The RAF-like mitogen-activated protein kinase kinase kinases RAF22 and RAF28 are required for the regulation of embryogenesis in Arabidopsis.

In Arabidopsis, embryonic development follows a stereotypical pattern of cell division. Although many factors have been reported to participate in establishment of the proper embryonic pattern, the molecular mechanisms underlying pattern formation are unclear. In this study we showed that RAF22 and RAF28, two RAF-like mitogen-activated protein kinase kinase kinases (MAPKKKs) in Arabidopsis, are involved in the regulation of embryogenesis. The double knockout mutant of RAF22 and RAF28 was embryo lethal. A large proportion of the raf22-/- raf28+/- mutant embryos exhibited various defects, including disordered proembryo cell divisions, disruption of the bilaterally symmetrical structure, abnormally formative divisions of hypophysis and exaggerated suspensor growth. Whereas the kinase active form of RAF22 could complement these embryonic aberrant phenotypes, the kinase inactive form could not. The restrictive expression of the basal cell fate marker WOX8 in the abnormally dividing suspensor cells and the apical cell linage marker WOX2 in the abnormal proembryos indicated that apical and basal cell fates were unchanged in the abnormal embryos. The polar distribution of the auxin maxima and the PIN1 and PIN7 auxin transporters was markedly altered in the abnormal embryos. Our results suggest that RAF22 and RAF28 are important components of embryogenesis and that auxin polar transport may be involved in this regulation.

Arabidopsis thaliana Raf22 protein kinase maintains growth capacity during postgerminative growth arrest under stress.

When seeds are exposed to drought and salinity during germination, newly germinated embryos stop growth and enter a quiescent state called postgerminative growth (PGG) arrest. PGG arrest protects embryos from the stress, but it is not known how PGG is restored from the arrest when stress is eased. In this study, we show that under stress- or abscisic acid-induced PGG arrest conditions, Arabidopsis thaliana Raf-type protein kinase 22 (AtRaf22) overexpression accelerated photoautotrophic seedling establishment, whereas atraf22 knockout mutations enhanced the arrest. Furthermore, when the stress intensity was reduced subsequently, AtRaf22 overexpression plants resumed growth and accomplished photoautotrophic transition much faster than the knockout or wild-type plants. These results suggest that AtRaf22 activity is important for maintaining growth capacity during stress-induced PGG arrest, which is most likely critical for competitive growth when the stress subsides and plants resume growth. Such a factor has not been reported before.

RAF22, ABI1 and OST1 form a dynamic interactive network that optimizes plant growth and responses to drought stress in Arabidopsis.

Plants adapt to their ever-changing environment via positive and negative signals induced by environmental stimuli. Drought stress, for instance, induces accumulation of the plant hormone abscisic acid (ABA), triggering ABA signal transduction. However, the molecular mechanisms for switching between plant growth promotion and stress response remain poorly understood. Here we report that RAF (rapidly accelerated fibrosarcoma)-LIKE MITOGEN-ACTIVATED PROTEIN KINASE KINASE KINASE 22 (RAF22) in Arabidopsis thaliana physically interacts with ABA INSENSITIVE 1 (ABI1) and phosphorylates ABI1 at Ser416 residue to enhance its phosphatase activity. Interestingly, ABI1 can also enhance the activity of RAF22 through dephosphorylation, reciprocally inhibiting ABA signaling and promoting the maintenance of plant growth under normal conditions. Under drought stress, however, the ABA-activated OPEN STOMATA1 (OST1) phosphorylates the Ser81 residue of RAF22 and inhibits its kinase activity, restraining its enhancement of ABI1 activity. Taken together, our study reveals that RAF22, ABI1, and OST1 form a dynamic regulatory network that plays crucial roles in optimizing plant growth and environmental adaptation under drought stress.