| Nuclear pore complex (NPC) embeds in the double membrane nuclear envelope (NE), the physical barrier separating the nucleus from cytosol, and serves as the sole channel for nucleocytoplasmic transport of macromolecules essential for cell survival (i.e. proteins and RNAs). Because of the limitations of research tools and materials, it was not until the last decade that the biological functions of these individual nucleoporins just began to be understood.Previously in our lab, a set of candidate nucleoporin genes in Arabidopsis genome has been identified using bioinformatical approaches. After screening the Arabidopsis mutants for those candidate genes, a few lines with discernible developmental phenotypes have been isolated for further analysis and nup96in this thesis is one of them. Nup96is a single copy gene in Arabidopsis genome and encodes a protein whose sequence shows strong similarity with vertebrates Nup96and yeast Nup145C. Nup96is one of the stable constituents of Nup107-160subcomplex in NPC, which interacts with Arabidopsis SEC13protein both in vitro and in vivo. A conserved peptidase domain is included in the N terminus of Arabidopsis Nup96, which remains autoproteolytic activities both in prokaryotic and plant cells. Failure to complete autoproteolysis would not affect the functions of Nup96in Arabidopsis, therefore, the biological function of this proteolysis still needs to be elucidated.Loss-of-function alleles of nup96exhibit a strong early-flowering phenotype regardless of photoperiod, which can be strongly suppressed by co mutation under long days. The flowering of nup96mutants is also non-responsive to different ambient temperatures. Moreover, our data shows that Nup96is required for FLC activation partially through a mechanism involving histone acetylation of FLC chromatin. Nup96mutations also reduce the expression level of miR156genes. In addition, expression of Nup96in both vascular tissue and shoot apical meristems could rescue the early-flowering phenotype of nup96mutants. Thus, we proposed that Nup96could negatively regulate flowering in Arabidopsis through multiple pathways.Mutation of Nup96also has a broad affect on circadian clock function in Arabidopsis. Given by the leaf movement assay and bioluminescence of clock gene promoter activity, we found that nup96lengthens the circadian period both in continuous light and dark, suggesting that Nup96plays an important role in determining the circadian period of circadian clock. Phase response curve shows that nup96mutants are more sensitive to light pulses in the daytime than wild type, indicating that nup96could negatively regulate clock sensitivity for gating light signals, In addition, nup96mutation also results in a temperature overcompensation phenotype and low temperatures completely rescue the long period phenotype of nup96plants. Moreover, nup96mutation also lengthens the circadian period of clock gene expressions. Taken together, we demonstrate that Nup96is a novel regulator in clock period determination. |
