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Abstract

The control of cell division plane orientation and precise organization of growing structures are fundamental processesin developmental biology. In plants, radial growth is mediated by the cambium, a stem cell niche continuously producing xylem and phloem in a strictly bidirectional manner. Cambium stem cells are particularly unusual as they are embedded deeply in very rigid tissues and divide along their longest axis in a highly ordered manner. While several intercellular signaling cascades have been identified to regulate activity and differentiation of cambium stem cells and, thereby, tissue organization during radial growth, the process of guiding cell division orientation and the role of microtubule organization in this context is still unclear. In my study, I characterized microtubule organization in dividing cambium stem cells: during interphase, cortical microtubules were twisted spirally around cambium stem cells. During prophase, the preprophase band (PPB) was absent in cambium stem cells but was replaced by perinuclear microtubules. During metaphase, initial spindle orientation in cambium stem cells was remarkably flexible. In anaphase and telophase, phragmoplast microtubule arrays distinguished periclinal from anticlinal cell divisions. Additionally, I found the orientation of division planes was preserved by the cortical division zone (CDZ) in dividing cambium stem cells in the absence of the PPB. In addition, I compared anatomical analyses with a cell-based vertex model generated by Mathias Höfler (Technische Universität München, TUM) to investigate the nature of mechanical stress at the cell and tissue scale in hypocotyls. Using realistic cellular outlines of the radially growing hypocotyl, Mathias revealed a distinct stress pattern with circumferential stresses in cambium stem cells and typical cells files in the cambium only emerging when divisions were guided by the direction of stress. Supporting the predictive power of the established model, removal of xylem cells in simulations and in plants, lead to the same circumferential realignment of simulated stress patterns as indicated in plants by microtubule reorientation. Moreover, the same alteration in the division orientation of cambium stem cells upon mechano-perturbation in silico and in planta supported an instructive role of mechanical forces in cambium organization. Furthermore, I identified an uncharacterized gene, JING1, to be expressed in cambium stem cells encoding for a plasma membrane-associated protein. Furthermore, JING1 responded to exogenous mechanical stress in a CAMTA3-dependent way. In summary, my results characterize cambium stem cells regarding the cell division process. Cambium stem cells conduct PPB-independent cell divisions and the CDZ is involved in determining the future division plane orientation. My work also underline the significance of mechanical forces in determining the division plane orientation of cambium stem cells through self-emerging stress patterns upon cell proliferation during radial growth. Lastly, my results characterize a cambium stem cell-specific gene, JING1, to be a CAMTA3-dependent, mechanical stress-responsive gene active in cambium stem cells.