Mechanistic insight into the MAP kinase signaling pathway during zygote polarization in Arabidopsis thaliana

Abstract

In Arabidopsis, the YDA pathway regulates stomata formation, inflorescence architecture and zygote polarity. Although many upstream components of the YDA pathway during stomata patterning have been revealed, upstream components of the embryonic YDA pathway are still largely unknown. In addition, how the YDA signaling confers zygote polarity is still elusive. In the first project I was involved in, we found that BSK1/2 function upstream of the YDA cascade in addition to SSP. The bsk1 bsk2 ssp triple mutant shows severe zygote polarity defects, recapitulating the yda phenotype. It has been suggested that the intramolecular interaction between the BSK kinase domain and TPR domain represses BSK activity. We confirmed this repressive interaction within BSK1. Intriguingly, SSP lost this interaction, which confers a constitutive activity. Further swapping experiments between BSK1 and SSP indicate that the first and second TPR motifs of SSP are responsible for its hyperactivity. Since BSKs and MAPK signals function downstream of receptors, we then wondered which receptor regulates the embryonic YDA pathway. I showed ER functions upstream of YDA with a sporophytic maternal effect. The functional ER mRNAs/proteins in the zygote are likely inherited pre-meiotically from the megaspore mother cell. The sporophytic maternal effect was also observed for BSK1. Furthermore, I confirmed the paternal regulator SSP can function in an ERf-independent manner. My results demonstrate that independent parental signal inputs converge on YDA activation to modulate zygote polarity, reminiscent of the parental conflict theory. To detect whether other receptors can also activate YDA, we collaborated with Prof. Dr. Michael Hothorn to design an approach for constitutive activation of SERK-dependent LRR-RK pathways. We fused the BIR3 ectodomain with the kinase domain of several LRR-RKs and confirmed their constitutive activity in the corresponding tissues. In particular, when expressed in the epidermal meristemoid, oBIR3-iER blocked stomata formation while oBIR3-iFLS2 had no apparent influence, suggesting the signal specificity of BIR3 chimeras. Then I used this approach to screen the HAESA family receptors and found that most receptors of this family can potentially activate YDA. In yda-CA transgenic lines, long filamentous embryos are formed. However, what identity these cells possess has not been studied. I designed a filamentous-embryo system to study the suspensor-embryo transition. Twin embryos are frequently developed from the early filamentous embryo. Then we proved that the early filamentous embryo has the identity of early basal cells. In addition, the maximum auxin response is shifted from the hypophysis to basal cells, which may directly contribute to the suspensor-embryo transition. We then investigate how the YDA signal confers zygote polarity. I showed that SSP is polarly localized in the basal plasma membrane of the elongating zygote. Depolarized localization of SSP leaded to a severe zygote polarity defect, indicating that its asymmetric localization is potent for zygote polarity. As SSP interacts directly with YDA and functions in an ERf-independent way, these results shed light on a polarized YDA activity in the zygote. These works collectively improved our understanding of the mechanism of zygote polarization.