Body of Abstract: Cell polarity fundamentally shapes the development of multicellular organisms. Preliminary data from my research group suggest that an environmental signal – white light – can induce polarity in Marchantia polymorpha spores. We identified directional blue light as the main polarizing cue.
Based on steady-state mRNA level measurements during spore polarization, I hypothesize that the blue light receptors PHOTOTROPIN (MpPHOT), CRYPTOCROME (MpCRY), and FLAVIN-BINDING KELCH REPEAT F-BOX PROTEIN (MpFKF) play a role in translating these directional blue light cues into de novo polarity in developing spores. To test my hypothesis, I generated mutant lines with loss of function mutations in the MpPHOT, MpCRY, and MpFKF genes employing the CRISPR-Cas9 system.
I am characterizing cell polarization and initial divisions of these mutant spores under blue light conditions. Preliminary data are consistent with the hypothesis that a blue light receptor is required for polarization. While the cell plate that forms during the first cell division in wild-type spores is always oriented perpendicular to the direction of blue light, the cell plates that form in Mpphot spores are randomly oriented. Through the generation of MpPHOT-VENUS (YFP) protein fusions, I am studying the subcellular localization of MpPHOT during spore polarization. These reporter lines facilitate testing the hypothesis whether specific localization of MpPHOT sets up polarity de novo.
Using a combination of genetic and imaging approaches, I aim to define the role and molecular networks of blue light receptors in de novo polarity formation in Marchantia polymorpha. For example, I am employing a proximity labeling approach to identify interaction partners of MpPHOT. Functional characterization of strong gene candidates will define the molecular mechanism for MpPHOT-mediated polarization. Ultimately, my work proposes to describe how blue light receptors control de novo polarization in a single isolated cell that eventually gives rise to a complex multicellular plant.