Actin filaments mediated root growth inhibition by changing their distribution under UV-B and hydrogen peroxide exposure in Arabidopsis

BACKGROUND: UV-B signaling in plants is mediated by UVR8, which interacts with transcriptional factors to induce root morphogenesis. However, research on the downstream molecules of UVR8 signaling in roots is still scarce. As a wide range of functional cytoskeletons, how actin filaments respond to UV-B-induced root morphogenesis has not been reported. The aim of this study was to investigate the effect of actin filaments on root morphogenesis under UV-B and hydrogen peroxide exposure in Arabidopsis. RESULTS: A Lifeact-Venus fusion protein was used to stain actin filaments in Arabidopsis. The results showed that UV-B inhibited hypocotyl and root elongation and caused an increase in H2O2 content only in the root but not in the hypocotyl. Additionally, the actin filaments in hypocotyls diffused under UV-B exposure but were gathered in a bundle under the control conditions in either Lifeact-Venus or uvr8 plants. Exogenous H2O2 inhibited root elongation in a dose-dependent manner. The actin filaments changed their distribution from filamentous to punctate in the root tips and mature regions at a lower concentration of H2O2 but aggregated into thick bundles with an abnormal orientation at H2O2 concentrations up to 2 mM. In the root elongation zone, the actin filament arrangement changed from lateral to longitudinal after exposure to H2O2. Actin filaments in the root tip and elongation zone were depolymerized into puncta under UV-B exposure, which showed the same tendency as the low-concentration treatments. The actin filaments were hardly filamentous in the maturation zone. The dynamics of actin filaments in the uvr8 group under UV-B exposure were close to those of the control group. CONCLUSIONS: The results indicate that UV-B inhibited Arabidopsis hypocotyl elongation by reorganizing actin filaments from bundles to a loose arrangement, which was not related to H2O2. UV-B disrupted the dynamics of actin filaments by changing the H2O2 level in Arabidopsis roots. All these results provide an experimental basis for investigating the interaction of UV-B signaling with the cytoskeleton.

A simple and effective pressure culture system modified from a transwell cell culture system

Mechanical pressure plays an important role in many physiological and pathological processes. Mimicking the mechanical pressure present in vitro is necessary for related research, but usually requires expensive and complicated equipment. In this study we created a simple pressure culture system based on the transwell culture system. By cutting off the top rim of the transwell insert, the cells were compressed between the insert membrane and the well floor. The new pressure culture system was proven effective in that it induced cell morphological change, integrin β1 upregulation, actin polymerization and growth change in rat retinal ganglion cells, human nasopharyngeal carcinoma cells and mice embryonic fibroblasts. Though the pressure value is immeasurable and inhomogeneous, the easily available culture system still provides a choice for the laboratories that do not have access to the better, but much more expensive pressure culture equipment.

Molecular imaging of membrane proteins and microfilaments using atomic force microscopy

Atomic force microscopy (AFM) is an emerging technique for a variety of uses involving the analysis of cells. AFM is widely applied to obtain information about both cellular structural and subcellular events. In particular, a variety of investigations into membrane proteins and microfilaments were performed with AFM. Here, we introduce applications of AFM to molecular imaging of membrane proteins, and various approaches for observation and identification of intracellular microfilaments at the molecular level. These approaches can contribute to many applications of AFM in cell imaging.

beta PAK-interacting exchange factor may regulate actin cytoskeleton through interaction with actin

p21-activated kinase (PAK)-interacting exchange factor (PIX) is known to be involved in regulation of Cdc42/Rac GTPases and PAK activity. PIX binds to the proline-rich region of PAK, and regulates biological events through activation of Cdc42/Rac GTPase. To further investigate the role of PIX we produced monoclonal antibodies (Mab) against beta PAK. Three clones; N-C6 against N-terminal half and C-A3 and C-B7 against C- terminal half of beta PAK were generated and characterized. N-C6 Mab detected beta PAK as a major band in most cell lines. C-A3 Mab recognizes GIT-binding domain (GBD), but it does not interfere with GIT binding to beta PAK. Using C-A3 Mab possible beta PAK interaction with actin in PC12 cells was examined. beta PAK Mab (C-A3) specifically precipitated actin of the PC12 cell lysates whereas actin Mab failed to immunoprecpitate beta PAK. Co-sedimentation of PC12 cell lysates with the polymerized F-actin resulted in the recovery of most of beta PAK in the cell lysates. These results suggest that beta PAK may not interact with soluble actin but with polymerized F-actin and revealed that beta PAK constitutes a functional complex with actin. These data indicate real usefulness of the beta PAK Mab in the study of beta PAK role(s) in regulation of actin cyoskeleton.

The cytoskeleton of the electric tissue of Electrophorus electricus, L

The electric eel Electrophorus electricus is a fresh water teleost showing an electrogenic tissue that produces electric discharges. This electrogenic tissue is distributed in three well-defined electric organs which may be found symmetrically along both sides of the eel. These electric organs develop from muscle and exhibit several biochemical properties and morphological features of the muscle sarcolema. This review examines the contribution of the cytoskeletal meshwork to the maintenance of the polarized organization of the electrocyte, the cell that contains all electric properties of each electric organ. The cytoskeletal filaments display an important role in the establishment and maintenance of the highly specialized membrane model system of the electrocyte. As a muscular tissue, these electric organs expresses actin and desmin. The studies that characterized these cytoskeletal proteins and their implications on the electrophysiology of the electric tissues are revisited.