-resolution imaging of root nuclei.
On a global scale we assessed changes in gene expression that occur when the function of the NPC is compromised by performed both microarray and a RNA-sequencing (RNAseq) experiments. The former experiment gave the interesting and surprising result showing that in nup mutants, a number of genes involved in nuclear transport are upregulated. This hints towards a previously uncharacterised mode of regulation of nuclear transport and suggests a link between the function of the NPC and the transport processes it helps control.
In order study this phenomenon in more detail we have performed an RNAseq experiment using RNA samples isolated from ribosomes, with the aim of deciphering if there is preferential nuclear export of transcripts in different nup mutants. The information provided by the RNAseq data will set up the future result in the lab as will provide important insight into the specificity that occurs at the level of the NPC. Both the work that has occurred during the CIG-funded period and in the future experiments provided for by the CIG-funding will hopefully enable the establishment of a productive laboratory to investigate plant nuclear transport, a topic on which we are a world leading authority.