The first MAPK identified from rice is the OsBWMK1 (where Os denotes Oryza sativa), whose expression is induced upon infection by blast fungus Magnaporthe grisea and mechanical wounding (He et al., 1999). This study
MAPKs in Plant Defence Responses 49 provides the first evidence for the existence of a MAPK cascade component in rice and the possible involvement in rice defence mechanism(s). With the completion of the rice genome sequencing project, extensive bioinformatics analyses identified 17 MPKs and eight MKKs in rice (Hamel et al., 2006;
Reyna and Yang, 2006). Compared with the significant progress on the biological function of the MAPK cascades in Arabidopsis, little is known about the function and regulation of MAPKs in rice. Up to date, only a few of them have been characterized in detail for their biological function in rice growth, development and response to environmental cues.
Identification of MAPKs and expression patterns in defence responses
Phylogenetic analysis and pairwise comparison of Arabidopsis and rice MAPKs revealed that 11 rice MAPKs contain the TDY activation motif and six MAPKs contains the TEY motif. This is different from those in Arabidopsis, which contains more MAPKs with the TEY motif than the MAPKs with the TDY motif. Genome-wide expression analyses by qRT-PCR indicate that, upon inoculation with M. grisea, nine of 17 OsMPK genes are induced at the mRNA level during either early, late, or both stages of infection. Four of the M. grisea-induced OsMPK genes are associated with host-cell death in the lesion-mimic rice mutant, and eight of them are differentially induced in response to defence signal molecules such as JA, SA and ET (Reyna and Yang, 2006).
OsMPK12 (OsBWMK1) is induced by both virulent and avirulent isolates of M. grisea and by treatments with SA, JA and 1-aminocyclopropane-1-carboxylic acid (ACC) within 24 h after inoculation/treatment (He et al., 1999;
Cheong et al., 2003; Reyna and Yang, 2006). The OsMPK12 protein levels do not change in rice leaf tissues after treatments with different defence signalling molecules, indicating that the OsMPK12 activation is primarily achieved by post-translational modification (Cheong et al., 2003). The OsMPK12 gene has three alternatively spliced transcripts, OsBWMK1L, OsBWMK1M and OsBWMK1S, but only the OsBWMK1 transcripts are differentially expressed in tissues and under various stress conditions. Alternative splicing of OsBWMK1 generates three different transcript variants that produce proteins with different subcellular localizations (Koo et al., 2007). OsMPK5 (also known as OsMSRMK2, OsMAPK2, OsBIMK1 or OsMAP1) is induced by various biotic and abiotic stresses, its induction by M. grisea and some defence signalling molecules including SA, JA and ACC is well documented by several research groups (Agrawal et al., 2002; Song and Goodman, 2002;
Xiong and Yang, 2003; Reyna and Yang, 2006). The OsMPK5 gene generates at least two alternatively spliced transcripts, OsMPK5a and OsMPK5b. Kinase activity assays revealed that only OsMAPK5a exhibits autophosphorylation activity in vitro (Xiong and Yang, 2003). OsMAPK5 kinase activity is induced significantly by blast fungus infection and the early transient activation of OsMAPK5 activity probably is related to the resistance response to avirulent blast isolates (Xiong and Yang, 2003). OsMPK13 (also known as OsBIMK2) is induced within 48 h during an incompatible interaction between rice and M.
50 F. Song et al.
grisea, and by treatment with SA, benzothiadiazole (BTH) and ACC, but not by treatment with JA (Reyna and Yang, 2006; Song et al., 2006). In vitro kinase assay revealed that OsBIMK2 has an autophosphorylation activity (Song et al., 2006). OsMPK1 (also known as OsMAPK6) was reported to be post-translationally activated by a sphingolipid elicitor and regulated by the small GTPase OsRac1 and heterotrimeric G-protein (Lieberherr et al., 2005).
Genome-wide analyses also revealed that six other rice MAPK genes, OsMPK2, OsMPK4, OsMPK7, OsMPK8, OsMPK15 and OsMPK17, are induced by M.
grisea and defence signalling molecules, suggesting that they may also play roles in defence signal transduction (Reyna and Yang, 2006).
Very little is known about MKKs and MEKKs in biotic defence response in rice. OsMKK1 is induced in rice leaves after infection by M. grisea, feeding by insect (Nilaparvata lugens) and treatment with SA, JA and ET (You et al., 2007). OsEDR1, an orthologue of Arabidopsis AtEDR1, has a constitutive expression in seedling leaves and is further upregulated by treatment with JA, SA and ET (Kim, J.A. et al., 2003). These preliminary observations may pro-vide starting points to investigate defence-related MAPK cascades in rice disease resistance.
However, the involvement of most of the rice MAPKs in disease resistance was established mainly based on their inducible expression patterns in response to infection by M. grisea and to treatments with defence signalling molecules.
Further biochemical, genetics and functional analyses are required to clarify the biological functions and mechanisms of MAPK cascades in rice disease resistance.
Functions of MAPKs in rice disease resistance
Although a number of MAPKs have been implicated in rice defence responses, only a few of them have been characterized for their biological functions in rice disease resistance through functional genomics approaches. Suppression of OsMPK5 expression and its kinase activity results in the constitutive expression of defence genes in dsRNAi transgenic plants and significantly enhances resistance to fungal (M. grisea) and bacterial (Burkholderia glumae) pathogens.
However, these same dsRNAi lines have significant reductions in drought, salt and cold tolerance. By contrast, OsMPK5-overexpressing plants exhibit increased kinase activity and increased tolerance to drought, salt and cold stresses. Based on these findings, it was proposed that OsMAPK5 negatively modulates defence gene expression and broad-spectrum disease resistance and positively regulates abiotic stress tolerance (Xiong and Yang, 2003). It is likely that suppressing or knocking out of OsMPK6, an orthologue of Arabidopsis AtMPK4, enhances resistance to different races of Xanthomonas oryzae pv.
oryzae, causing bacterial blight disease. The OsMPK6-suppressed plants show increased expression of a subset of defence-responsive genes functioning in the NH1 (an Arabidopsis NPR1 orthologue)-involved defence signal transduc-tion pathway. These findings support that OsMPK6 functransduc-tions as a repressor to regulate rice defence responses upon bacterial infection (Yuan et al., 2007).
MAPKs in Plant Defence Responses 51 Ectopic expression in tobacco was also used to study the function of some rice MAPK genes in disease resistance responses. Overexpression of the OSBIMK2 gene in transgenic tobacco results in enhanced disease resistance against tomato mosaic virus and Alternaria alternata (Song et al., 2006).
OsMPK12 (OsBWMK1) interacts with and phosphorylates OsEREBP1, a transcription factor belonging to the ethylene-responsive transcriptional factor family. Phosphorylation of OsEREBP1 by OsMPK12 enhances its in vitro DNA-binding activity to the GCC box element (AGCCGCC) in promoters of several defence genes. Transgenic tobacco plants overexpressing OsMPK12 display increased expression of many defence genes, induced HR-like cell death and enhanced disease resistance against H. parasitica var. nicotianae and P. syringae pv. tabacci. Therefore, it is likely that OsMPK12 contributes to plant defence signal transduction by phosphorylating one or more tran-scription factors (Cheong et al., 2003). This is similar to the observations that, in tobacco and Arabidopsis, MAPK cascades regulate defence responses by phosphorylating downstream WRKY transcription factors. Together, these find ings support the idea that MAPK cascades regulate transcription of a variety of stress responsive genes through modulation of corresponding tran-scription factors, although the target trantran-scription factors might be unique to different plant species.