Assessing cell proliferation dynamics is crucial to understand the spatiotemporal control of organogenesis. Here we have generated a versatile fluorescent sensor, PlaCCI (plant cell cycle indicator) on the basis of the expression of CDT1a-CFP, H3.1-mCherry and CYCB1;1-YFP, that identifies cell cycle phases in Arabidopsis thaliana. This tool works in a variety of organs, and all markers and the antibiotic resistance are expressed from a single cassette, facilitating the selection in mutant backgrounds. We also show the robustness of PlaCCI line in live-imaging experiments to follow and quantify cell cycle phase progression.Development in multicellular organisms depends on the highly regulated formation of new organs. Organogenesis requires the production of a sufficient number of cells in a temporally and spatially regulated manner, that eventually will constitute the cohort of different cell types present in the organ. Therefore, understanding morphogenesis requires the availability of confident markers to assess cell proliferation and identify unequivocally the different cell cycle phases in a living cell environment.Efforts in several animal models have led to the development of the FUCCI (fluorescent, ubiquitination-based cell cycle indicator) systems that identify each cell cycle phase [1][2][3][4] . Comparable advances for live-imaging of cell proliferation studies in plants lag behind. One reason is that plants lack the CDT1a-interacting protein, geminin, a key component of the FUCCI system, although they contain a CDT1a-interacting protein (GEM) that is structurally unrelated to animal geminin 5,6 . Attempts to circumvent this have been reported. Thus, in addition to the tags used for G2 + M (refs. 7,8 ), the expression of a truncated version of CDT1a, a DNA replication initiator protein, from a heterologous promoter has been reported as an S + G2 marker 9 . In another approach, the fluorescent protein Venus fused to the destruction box of CYCB1;1 was expressed under the control of H4 promoter and labelled cells in S + G2 + M (ref. 10 ). However, a comprehensive cell cycle marker line should ideally (1) be able to distinguish in an unequivocal manner all cell cycle phases to follow individual cells in a growing organ and (2) offer the possibility of using advanced live-imaging strategies. This is precluded when using nucleoside analogues for labelling the S-phase, since they require specific detection protocols incompatible with live imaging.Still, one major drawback of cell cycle marker lines developed so far is the lack of appropriate markers to identify unequivocally cells in G1. We have recently analysed in detail CDT1a dynamics in proliferating cells and demonstrated that CDT1a starts to be loaded into chromatin shortly after mitosis, reaches a maximum during G1 and is rapidly degraded only minutes after S-phase initiation 11 , expanding previous claims of its proteasome-dependent degradation 12 . Importantly, we demonstrated that CDT1a accumulates rapidly after the S-phase of endocycling cells and is...

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Mammary stroma is essential for epithelial morphogenesis and development. Indeed, postnatal mammary gland (MG) development is controlled locally by the repetitive and bi-directional cross-talk between the epithelial and the stromal compartment. However, the signalling pathways involved in stromal–epithelial communication are not entirely understood. Here, we identify Sfrp3 as a mediator of the stromal–epithelial communication that is required for normal mouse MG development. Using Drosophila wing imaginal disc, we demonstrate that Sfrp3 functions as an extracellular transporter of Wnts that facilitates their diffusion, and thus, their levels in the boundaries of different compartments. Indeed, loss of Sfrp3 in mice leads to an increase of ductal invasion and branching mirroring an early pregnancy state. Finally, we observe that loss of Sfrp3 predisposes for invasive breast cancer. Altogether, our study shows that Sfrp3 controls MG morphogenesis by modulating the stromal-epithelial cross-talk during pubertal development.