Abstract
The mitogen-activated protein kinase (MAPK) LjMPK6 is a phosphorylation target of SIP2, a MAPK kinase that interacts with SymRK (symbiosis receptor-like kinase) for regulation of legume-rhizobia symbiosis. Both LjMPK6 and SIP2 are required for nodulation in Lotus japonicus. However, the dephosphorylation of LjMPK6 and its regulatory components in nodule development remains unexplored. By yeast two-hybrid screening, we identified a type 2C protein phosphatase, LjPP2C, that specifically interacts with and dephosphorylates LjMPK6 in vitro. Physiological and biochemical assays further suggested that LjPP2C phosphatase is required for dephosphorylation of LjMPK6 in vivo and for fine-tuning nodule development after rhizobial inoculation. A non-phosphorylatable mutant variant LjMPK6 (T224A Y226F) could mimic LjPP2C functioning in MAPK dephosphorylation required for nodule development in hairy root transformed plants. Collectively, our study demonstrates that interaction with LjPP2C phosphatase is required for dephosphorylation of LjMPK6 to fine tune nodule development in L. japonicus.
Highlights
Root nodule symbiosis (RNS) between legumes and rhizobia allows conversion of atmospheric nitrogen into ammonia absorbed by plants
To study proteins interacting with the mitogen-activated protein kinase (MAPK) kinase LjMPK6, we identified several protein candidates from a Lotus cDNA library by yeast two-hybrid (Y2H) screening
SymRK-interacting protein SIP2 is required for early symbiotic signal transduction and MAPK signaling during nodule symbiosis [24]
Summary
Root nodule symbiosis (RNS) between legumes and rhizobia allows conversion of atmospheric nitrogen into ammonia absorbed by plants. Host plants secrete flavonoids, inducing rhizobia to synthesize and secrete Nod factors (NFs), a type of lipo-chitooligosaccharide molecules [1,2]. Two plant LysM-type serine/threonine receptor kinases, NFR1 and NFR5 in Lotus japonicus (Nod Factor Receptor 1 and 5) [3,4,5], cooperating with the leucine-rich repeat (LRR) receptor-like kinase SymRK (Symbiosis Receptor-like Kinase) [6], recognize NFs and initiate the NF signaling pathway [7,8,9,10]. The core component of the NF signaling pathway is a calcium oscillation which is decoded by the nucleus-localized CCaMK (calcium/calmodulin-dependent protein kinase) [11,12]. CCaMK phosphorylates and activates CYCLOPS [13,14], which binds to and activates the nodule inception (NIN) gene essential for rhizobial colonization, nodule organogenesis and ultimate nitrogen fixation in mature nodules [15,16]
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