title: Mlo, a modulator of plant defense and cell death, is a novel calmodulin-binding protein. Isolation and characterization of a rice Mlo homologue.
abstract: Transient influx of Ca(2+) constitutes an early event in the signaling cascades that trigger plant defense responses. However, the downstream components of defense-associated Ca(2+) signaling are largely unknown. Because Ca(2+) signals are mediated by Ca(2+)-binding proteins, including calmodulin (CaM), identification and characterization of CaM-binding proteins elicited by pathogens should provide insights into the mechanism by which Ca(2+) regulates defense responses. In this study, we isolated a gene encoding rice Mlo (Oryza sativa Mlo; OsMlo) using a protein-protein interaction-based screening of a cDNA expression library constructed from pathogen-elicited rice suspension cells. OsMlo has a molecular mass of 62 kDa and shares 65% sequence identity and scaffold topology with barley Mlo, a heptahelical transmembrane protein known to function as a negative regulator of broad spectrum disease resistance and leaf cell death. By using gel overlay assays, we showed that OsMlo produced in Escherichia coli binds to soybean CaM isoform-1 (SCaM-1) in a Ca(2+)-dependent manner. We located a 20-amino acid CaM-binding domain (CaMBD) in the OsMlo C-terminal cytoplasmic tail that is necessary and sufficient for Ca(2+)-dependent CaM complex formation. Specific binding of the conserved CaMBD to CaM was corroborated by site-directed mutagenesis, a gel mobility shift assay, and a competition assay with a Ca(2+)/CaM-dependent enzyme. Expression of OsMlo was strongly induced by a fungal pathogen and by plant defense signaling molecules. We propose that binding of Ca(2+)-loaded CaM to the C-terminal tail may be a common feature of Mlo proteins.
authors.affiliation: Division of Applied Life Science, Plant Molecular Biology and Biotechnology Research Center, Gyeongsang National University, Chinju 660-701, Korea.
authors.forename: Min Chul
authors.initials: MC
authors.lastname: Kim
authors.forename: Sang Hyoung
authors.initials: SH
authors.lastname: Lee
authors.forename: Jong Kyong
authors.initials: JK
authors.lastname: Kim
authors.forename: Hyun Jin
authors.initials: HJ
authors.lastname: Chun
authors.forename: Man Soo
authors.initials: MS
authors.lastname: Choi
authors.forename: Woo Sik
authors.initials: WS
authors.lastname: Chung
authors.forename: Byeong Cheol
authors.initials: BC
authors.lastname: Moon
authors.forename: Chang Ho
authors.initials: CH
authors.lastname: Kang
authors.forename: Chan Young
authors.initials: CY
authors.lastname: Park
authors.forename: Jae Hyuk
authors.initials: JH
authors.lastname: Yoo
authors.forename: Yun Hwan
authors.initials: YH
authors.lastname: Kang
authors.forename: Seong Cheol
authors.initials: SC
authors.lastname: Koo
authors.forename: Yoon Duck
authors.initials: YD
authors.lastname: Koo
authors.forename: Jae Cheol
authors.initials: JC
authors.lastname: Jung
authors.forename: Sun Tae
authors.initials: ST
authors.lastname: Kim
authors.forename: Paul
authors.initials: P
authors.lastname: Schulze-Lefert
authors.forename: Sang Yeol
authors.initials: SY
authors.lastname: Lee
authors.forename: Moo Je
authors.initials: MJ
authors.lastname: Cho
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001667:Binding, Competitive; D015152:Blotting, Northern; D015139:Blotting, Southern; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000737:chemistry / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D003001:Cloning, Molecular; D003593:Cytoplasm / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D015723:Gene Library; D017343:Genes, Plant; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D012275:Oryza / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D010802:Phylogeny; D010940:Plant Proteins / Q000378:metabolism* / Q000502:physiology*; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D013347:Subcellular Fractions / Q000378:metabolism; D013997:Time Factors
chemical_list: D002147:Calmodulin; D018076:DNA, Complementary; C104922:MLO protein, Hordeum vulgare; D010455:Peptides; D010940:Plant Proteins; D020033:Protein Isoforms; D011994:Recombinant Proteins; D010727:Phosphoric Diester Hydrolases; D002118:Calcium