Date of Award

2020

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Biology

First Advisor

Wagner Doris

Second Advisor

Brian Gregory

Abstract

A key question in multicellular eukaryote development is how master transcription factors can activate silenced genes during cell fate reprogramming in the context of chromatin. A special class of transcription factors, termed pioneer transcription factors, is ideally suited to execute this role because they can bind their target sites in nucleosomes. Although plants frequently reprogram cell fate in response to a changing environment, pioneer transcription factor activity has not been tested yet in this kingdom. Here we identify the master transcription factor LEAFY (LFY), which reprograms cells to flower fate, as a pioneer transcription factor. LFY binds in vitro in a sequence-specific manner and with high affinity to native target locus DNA assembled into a nucleosome. Genomic analyses and sequential chromatin immunoprecipitation reveal that LFY also binds nucleosome occupied target sites in vivo. Upon association with nucleosome enriched binding sites, LFY displaces the H1 linker histone and recruits SWI/SNF chromatin remodelers. The combined data suggest that the pioneer transcription factor LFY acts as a licensing factor for floral fate that facilitates binding of additional non pioneer transcription factors, chromatin opening and cell fate reprogramming.

We are also interested in understanding the physiochemical properties of LFY and wondered whether its unique structural property, a long central disordered domain in between two multivalent functional domains, could support the formation of membrane-less compartments in cells and offer an additional layer of regulation through phase separation. By transiently expressing various LFY proteins in protoplast systems, we observed cytoplasmic droplet formation of LFY and came to the conclusions that all domains in LFY are important for such behavior. Additionally, these droplets co-localize with processing bodies and co-expression of LFY co-regulator drove even bigger puncta. Our data suggested that droplet formation might be a regulatory mechanism to prevent nuclear entry of LFY when not needed and that proteasome mediated degradation might be involved when an exuberated amount is present.

Embargoed

Available to all on Friday, June 10, 2022

Files over 3MB may be slow to open. For best results, right-click and select "save as..."

Included in

Biology Commons

Share

COinS