RESUMO
BACKGROUND AND AIMS: CRABS CLAW (CRC) is a member of the YABBY family of transcription factors involved in carpel morphogenesis, floral determinacy and nectary specification in arabidopsis. CRC orthologues have been functionally characterized across angiosperms, revealing additional roles in leaf vascular development and carpel identity specification in Poaceae. These studies support an ancestral role of CRC orthologues in carpel development, while roles in vascular development and nectary specification appear to be derived. This study aimed to expand research on CRC functional conservation to the legume family in order to better understand the evolutionary history of CRC orthologues in angiosperms. METHODS: CRC orthologues from Pisum sativum and Medicago truncatula were identified. RNA in situ hybridization experiments determined the corresponding expression patterns throughout flower development. The phenotypic effects of reduced CRC activity were investigated in P. sativum using virus-induced gene silencing. KEY RESULTS: CRC orthologues from P. sativum and M. truncatula showed similar expression patterns, mainly restricted to carpels and nectaries. However, these expression patterns differed from those of other core eudicots, most importantly in a lack of abaxial expression in the carpel and in atypical expression associated with the medial vein of the ovary. CRC downregulation in pea caused defects in carpel fusion and style/stigma development, both typically associated with CRC function in eudicots, but also affected vascular development in the carpel. CONCLUSIONS: The data support the conserved roles of CRC orthologues in carpel fusion, style/stigma development and nectary development. In addition, an intriguing new aspect of CRC function in legumes was the unexpected role in vascular development, which could be shared by other species from widely diverged clades within the angiosperms, suggesting that this role could be ancestral rather than derived, as so far generally accepted.
Assuntos
Flores/genética , Regulação da Expressão Gênica de Plantas , Medicago truncatula/genética , Pisum sativum/genética , Proteínas de Plantas/metabolismo , Sequência de Aminoácidos , Arabidopsis/citologia , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/fisiologia , Evolução Molecular , Flores/crescimento & desenvolvimento , Flores/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Medicago truncatula/citologia , Medicago truncatula/crescimento & desenvolvimento , Medicago truncatula/fisiologia , Dados de Sequência Molecular , Pisum sativum/citologia , Pisum sativum/crescimento & desenvolvimento , Pisum sativum/fisiologia , Filogenia , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/fisiologia , Proteínas de Plantas/genética , Feixe Vascular de Plantas/genética , Feixe Vascular de Plantas/crescimento & desenvolvimento , Feixe Vascular de Plantas/fisiologia , Alinhamento de Sequência , Análise de Sequência de DNA , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Delivery of secretory immunoglobulin A (sIgA) to mucosal surfaces as a passive immunotherapy agent is a promising strategy to prevent infectious diseases. Recombinant sIgA production in plants requires the co-expression of four transcriptional units encoding the light chain (LC), heavy chain (HC), joining chain (JC) and secretory component (SC). As a way to optimize sIgA production in plants, we tested the combinatorial expression of 16 versions of a human sIgA against the VP8* rotavirus antigen in Nicotiana benthamiana, using the recently developed GoldenBraid multigene assembly system. Each sIgA version was obtained by combining one of the two types of HC (α1 and α2) with one of the two LC types (k and λ) and linking or not a KDEL peptide to the HC and/or SC. From the analysis of the anti-VP8* activity, it was concluded that those sIgA versions carrying HCα1 and LCλ provided the highest yields. Moreover, ER retention significantly increased antibody production, particularly when the KDEL signal was linked to the SC. Maximum expression levels of 32.5 µg IgA/g fresh weight (FW) were obtained in the best performing combination, with an estimated 33% of it in the form of a secretory complex.