Thesis or Dissertation Molecular genetic study on gravitropism and gravitropic growth habit of the Arabidopsis shoot

佐々木, 秋

Gravitropic response is a control of growth direction with respect to the gravity vector. Gravity sensing process starts with downward movement of amyloplasts in statocytes and subsequent physiological signaling is presumed to regulate auxin transport. Putative signaling components, Arabidopsis LAZY1 (AtLAZY1) and ALTERED RESPONSE TO GRAVITY 1 (ARG1) are involved in negative gravitropism (upward bending) of Arabidopsis shoots. However, little is known about relationships between them. In this study, I examined genetic interaction between AtLAZY1 and ARG1 using various mutants of Arabidopsis. The double mutant atlazy1 arg1 showed synergistic loss of stem gravitropism, implicating that there are multiple pathways of gravitropic signaling. Auxin transporter genes PIN-FORMED 3 (PIN3), PIN4 and PIN7 did not genetically interact with AtLAZY1. I isolated a novel mutant dsl1 that suppressed atlazy1 defect. dsl1 did not suppressed defects of sgr2 and eal1 that were mutants of amyloplast sedimentation, suggesting that dsl1 specifically suppresses the AtLAZY1-dependent signaling pathway. dsl1 encoded ASL5/LBD12 transcription factor and microarray analyses revealed that secondary cell wall deposition was significantly upregulated in dsl1, while expression of auxin signaling components was not altered. Moreover, I found that impaired secondary cellulose biosynthesis and inhibition of cellulose biosynthesis by isoxaben treatment attenuated stem gravitropism. These results indicate that cell wall biogenesis plays an important role in AtLAZY1-dependent gravitropism of Arabidopsis stems. To further investigate how AtLAZY1 functioned in gravitropism, we used fluorescence microscopy to examine the subcellular localization of AtLAZY1 and its truncated proteins fused to GFP in tobacco leaves. I found that AtLAZY1 localized to the plasma membrane through the C-terminal region, indicating that the putative transmembrane domain in the N-terminal half is not required for its localization. Next, I took a biochemical approach to investigate the membrane association of AtLAZY1. Transiently expressed AtLAZY1 in transgenic Arabidopsis was fractionated in an insoluble fraction that contained membranous compartments. The AtLAZY1 protein was solubilized by a non-ionic detergent or at a high pH condition, suggesting that AtLAZY1 is a peripheral membrane protein. I also found that when expressed in tobacco the C-terminal part of AtLAZY1 co-localized with microtubules. A microtubule binding assay showed that the C-terminal half of AtLAZY1, which localized to the plasma membrane, interacted with microtubules in vitro. These results suggest that AtLAZY1 may function with microtubules at the periphery of the plasma membrane in the gravity signaling process.
vii, 85p
Hokkaido University(北海道大学). 博士(生命科学)

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