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Functional characterization of tomato Sl-IAA3 and Sl-hls genes. Role ...

Functional characterization of tomato Sl-IAA3 and Sl-hls genes. Role ...

CHAPITRE III. HOOKLESS1,

CHAPITRE III. HOOKLESS1, an Integrator of Multiple Signaling Pathways in Arabidopsis Ethylene HLS1 Glucose GIN2/At-HXK ABA GIN6/ABI4 Germination Cotyledons greening Figure 6. A model for the role of HLS1 in enhancing glucose sensitivity compatible with ethylene being a repressor of glucose sensitivity Glucose enhances the the proteasome-mediated degradation of the nuclear EIN3 protein, while ethylene enhances its stability. Considering that HLS1 is homologue to diverse class of N-acetyltransferase, it is possible that HLS1 could be implicated in the regulation of protein stability of EIN3 (Ohto et al., 2006). Strikingly, expression analysis performed in this study indicate that HLS1 gene is not regulated by glucose at the transcriptional level and that gin1, ctr1, etr1 mutations do not affect the accumulation of HLS1 transcript in presence or absence of glucose (data not shown). We conclude that HLS1 implication in the glucose signalling pathway may be at the post transcriptional level. Taken together, these data clearly indicate that HLS1 gene is an integrator of many signalling pathways including ethylene, light, glucose, as well as gravitropic response. Efforts are currently being directed towards uncovering the molecular mechanisms by which HLS1 impacts all these singling pathways. 97

METHODS CHAPITRE III. HOOKLESS1, an Integrator of Multiple Signaling Pathways in Arabidopsis Plant Material hls1-1 in the Columbia ecotype was obtained from the Nottingham Arabidopsis Stock Center. Arabidopsis (Columbia) was always used as control. Plants were grown under standard green house condition. Greening Test Sterilized seeds were put on Murashige and Skoog agar medium plates and placed at 4°C for 2 days. After the seeds were kept at 25°C in the dark for a number of days. They were then exposed to white light with a fluence rate (µmol.m -2 .s -2 ) for 48 h. Each seedling was then assessed visually as to whether it was green or still yellow. The experiments were done independently more than 2 times. Glucose Sensitivity The glucose-resistant phenotype was scored by growing sterilized seeds on Murashige and Skoog agar medium containing 6.5 % (w/v) filter-sterilized Glc. After 2 days at 4°C, plates were placed in light in a sealed box containing either air or 10 µL L -1 ethylene. Evry 2 days, the boxes were opened to allow a renewal of the atmosphere and put back either with air or 10 µL L -1 ethylene. The experiment was stopped after 10 days of culture. ABA Treatment ABA (Lomon, bio) was prepared as a 10 mM stock in NAOH and diluted into appropriate concentrations (100 nM). ABA sensitivity tests were done in darkness using agar medium supplemented with 100 nM of ABA and scored after 5 days after sowing. REFERENCES Achard, P., Vriezen, W.H., Van Der Straeten, D. and Harberd, N.P. (2003) Ethylene regulates Arabidopsis development via the modulation of DELLA protein growth repressor function. Plant Cell 15: 2816–2825. Arenas-Huertero, F., Arroyo, A., Zhou, L., Sheen, J. and Leon, P. (2000). Analysis of Arabidopsis glucose insensitive mutants, gin5 and gin6, reveals a central role of the plant hormone ABA in the regulation of plant vegetative development by sugar. Genes Dev. 14, 2085– 2096 98

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