A gas-and-brake mechanism of bHLH proteins modulates shade avoidance in Arabidopsis

Abstract

The shade avoidance syndrome (SAS) is typical of crops and wild plants grown at high densities. Common phenotypic traits of the SAS are petiole and hypocotyl elongation, upward movement of the leaves (hyponastic growth) and early flowering. This syndrome is often seen in crops, which are usually grown in high densities, and leads to losses in harvestable organs. Several attempts have been made to understand the molecular mechanisms of how SAS is controlled, ultimately with the aim to improve crop species for these traits. Most of the studies have been performed on the model species Arabidopsis thaliana. More recently, a different approach was taken, using two species of the Geranium genus that were selected for their antithetical response to shade: G. pyrenaicum is shade avoiding and can be found in open fields; G. robertianum is shade tolerant and is, amongst others, typical of forest understory. This alternative study had the aim to unravel the molecular mechanisms employed by shade tolerant species to suppress SAS. Through a transcriptome analysis a few candidate genes were selected and identified as new regulators of SAS. In this study we focused the attention to one of these candidates called KIDARI (KDR), an atypical basic helix-loop-helix (bHLH) protein. We studied the response of gain and loss of function lines of KDR of A. thaliana upon exposure to low R:FR treatment. We discovered that KDR acts as a positive regulator of SAS. We then identified novel interactors of KDR of A. thaliana. Two of the identified interactors of KDR were well established negative regulators of SAS, while others were not directly associated with shade avoidance responses but were characterized as negative regulators of cell elongation. We discovered also that all targets of KDR seem to be localized in the nucleus when transiently expressed in Nicotiana benthamiana leaves. On the other hand, KDR was localized in both the cytoplasm and the nucleus when transiently expressed in N. benthamiana. More interestingly, the localization of KDR was found to be primarily nuclear when coexpressed with the strong interactors, suggesting that the binding with partners leads to the translocation of KDR to the nucleus. In order to have a broader overview of the network of interactions in which KDR is involved, we identified interactor proteins of the newly identified KDR targets. Some of the interactors identified were already described as positive growth regulators and associated with shade avoidance. Other proteins were identified instead in other abiotic stresses but never in relation to shade responses and our findings showed that they can be potentially involved also in the regulation of cell elongation. The results presented in this thesis show that the response of plants to different stimuli is highly regulated by a complex network of interactions composed by bHLH transcription factors, allowing plants to tightly modulate the strength of their shade responses, for example relative to the intensity of signal inputs and other ecological parameters.