Suspension cell culture as a tool for the characterization of class III peroxidases in sugarcane

Autor(es)

Cesarino, Igor ^[1] ; Araujo, Pedro ^[1] ; Paes Leme, Adriana Franco ^[2] ; Creste, Silvana ^[3] ; Mazzafera, Paulo ^[1]

Afiliação do(s) autor(es)

^[1] Univ Estadual Campinas, Inst Biol, Dept Biol Vegetal, BR-13083970 Campinas, SP - Brazil

^[2] Lab Nacl Biociencias CNPEM ABTLuS, Campinas, SP - Brazil

^[3] Inst Agron Campinas, Ctr Cana, BR-14001970 Ribeirao Preto, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: Plant Physiology and Biochemistry; v. 62, p. 1-10, JAN 2013.
• Citações Web of Science: 19

Resumo

Secreted class III peroxidases (EC 1.11.1.7) are implicated in a broad range of physiological processes throughout the plant life cycle. However, the unambiguous determination of the precise biological role of an individual class III peroxidase isoenzyme is still a difficult task due to genetic redundancy and broad substrate specificity in vitro. In addition, many difficulties are encountered during extraction and analysis of cell wall proteins. Since class III peroxidases are also secreted into the apoplast, the use of suspension cell cultures can facilitate isolation and functional characterization of individual isoforms. Here, we report on the characterization of class III peroxidases secreted in the spent medium of sugarcane suspension cell cultures. After treatment with specific inducers of cell wall lignification, peroxidases were isolated and activities assayed with guaiacol, syringaldazine and coniferyl alcohol. Enzymatic activity was not significantly different after treatments, regardless of the substrate, with the exception of methyl-jasmonate treatment, which led to a decreased guaiacol peroxidase activity. Remarkably, peroxidases isolated from the medium were capable of oxidizing syringaldazine, an analog to sinapyl alcohol, suggesting that sugarcane cultures can produce peroxidases putatively correlated to lignification. A proteomic approach using activity staining of 2-DE gels revealed a complex isoperoxidase profile, composed predominantly of cationic isoforms. Individual spots were excised and analyzed by LC-ESI-Q-TOF and homology-based search against the Sugarcane EST Database resulted in the identification of several proteins. Spatio-temporal expression pattern of selected genes was determined for validation of identified class III peroxidases that were preferentially expressed during sugarcane stem development. (C) 2012 Elsevier Masson SAS. All rights reserved. (AU)

• Processo FAPESP: 08/58035-6 - Control of lignin biosynthesis in sugar cane: many gaps still to be filled
• Beneficiário: Paulo Mazzafera
• Linha de fomento: Auxílio à Pesquisa - Programa BIOEN - Temático