nemhauser lab
department of biology
university of washington
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Building tools to study signaling dynamics

Summary: There is an often-told allegory about the differences in methodology between genetics and biochemistry that involves answering the question, ‘How does a car work?’. In brief, the geneticist approaches the problem by removing parts one-by-one. By eliminating the steering wheel, for example, the geneticist learns what controls the direction the car will travel. In contrast, the hapless biochemist completely disassembles the car and attempts to intuit function of each part by studying it in isolation. While this historic division between disciplines has largely been erased (and researchers have largely stopped publishing variations of the story that highlight the foolishness of the other discipline), a new challenge to understanding complex biological systems has emerged: once you know all of the important parts of an organism, how do you connect their static function with the dynamic, integrative operations required for survival? To keep with the metaphor: what allows a car to maintain speed while navigating a winding stretch of highway? We are taking a variety of approaches to bring the current understanding of plant biology to this next level.

Building dynamic networks from the ground up. Auxin is a plant hormone that plays a key role in nearly every aspect of plant biology. Direct experimental tests of signaling dynamics in this crucial pathway are confounded by the ubiquity of auxin response in plant cells. In collaboration with Eric Klavins in the UW Electrical Engineering Department, we have developed an alternative approach where we are systematically transplanting the auxin response pathway from Arabidopsis into the single-celled yeast Saccharomyces cerevisiae. An analogy to our approach is trying to understand how a radio works by removing components one by one, reconnecting each part in a simple setting, and characterizing the resulting circuits in great detail. We have successfully transferred the nuclear auxin response pathway from auxin perception through activation of transcription—a rather remarkable feat highlighting the fundamental conservation of core eukaryotic cell biology. This trans-kingdom transplant of a multi-component pathway has led to a number of novel insights into auxin signaling dynamics and produced a toolkit to examine diverse facets of cell function with unprecedented accuracy.

Synthetic auxin-induced transcription in yeast. All of the parts of the network transferred to yeast are shown as a circuit diagram to the left. Auxin addition to the yeast culture at time 0 triggers degradation of repressors (labeled with YFP, a protein that fluoresces yellow) and activation of a reporter (CFP, a protein that fluoresces cyan).

Dynamics of growth control during development. Using time-lapse analysis of plant growth in combination with a range of molecular genetic tools, we have found that seedling growth dynamics vary over developmental time and that the specific proteins that control growth also vary over time. We are excited about the emerging picture of a fluid transcription factor landscape directing dynamic fluctuations in both transcription and growth.

Dynamic DNA. The orchestrated binding of transcriptional activators and repressors to DNA sequences defines the regulatory program of eukaryotic genomes. A large number of previous studies tell us which genes may be important for plant responses, but we know very little about the regulatory regions orchestrating these global transitions. In collaboration with John Stamatoyannopoulos and Christine Queitsch in the UW Genome Sciences Department, the Nemhauser Lab has applied a high-throughput experimental technology called digital DNaseI footprinting to delineate regulatory DNA across the Arabidopsis genome. Using this approach, we have precisely mapped regulatory DNA dynamics in response to light and heat—two of the most important environmental cues for developing seedlings.

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