Study On The Physiological Function Of Arabidopsis Actin Binding Protein Profilin3in

The actin cytoskeleton has emerged as a very pivotal component of eukaryotic cells, which presents a highly dynamic three-dimensional mesh networks. The actin cytoskeleton is not only offers internal mechanical support for cells but also participates in a range of important cellular processes. The function of actin cytoskeleton relies on both actin cytoskeleton own organization and actin dynamics which is regulated by multiple actin binding proteins (ABPs) and many different signals. In plant cells, a large number of ABPs have been identified and involved in different physiological functions. Profilin (PRFs) is a small monomer actin sequestering protein family in all eukaryotes, which is found earlier, highly conserved and widely expressed. The Arabidopsis PRF family contains five highly conservate isoforms. The biochemical functions of this family have been studied for a long time, the physiological function are unclear. PRF3, one of the vegetative isoforms, has36more N-terminal amino acid residues than other PRFs. The physiological and biochemical functions of PRF3are completely unknown. In this study, for the first time, we systematically studied the tissue localization of PRF3, and its physiological function, etc. The major results are as follows:1. PRF3s a vegetative profilin gene. GUS activities and real time PCR analysis were detected in the most parts of the plants demonstrated that PRF3was strongly expressed in young seedlings, rosette leaves, and cauline leaves but weakly expressed in14-d-old seedlings and flowers and that its expression was enhanced by the first intron of PRF3.2. PRF3increased the critical concentration (Cc) of actin assembly in vitro. PRF3-AN36increased the Cc of actin assembly, which was consistent with PRF1and PRF2in Arabidopsis. The Cc of N36was closer to actin-only controls, and the Cc of PRF3was between them. This result may indicate that the N36of PRF3protein has a role for its functions.3. Overexpression of the full-length PRF3cDNA decreased the lengths of roots and hypocotyls and delayed germination, but PRF3-ΔN36transgenic plants and pr/3-1mutants were not, compared with wild-type plants. These results showed that overexpression of the full-length PRF3cDNA affected the lengths of roots and hypocotyls, and PRF3was a key regulator of root and hypocotyl lengths.4. Using light microscopy and scanning electron microscopy, we found that the cell lengths of OE-PRF3-3hypocotyls were much shorter than those in the other three plant lines and that the dwarf phenotype of the OE-PRF3seedlings can be largely accounted for by the reduction in cell length. The results implated that PRF3was important for the elongation of the hypocotyl cells.5. Confocal observation showed that thick actin cables were absent and fine actin filaments appeared less affected in the hypocotyl cells of the OE-PRF3-3seedlings. These results showed that overexpression of PRF3induce F-actin arrangement in hypocotyl cells and that thick actin cables might be associated with cell elongation.6. We observed that reorganization of the actin cytoskeleton was triggered by cold or heat stress. The actin cytoskeletonal organization responds to temperature stresses. The MFs were considerably rearranged, and some MFs were depolymerized under cold stress; the applied heat shock stress led to changes in the MF arrays in Arabidopsis hypocotyl cells.7. Facilitation of microfilament (MF) assembly by phalloidin treatment increased the resistance of Arabidopsis seedlings to temperature stress, whereas blocking MF assembly with latrunculin B decreased their temperature tolerance.8. Using real-time PCR, we found that the expression patterns of some of the vegetative profilins and actin depolymerizing factors (ADFs) were specifically altered under various stresses, such as ADF6. These results showed that the expression profiles of different genes were different under the same stress, and different expression patterns for the same gene were also observed. Thus, our results show that PRF3is vital for the development of Arabidopsis seedlings., especially the growth of roots, root hairs, and hypocotyls. The dwarf phenotype of the PRF3-overexpression seedlings can be largely accounted for by the reduction in cell length and F-actin arrangement, especially the disappearance of thick, longitudinally oriented cables. The in vivo results may indicate that the N36of PRF3protein has a role for its functions. Our results show that the actin cytoskeleton is vital for the tolerance of Arabidopsis seedlings to heat/cold stresses. The results of real time PCR will be valuable for studying the functions of ABPs in abiotic stresses and uncovering new components of stress signal transduction. In this study, we provide a theoretical foundation for studying the physiological function of the plant actin cytoskeleton and its binding proteins, and exploring the mechanism of epigenetic regulation in plant abiotic stresses tolerance.