Minisymposium 21: Protein and Vesicle Trafficking
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Abs #
M2104: Chemical Genomics of an inhibitor of tonoplast protein trafficking in Arabidopsis.
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Presenter: |
Rojas-Pierce, Marcela Contact Presenter |
Authors | Rojas-Pierce, Marcela (A) Sohn, Eun-Ju (A) Girke, Thomas (A) Raikhel, Natasha V. (A) | | Affiliations: |
(A): Department of Botany and Plant Sciences and Center for Plant Cell Biology (CEPCEB), University Of California
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| Web Site: | http://cepceb.ucr.edu/members/raikhel.htm | |
Lethality and redundancy are common and challenging descriptors in genetic studies of the endomembrane system. Chemical genomics can be used to overcome these challenges and study protein trafficking mechanisms. Chemical 5850247 inhibits the gravitropic response and induces the accumulation of the tonoplast marker GFP-δ-TIP in the Endoplasmic Reticulum (ER) of root cells (Surpin et al., PNAS, 102: 4902). Trafficking of another tonoplast marker, GFP-γTIP-like, is also blocked by chemical treatment. However, only the GFP-δTIP marker displays an ER pattern, suggesting that these two proteins traffic by different pathways. The localization and appearance of markers that label ER, plasma membrane, Golgi apparatus and actin cytoskeleton did not change after treatment with chemical 5850247, indicating that the effect of this chemical is specific to tonoplast protein trafficking. Also, markers for soluble vacuolar proteins accumulated in the proper compartment even though at higher levels when compared to control. Analogs of this chemical were tested to identify the chemical structures that are responsible for bioactivity of this molecule. The analogs displayed different levels of potency for both the gravitropic and the protein targeting phenotypes. Screens for resistant and hypersensitive mutants were carried out with the goal of identifying putative targets. Nine resistant and 14 hypersensitive mutants were identified from screens of EMS mutagenized seeds carrying the GFP-δTIP construct. Characterization and mapping of the most robust mutants is under way.
