Proteomic Analysis Of Arabidopsis In Response To Altered Gravity

Proteomic approaches were applied to analyze responses of Arabidopsis thaliana callus cells and root tips to altered gravity.Callus cells of Arabidopsis thaliana (cv. Landsberg erecta) were exposed for 8 h to a horizontal clinostat rotation. The amount of glucose and fructose apparently decreased, while starch content increased in the H(horizontal) compared with the V-(vertiacal) and S-(still control) treated cells. In order to investigate the influences of clinostat rotation on the cellular proteome further, the proteome alterations induced by horizontal and vertical clinostat rotation have been comparatively analyzed by two-dimensional (2-D) gel electrophoresis and mass spectrometry. Image analysis of silver stained 2-D gels revealed that 80 protein spots showed quantitative and qualitative variations that were significantly (p<0.01) and reproducibly different between the clinorotated (H or V) and the stationary control samples. MS analysis revealed that 18 protein spots showed significant expression alteration only under the H condition compared with those under V and S conditions. These proteins functionally related to general stress responses, metabolic pathways, gene activation/transcription, protein synthesis, and cell wall biosynthesis.Five-day-old seedlings of WT and pin2 mutant were exposed for 12 h to horizontal clinostat rotation (H, simulated weightlessness), vertical clinostat rotation(V, clinostat control), 0.5 h to altered g-forces by centrifugation (7×g, hypergravity), or a stationary control grown conditions. Total proteins of root tips were extracted and separated by two-dimensional gel electrophoresis. About 1300-1500 protein spots were reproducibly detected by ImageMaster 2D Platinum software on silver-stained gels. Image analysis revealed that 54 protein spots in WT and 64 protein spots in pin2 mutant changed their intensities significantly (p<0.05) by more than 1.5-fold and / or their positions after at least one treatment. 38 protein spots were excised from preparative gels and analyzed by LC-IT-MS. 32 protein spots represent 28 proteins were successfully identified, and they falls into several functional categories including stress response, metabolism, protein modification, gravity-sensing and others. The different expression pattern of these proteins in samples treated with clinorotation and centrifugation suggest that changed gravity both in direction and magnitude can induce plant stress response, but different signal pathway and metabolic modulation were involved in the responses. The different expression pattern of these proteins in WT and mutant may be related to repressed transportation of auxin in mutant root tips.