Expression and functional analyses of the mitogen-activated protein kinase (MPK) cascade genes in response to phytohormones in wheat ( Triticum aestivum L.)

Mitogen-activated protein kinase （MPK） cascades consist of a set of kinase types （MPKKKs, MPKKs, MPKs） to establish conserved signal-transducing modules mediating plant growth, development as well as responses to internal and external cues. In this study, the expression patterns of six MPKKK, two MPKK, and 11 MPK genes in wheat in responses to external treatments of phytohormones, including naphthylacetic acid （NAA）, abscisic acid （ABA）, 6-benzyladenine （6-BA）, gibber- ellin （GA3）, salisylic acid （SA）, jasmonic acid （JA）, and ethylene （ETH）, were investigated. Expression analysis revealed that several of the MPK cascade genes are responses to the external phytohormone signaling. Of which, TaMPKKKA;3 is induced by 6-BA and NAA while TaMPK4 repressed by ETH, GA3, SA, and JA; TaMPKKKA, TaMPKKKA;3 and TaMPK1 are down-regulated by ETH and GA3whereas TaMPK9 and TaMPK12 repressed by ETH and JA in addition that TaMPK12 also repressed by GA3; TaMPK12;1 is down-regulated by ABA, GA3 and SA while TaMPK17 repressed by all exogenous phytonormones examined. TaMPK4, a MPK type gene previously characterized to mediate tolerance to phosphate （Pi） deprivation, was functionally evaluated for its role in mediation of responses of plants to exogenous GA3, ETH, SA, and JA. Results indicated that overexpression and antisense expression of TaMPK4 in tobacco dramatically modify the growth of seedlings upon treatments of GA3, SA and JA, in which the overexpressors behaved deteriorated growth feature whereas the seedlings with antisense expression of TaMPK4 exhibited improved seedling phenotype. The growth behaviors in lines overexpressing or antisensely expressing TaMPK4 are closely associated with the biomass and the corresponding hormone-associated parameters. These results demonstrated that TaMPK4 acts as a critical player in mediating the phyto- hormone signaling. Our findings have identified the phytohormone-responsive MPK cascade genes in wheat and provided a connection between the phytohormone-mediated responses and the MPK cascade pathways.

Ethylene-induced stomatal closure is mediated via MKK1/3–MPK3/6 cascade to EIN2 and EIN3

Mitogen-activated protein kinases(MPKs)play essential roles in guard cell signaling,but whether MPK cascades participate in guard cell ethylene signaling and interact with hydrogen peroxide(H2O2),nitric oxide(NO),and ethylene-signaling components remain unclear.Here,we report that ethylene activated MPK3 and MPK6 in the leaves of wild-type Arabidopsis thaliana as well as ethylene insensitive2(ein2),ein3,nitrate reductase1(nia1),and nia2 mutants,but this effect was impaired in ethylene response1(etr1),nicotinamide adenine dinucleotide phosphate oxidase AtrbohF,mpk kinase1(mkk1),and mkk3 mutants.By contrast,the constitutive triple response1(ctr1)mutant had constitutively active MPK3 and MPK6.Yeast two-hybrid,bimolecular fluorescence complementation,and pull-down assays indicated that MPK3 and MPK6 physically interacted with MKK1,MKK3,and the C-terminal region of EIN2(EIN2 CEND).mkk1,mkk3,mpk3,and mpk6 mutants had typical levels of ethylene-induced H2O2 generation but impaired ethylene-induced EIN2 CEND cleavage and nuclear translocation,EIN3 protein accumulation,NO production in guard cells,and stomatal closure.These results show that the MKK1/3–MPK3/6 cascade mediates ethylene-induced stomatal closure by functioning downstream of ETR1,CTR1,and H2O2 to interact with EIN2,thereby promoting EIN3 accumulation and EIN3-dependent NO production in guard cells.