RESUMO
Extracellular perception of auxin, an essential phytohormone in plants, has been debated for decades. Auxin-binding protein 1 (ABP1) physically interacts with quintessential transmembrane kinases (TMKs) and was proposed to act as an extracellular auxin receptor, but its role was disputed because abp1 knockout mutants lack obvious morphological phenotypes. Here, we identified two new auxin-binding proteins, ABL1 and ABL2, that are localized to the apoplast and directly interact with the extracellular domain of TMKs in an auxin-dependent manner. Furthermore, functionally redundant ABL1 and ABL2 genetically interact with TMKs and exhibit functions that overlap with those of ABP1 as well as being independent of ABP1. Importantly, the extracellular domain of TMK1 itself binds auxin and synergizes with either ABP1 or ABL1 in auxin binding. Thus, our findings discovered auxin receptors ABL1 and ABL2 having functions overlapping with but distinct from ABP1 and acting together with TMKs as co-receptors for extracellular auxin.
Assuntos
Arabidopsis , Ácidos Indolacéticos , Reguladores de Crescimento de Plantas , Ácidos Indolacéticos/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Arabidopsis/química , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismoRESUMO
Transmembrane kinases (TMKs) play important roles in plant growth and signaling cascades of phytohormones. However, its function in the regulation of early leaf senescence (ELS) of plants remains unknown. Here, we report the molecular cloning and functional characterization of the WATER-SOAKED SPOT1 gene which encodes a protein belongs to the TMK family and controls chloroplast development and leaf senescence in rice (Oryza sativa L.). The water-soaked spot1 (oswss1) mutant displays water-soaked spots which subsequently developed into necrotic symptoms at the tillering stage. Moreover, oswss1 exhibits slightly rolled leaves with irregular epidermal cells, decreased chlorophyll contents, and defective stomata and chloroplasts as compared with the wild type. Map-based cloning revealed that OsWSS1 encodes transmembrane kinase TMK1. Genetic complementary experiments verified that a Leu396Pro amino acid substitution, residing in the highly conserved region of leucine-rich repeat (LRR) domain, was responsible for the phenotypes of oswss1. OsWSS1 was constitutively expressed in all tissues and its encoded protein is localized to the plasma membrane. Mutation of OsWSS1 led to hyper-accumulation of reactive oxygen species (ROS), more severe DNA fragmentation, and cell death than that of the wild-type control. In addition, we found that the expression of senescence-associated genes (SAGs) was significantly higher, while the expression of genes associated with chloroplast development and photosynthesis was significantly downregulated in oswss1 as compared with the wild type. Taken together, our results demonstrated that OsWSS1, a member of TMKs, plays a vital role in the regulation of ROS homeostasis, chloroplast development, and leaf senescence in rice.
RESUMO
Transmembrane kinases (TMKs) are members of the plant receptor-like kinase (RLK) family. TMKs are characterized by an extracellular leucine-rich-repeat (LRR) domain, a single transmembrane region and a cytoplasmic kinase domain. TMKs have been shown to act as critical modulators of cell expansion and cell proliferation. Here, the crystal structure of the extracellular domain of TMK3 (TMK3-ECD) was determined to a resolution of 2.06â Å, with an Rwork of 17.69% and an Rfree of 20.58%. Similar to the extracellular domain of TMK1, the TMK3-ECD structure contains two solenoids with 13 LRRs and a non-LRR region (316-364) between the tenth and 11th LRRs. A comparison of TMK3-ECD with other LRR-RLKs that contain a non-LRR region indicates that the non-LRR region plays a critical role in structural integrity and may contribute to ligand interactions. The non-LRR region of TMK3-ECD is characterized by two disulfide bonds that may have critical biological implications.
Assuntos
Proteínas de Arabidopsis/química , Arabidopsis/química , Dissulfetos/química , Proteínas Serina-Treonina Quinases/química , Sequência de Aminoácidos , Arabidopsis/enzimologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Sítios de Ligação , Clonagem Molecular , Cristalografia por Raios X , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Modelos Moleculares , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia Estrutural de ProteínaRESUMO
Transmembrane kinases of Entamoeba histolytica are known to play a wide range of roles from virulence, phagocytosis, and proliferation to stress response. Transmembrane kinase EhTMKB1-9 is thought to be involved in early proliferative response and it was originally identified as a serum inducible gene. Ability to stimulate EhTMKB1 expression of serum starved cells resides in unsaturated fatty acids associated with albumin fraction of serum and the mechanism of stimulation follows activation of EhTMKB1-9 promoter. Gel shift assay showed the presence of proteins that bind to the specific site of EhTMKB1-9 upstream region and the concentration of these protein(s) go down on serum starvation, but level of binding protein(s) go up on serum or fatty acid replenishment. This increase in concentration of binding molecule(s) is due to new synthesis rather than activation of existing molecule(s) as a protein synthesis inhibitor blocked enhanced level of gel shifted material on replenishment. The stimulating activity resides in the fatty acyl chain, but not in the head group. Moreover, the fatty acid initiates signaling through class I PI3 kinases that result in activation of EhTMKB1-9 expression. These results suggest a novel mechanism of gene regulation in E. histolytica, and unsaturated fatty acids as potential new signaling molecules.