Members of the TRANSPORT INHIBITOR RESPONSE1/AUXIN SIGNALING F-BOX PROTEIN (TIR1/AFB) family are known auxin receptors. To analyze the possible receptor function of AUXIN BINDING PROTEIN1 (ABP1), an auxin receptor currently under debate, we performed different approaches. We performed a pharmacological approach using α-(2,4-dimethylphenylethyl-2-oxo)-indole-3-acetic acid (auxinole), α-(phenylethyl-2-oxo)-indole-3-acetic acid (PEO-IAA), and 5-fluoroindole-3-acetic acid (5-F-IAA) to discriminate between ABP1- and TIR1/AFB-mediated processes in Arabidopsis (). We used a peptide of the carboxyl-terminal region of AtABP1 as a tool. We performed mutant analysis with the null alleles of , and , and the TILLING mutant We employed Coimbra, an accession that exhibits an amino acid exchange in the auxin-binding domain of ABP1. We measured either volume changes of single hypocotyl protoplasts or hypocotyl growth, both at high temporal resolution. 5-F-IAA selectively activated the TIR1/AFB pathway but did not induce protoplast swelling; instead, it showed auxin activity in the hypocotyl growth test. In contrast, PEO-IAA induced an auxin-like swelling response but no hypocotyl growth. The carboxyl-terminal peptide of AtABP1 induced an auxin-like swelling response. In the -related mutants and Coimbra, no auxin-induced protoplast swelling occurred. ABP1 seems to be involved in mediating rapid auxin-induced protoplast swelling, but it is not involved in the control of rapid auxin-induced growth.
In an era of genomics, proteomics, and metabolomics a large number of mutants are available. The discovery of their phenotypes is fast becoming the bottleneck of molecular plant physiology. This crisis can be overcome by imaging-based phenotyping, an emerging, rapidly developing and innovative approach integrating plant and computer science. A tremendous amount of digital image data are automatically analysed using techniques of 'machine vision'. This minireview will shed light on the available imaging strategies and discuss standard methods for the automated analysis of images to give the non-bioinformatic reader an idea how the new technology works. A number of successful platforms will be described and the prospects that image-based phenomics may offer for elucidating hormonal cross-talk and molecular growth physiology will be discussed.
we limited the maximum time of the season to 10% of the weeks per municipality. And in the third constraint we limited the season to the weeks 0-14 (See Fig. 4 and "Methods" for details).
Next generation phenotyping of auxin response mutants will be greatly facilitated by the ability to record rapid growth responses in roots and hypocotyls at high throughput and at high temporal resolution. As Arabidopsis seedlings are very tiny and fragile, imaging is the only adequate way for data acquisition. As camera-based systems described before have a limited throughput, we used commercial flatbed scanners to record a large number of simultaneous experiments. We developed Hansa Trace, software for automatically detecting and measuring hypocotyl segments and roots in the images. We validated this system by measuring some well-characterized growth responses to auxins, non-auxins, ATPase activators and apoplastic acidification. The method can be shared on a cooperation basis and is able to perform measurements with minimal user intervention.
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