Minisymposium 21: Emerging technologies
Abs #
42002: Laser-Guided Cell Capture Methods for Abscission Studies
Plant organ detachment is dependent upon separation of discrete cell layers within an abscission zone (AZ). AZ cells exhibit a unique hormone response capacity relative to surrounding cells. For example, cell expansion in AZs is promoted by ethylene and suppressed by auxin, behavior distinct from that of most plant cells. Our group is interested in identifying cell-specific mechanisms that regulate plant organ shed. Arabidopsis is an ideal system in which to exploit functional genomic strategies to reveal novel determinants of abscission control. But petal shed in this species takes place in a restricted tier of very small AZ cells whose manual dissection from surrounding tissues presents a technological hurdle. The historic absence of methods to isolate pure populations of AZ cells for biochemical and molecular analyses has greatly constrained progress in understanding key elements of abscission control. Now, published reports of laser-guided strategies to harvest individual plant cell types provide us with a foundation for developing Arabidopsis as a functional genomic abscission system. Using both the Arcturus PixCell laser capture microdissection (LCM) system (for background information see www.arctur.com ) and Zeiss’s PALM system for laser pressure catapulting (www.palm-microlaser.com) we have developed protocols to laser-capture morphologically distinct petal AZ cells from paraffin sections of flowers engaged in early stages of developmental abscission. Early analyses of RNA from pools of laser-captured abscission zones and surrounding cells established the high RNA purity and integrity required for genomic studies, and we have now captured sufficient numbers of AZ cells to generate RNA for preparing hybridization targets in microarray studies. Gene profiling studies in progress are expected to establish the feasibility of using laser-captured AZ cells in an Affymetrix-based microarray system. In future work, our cell-specific approach will facilitate identification of genes and gene populations whose expression may contribute to the unique hormone responsiveness of AZ cells and regulate other key processes within the abscission program.
