Phosphorylation of the Polarity Protein BASL Differentiates Asymmetric Cell Fate through MAPKs and SPCH.

Cell polarization is commonly used for the regulation of stem cell asymmetric division in both animals and plants. Stomatal development in Arabidopsis, a process that produces breathing pores in the epidermis, requires asymmetric cell division to differentiate highly specialized guard cells while maintaining a stem cell population [1, 2]. The BREAKING OF ASYMMETRY IN THE STOMATAL LINEAGE (BASL) protein exhibits a polarized localization pattern in the cell and is required for differential cell fates resulting from asymmetric cell division [3]. The polarization of BASL is made possible by a positive feedback loop with a canonical mitogen-activated protein kinase (MAPK) pathway that recruits the MAPKK kinase YODA (YDA) and MAPK 6 (MPK6) to the cortical polarity site [4]. Here, we study BASL intracellular dynamics and show that the membrane-associated BASL is slowly replenished at the cortical polarity site and that the mobility is tightly linked to its phosphorylation status. Because BASL polarity is only exhibited by one daughter cell after an asymmetric cell division, we study how BASL differentially functions in the two daughter cells. The YDA MAPK cascade transduces upstream ligand-receptor signaling [5-13] to the transcription factor SPEECHLESS (SPCH), which controls stomatal initiation and is directly suppressed by MPK3/6-mediated phosphorylation [14, 15]. We show that BASL polarization leads to elevated nuclear MPK6 signaling and lowered SPCH abundance in one of the two daughter cells. Therefore, two daughter cells are differentiated by BASL polarity-mediated differential suppression of SPCH, which may provide developmental plasticity in plant stem cell asymmetric cell division (ACD).

Single-cell dynamics and variability of MAPK activity in a yeast differentiation pathway.

In response to pheromones, yeast cells activate a MAPK pathway to direct processes important for mating, including gene induction, cell-cycle arrest, and polarized cell growth. Although a variety of assays have been able to elucidate signaling activities at multiple steps in the pathway, measurements of MAPK activity during the pheromone response have remained elusive, and our understanding of single-cell signaling behavior is incomplete. Using a yeast-optimized FRET-based mammalian Erk-activity reporter to monitor Fus3 and Kss1 activity in live yeast cells, we demonstrate that overall mating MAPK activity exhibits distinct temporal dynamics, rapid reversibility, and a graded dose dependence around the KD of the receptor, where phenotypic transitions occur. The complex dose response was found to be largely a consequence of two feedbacks involving cyclin-mediated scaffold phosphorylation and Fus3 autoregulation. Distinct cell cycle-dependent response patterns comprised a large portion of the cell-to-cell variability at each dose, constituting the major source of extrinsic noise in coupling activity to downstream gene-expression responses. Additionally, we found diverse spatial MAPK activity patterns to emerge over time in cells undergoing default, gradient, and true mating responses. Furthermore, ramping up and rapid loss of activity were closely associated with zygote formation in mating-cell pairs, supporting a role for elevated MAPK activity in successful cell fusion and morphogenic reorganization. Altogether, these findings present a detailed view of spatiotemporal MAPK activity during the pheromone response, elucidating its role in mediating complex long-term developmental fates in a unicellular differentiation system.