RESUMO
Despite the existence of considerable research on somatic embryogenesis (SE), the molecular mechanism that regulates the biosynthesis of auxins during the SE induction process remains unknown. Indole-3-acetic acid (IAA) is an auxin that is synthesized in plants through five pathways. The biosynthetic pathway most frequently used in this synthesis is the conversion of tryptophan to indol-3-pyruvic acid (IPA) by tryptophan aminotransferase of Arabidopsis (TAA) followed by the conversion of IPA to IAA by enzymes encoded by YUCCA (YUC) genes of the flavin monooxygenase family; however, it is unclear whether YUC-mediated IAA biosynthesis is involved in SE induction. In this study, we report that the increase of IAA observed during SE pre-treatment (plants in MS medium supplemented with 1-naphthaleneacetic acid (NAA) 0.54 µM and kinetin (Kin) 2.32 µM for 14 days) was due to its de novo biosynthesis. By qRT-PCR, we demonstrated that YUC gene expression was consistent with the free IAA signal found in the explants during the induction of SE. In addition, the use of yucasin to inhibit the activity of YUC enzymes reduced the signal of free IAA in the leaf explants and dramatically decreased the induction of SE. The exogenous addition of IAA restored the SE process in explants treated with yucasin. Our findings suggest that the biosynthesis and localization of IAA play an essential role during the induction process of SE in Coffea canephora.
Assuntos
Coffea/embriologia , Coffea/metabolismo , Ácidos Indolacéticos/metabolismo , Reguladores de Crescimento de Plantas/biossíntese , Vias Biossintéticas/efeitos dos fármacos , Vias Biossintéticas/genética , Coffea/genética , Inibidores Enzimáticos/farmacologia , Perfilação da Expressão Gênica , Genes de Plantas , Oxigenases de Função Mista/antagonistas & inibidores , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Família Multigênica , Reguladores de Crescimento de Plantas/genética , Proteínas de Plantas/antagonistas & inibidores , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Técnicas de Embriogênese Somática de Plantas , Triazóis/farmacologiaRESUMO
Auxin and polar auxin transport have been implicated in controlling zygotic embryo development, but less is known about their role in the development of somatic embryos. The aim of this study was to determine if indole-3-acetic acid (IAA) and the PIN1 transporter participate in the induction of somatic embryogenesis (SE) and the development of somatic embryos. The results show that IAA levels gradually increase during pre-treatment and accumulate in the chloroplast. During pre-treatment and the globular stage of SE in C. canephora, auxin is distributed uniformly in all of the cells of the somatic embryo. During the subsequent stages of development, auxins are mobilized to the cells that will form the cotyledons and the root meristem. The location of the PIN transporters shifts from the plasmalemma of the protoderm cells during the globular stage to the plasmalemma of the cells that will give rise to the cotyledons and the vascular tissue in the late stages of somatic embryogenesis. The incubation of the explants in the presence of 2,3,5-triiodobenzoic acid (TIBA) produced aberrant somatic embryos, suggesting that PIN1 mediates the transport of IAA.