RESUMO
Olfaction plays a fundamental role in insect survival through resource location and intra and interspecific communications. We used RNA-Seq to analyze transcriptomes for odorant-binding proteins (OBPs) from major stink bug pest species in Brazil, Euschistus heros, Chinavia ubica, and Dichelops melacanthus, and from their egg parasitoid, Telenomus podisi. We identified 23 OBPs in E. heros, 25 OBPs in C. ubica, 9 OBPs in D. melacanthus, and 7 OBPs in T. podisi. The deduced amino acid sequences of the full-length OBPs had low intraspecific similarity, but very high similarity between two pairs of OBPs from E. heros and C. ubica (76.4 and 84.0%) and between two pairs of OBPs from the parasitoid and its preferred host E. heros (82.4 and 88.5%), confirmed by a high similarity of their predicted tertiary structures. The similar pairs of OBPs from E. heros and C. ubica may suggest that they have derived from a common ancestor, and retain the same biological function to bind a ligand perceived or produced in both species. The T. podisi OBPs similar to E. heros were not orthologous to any known hymenopteran OBPs, and may have evolved independently and converged to the host OBPs, providing a possible basis for the host location of T. podisi using E. heros semiochemical cues.
Assuntos
Heterópteros/genética , Óvulo/parasitologia , Parasitos/genética , Receptores Odorantes/metabolismo , Transcriptoma/genética , Clima Tropical , Sequência de Aminoácidos , Animais , Simulação por Computador , Biblioteca Gênica , Ontologia Genética , Interações Hospedeiro-Parasita/genética , Modelos Moleculares , Anotação de Sequência Molecular , Dados de Sequência Molecular , Filogenia , Estrutura Terciária de Proteína , Receptores Odorantes/química , Receptores Odorantes/genética , Alinhamento de Sequência , Análise de Sequência de RNA , Especificidade da EspécieRESUMO
An actual severe problem in agriculture consists of an expressive increase of economical losses caused by fungi and resistant bacteria toward antibiotics. In order to find a solution to this problem, several studies have been concentrating on the screening of novel plant defense peptides with antimicrobial activities. These peptides are commonly characterized by having low molecular masses and cationic charges. The present work reports the purification and characterization of a novel plant peptide with molecular mass of 5340 Da, named Cp-AMP, from seeds of C. pallida, a typical plant from Caatinga biome. Purification was achieved using a size exclusion S-200 column followed by reversed-phase chromatography on Vydac C18-TP column. In vitro assays indicated that Cp-AMP was able to inhibit the development of filamentous fungi Fusarium oxysporum as well as the gram-negative bacterium Proteus sp. The identification of Cp-AMP could contribute, in the near future, to the development of biotechnological products, such as transgenic plants with enhanced resistance to pathogenic fungi and/or of antibiotics production derived from plant sources in order to control bacterial infections.
Assuntos
Peptídeos Catiônicos Antimicrobianos/isolamento & purificação , Peptídeos Catiônicos Antimicrobianos/farmacologia , Crotalaria/química , Fusarium/efeitos dos fármacos , Proteus/efeitos dos fármacos , Sementes/química , Peptídeos Catiônicos Antimicrobianos/química , Cromatografia Líquida/métodos , Peso Molecular , Proteínas de Plantas/química , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/farmacologiaRESUMO
Cowpea seeds (Vigna ungiculata) are widely cultivated by poor farmers in Latin America and Africa and are often severely damaged by the cowpea weevil Callosobruchus maculatus. A proteinaceous inhibitor of cowpea weevil digestive enzymes, PpAI, was purified from white sucupira seeds (Pterodon pubescens) and biochemically characterized in this study. Proteins were extracted from seeds and precipitated with ammonium sulfate at 100% saturation. This fraction was applied onto a Red-sepharose CL-6B column, and the retained peak showed 70% inhibitory activity toward larval C. maculatus digestive alpha-amylases. The retained peak was then purified using an analytical reversed-phase HPLC column. Purified PpAI showed 65% inhibitory activity against larval C. maculatus enzymes. Enzymatic assays also showed that the purified P. pubescens inhibitor was unable to reduce the activity of mammalian alpha-amylases, suggesting specificity toward insect enzymes. Moreover, artificial seeds containing PpAI were able to reduce larval weight by 36% and cause 55% mortality. Mass spectrometry and SDS-PAGE analyses indicated that PpAI showed a molecular mass of approximately 5.0 kDa. This alpha-amylase inhibitor, coming from a native Cerrado plant, could be used to construct a genetically engineered cowpea with enhanced resistance against weevil pests.