75 Integrating Membrane Transport with Male Gametophyte ... - TAIR

111 Characterization of the Anti-microtubule Drug Supersensitive Arabidopsis Mutant 28-2b
Charitha Galva 1 , Alex Paradez 2 , John Sedbrook 1
1
Department of Biological Sciences, Illinois State University, Normal, Illinois 61790., 2 Carnegie Institution,
Department of Plant Biology, Stanford, California 94305.
Microtubules assemble into highly organized arrays that are essential for nuclear migration, cytokinesis, and cell
expansion. In plants, microtubules form a cortical array that lines the periphery of each cell and plays a role in regulating
and directing cell wall deposition and cell expansion. Little is known about the mechanisms underlying the organization
of this array, or even how it carries out its functions. To learn more, we are taking a molecular genetic approach by
characterizing an Arabidopsis mutant line 28-2b whose cortical microtubule arrays are hypersensitive to disruption by
the anti-microtubule drug oryzalin. On agar growth medium containing 150 nanomolar oryzalin, wild type Arabidopsis
roots grow normally, while 28-2b root tips swell and stop growing due to disruption of cell expansion and division.
The roots of the 28-2b mutant are visibly indistinguishable from wild type in the absence of the drug oryzalin. We have
mapped the 28-2b mutation to a 70 kb interval on the lower arm of chromosome three, and are working to identify the
affected gene. The efforts to clone this gene as well as the implications of the 28-2b oryzalin supersensitive phenotype
will be discussed.
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112 Making sense of the multitude of chloroplast protein import receptors in Arabidopsis
thaliana
Bianca Hust 1 , Andrea Voigt 2 , Angelika Schierhorn 3 , Mario Jakob 1 , Ralf Bernd Kloesgen 1 , Michael Gutensohn 1
1
Institute of Plant Physiology, Martin-Luther-University Halle-Wittenberg, Germany, 2 Plant Physiology,
Humboldt University Berlin, Germany, 3 Department of Biochemistry, Martin-Luther-University Halle-
Wittenberg, Germany
The majority of chloroplast proteins are encoded in the nucleus and therefore have to be imported into the organelle
after their synthesis in the cytosol as precursor proteins carrying N-terminal transit peptides. The transport of precursor
proteins across the outer and inner chloroplast envelope membrane is mediated by their interaction with two import
machineries, the Toc and Tic complex, respectively. The core of the Toc complex essentially consists of two receptor
proteins involved in the initial binding of precursor proteins at the chloroplast surface, Toc34 and Toc159, and the
translocation pore, Toc75. In the genome of Arabidopsis thaliana two homologs of Toc34 (atToc33, atToc34) and four
homologs of Toc159 (atToc159, atToc132, atToc120, atToc90) have been identified. For the functional characterization we
have isolated knockout mutants for each of the six Arabidopsis Toc receptors. These mutants show remarkably different
phenotypes suggesting more specialized functions for the import receptors. Examination of the proteome and gene
expression profiles of the atToc33, atToc34, atToc159 and atToc132 mutants showed that in each case the accumulation
of a specific subset of chloroplast proteins is affected. Further in vitro experiments using overexpressed receptor domains
and specific antibodies suggest that each of these Toc receptors preferentially interacts with different groups of precursor
proteins. Taken together the results indicate different substrate specificities of the chloroplast protein import receptors.