Auxin Immunolocalization in Coffea canephora Tissues.

Auxins are plant growth regulators that participate in a variety of biological mechanisms during the growth and development of plants. The most abundant natural auxin is indole-3-acetic acid (IAA). The physiological processes regulated by IAA depend on their temporal space accumulation in different tissues of a plant. This accumulation is regulated by its biosynthesis, conjugation, degradation, and transport. Therefore tools that allow us a qualitative and quantitative detection of IAA in plant tissues are very useful to understand the homeostasis of IAA during the life cycle of plants. In this protocol, the complete procedure for localization of IAA in different tissues of Coffea canephora is described using specific anti-IAA monoclonal antibodies.

Localization and transport of indole-3-acetic acid during somatic embryogenesis in Coffea canephora.

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.

Regulation of polar auxin transport in grapevine fruitlets (Vitis vinifera L.) and the proposed role of auxin homeostasis during fruit abscission.

BACKGROUND: Indole-3-acetic acid (IAA), the most abundant auxin, is a growth promoter hormone involved in several developmental processes. Auxin homeostasis is very important to its function and this is achieved through the regulation of IAA biosynthesis, conjugation, degradation and transport. In grapevine, IAA plays an essential role during initial stages of berry development, since it delays fruitlet abscission by reducing the ethylene sensitivity in the abscission zone. For this reason, Continuous polar IAA transport to the pedicel is required. This kind of transport is controlled by IAA, which regulates its own movement by modifying the expression and localization of PIN-FORMED (PIN) auxin efflux facilitators that localize asymmetrically within the cell. On the other hand, the hormone gibberellin (GA) also activates the polar auxin transport by increasing PIN stability. In Vitis vinifera, fruitlet abscission occurs during the first two to three weeks after flowering. During this time, IAA and GA are present, however the role of these hormones in the control of polar auxin transport is unknown. RESULTS: In this work, the use of radiolabeled IAA showed that auxin is basipetally transported during grapevine fruitlet abscission. This observation was further supported by immunolocalization of putative VvPIN proteins that display a basipetal distribution in pericarp cells. Polar auxin transport and transcripts of four putative VvPIN genes decreased in conjunction with increased abscission, and the inhibition of polar auxin transport resulted in fruit drop. GA3 and IAA treatments reduced polar auxin transport, but only GA3 treatment decreased VvPIN transcript abundance. When GA biosynthesis was blocked, IAA was capable to increase polar auxin transport, suggesting that its effect depends on GA content. Finally, we observed significant changes in the content of several IAA-related compounds during the abscission period. CONCLUSIONS: These results provide evidence that auxin homeostasis plays a central role during grapevine initial fruit development and that GA and IAA controls auxin homeostasis by reducing polar auxin transport.