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Eugenia Brun Hernandez

Role of a potential pioneer factor in the differentiation of plant stem cells

Published on 27 April 2018
Thesis presented April 27, 2018

LFY is a key transcription factor in plant development, and especially in flowering for angiosperms. It has an important role, first, in the establishment of floral meristems and later, in the specification of their floral organ identities. This activity implicates on cells’ nucleus major chromatin rearrangements. Target loci need to pass from a closed to an opened state. In the last two decades, Pioneer Transcription Factors (PTFs) have been studied because they can bind their target sites at nucleosomal DNA, they are able to overcome the steric constraints of nucleosomes and establish a “competent state” in a particular region, so it can be further regulated by other partners (Iwafuchi-Doi & Zaret, 2014). LFY has been demonstrated to physically and genetically interact with two ATPases belonging to ATP-dependent chromatin remodeling complexes, SYD and BRM (Wu et al., 2012). Besides, genome-wide data analyses strongly suggest that its N-terminal oligomerization domain, confers LFY access to closed regions of chromatin (Sayou et al., 2016). In this way, LFY presents common features with PTFs. We worked in order to better understand LFY’s mode of action in relation to the mentioned ATPases as well as with chromatin.
In Chapter I, through in vitro experiments, LFY’s potential interaction with nucleosomes, was assessed. We performed reconstituted nucleosomes by identifying nucleosome-enriched regions in Arabidopsis genome, efficiently targeted by LFY. These regions were selected used genome-wide data from ChIP-seq of LFY in overexpressing lines and DNAse-seq as well as MNase-seq data, which was useful to analyze chromatin landscape (T. Zhang, Zhang & Jiang, 2015; W. Zhang, Zhang, Wu & Jiang, 2012). A strong but non-specific binding of LFY from the gymnosperm Ginkgo biloba to nucleosomes was observed. However, LFY from Arabidopsis thaliana, showed a weak binding to nucleosomes.
In Chapter II, the aim was to map the minimal necessary interacting domains of LFY and the ATPases SYD and BRM. Using the HTRF technique, LFY’s C-terminal domain was shown to interact with BRM’s HSA domain. In addition, through an in vivo approach, we observed the loss of the 35S:LFY phenotype in the F1 plants from each of the three crosses: 35S:LFY syd-5, 35S:LFY brm-1 and 35S:LFY brm-3. This suggested a strong interaction, meaning that when BRM or SYD are not functional, LFY’s function is affected and no ectopic flowers are formed.

LEAFY, Pioneer factor, Nucleosome

On-line thesis.