The identity of plant cells is remarkably fluid. This property is dramatically demonstrated by the ability of an excised piece of differentiated plant tissue to regenerate into an entire plant. We are interested in characterizing the factors and mechanisms that govern plant cell identity and facilitate its flexible nature. To accomplish this goal, we are utilizing a mutant of Arabidopsis thaliana, pickle (pkl), in which the primary root differentiates improperly and expresses embryonic characteristics after germination. pkl primary roots that express embryonic differentiation characteristics are called pickle roots based on their appearance. Expression of the pickle root phenotype is dependent on gibberellin (GA), a plant growth regulator known to promote such diverse processes as germination, cell elongation, and initiation of flowering.
We have cloned PKL, and it codes for a predicted protein that is a new member of the CHD family. The CHD proteins derive their name from the possession of three domains: a chromo (chromatin organization modifier) domain, a SNF2-related helicase/ATPase domain, and a DNA-binding domain. CHD genes are well conserved throughout eukaryotes. Some CHD family members, referred to as CHD3 or mi-2 proteins, contain an additional domain of sequence homology: the PHD zinc finger. PKL contains this domain and thus is also a CHD3 protein. CHD3 proteins from Xenopus and human cell lines have been shown to associate with histone deacetylase indicating that they may act as negative regulators of transcription. Consistent with such a role for PKL in Arabidopsis, we have shown that LEC1, a promoter of embryonic identity, is derepressed in germinating pkl seedlings.
The cloning of the PKL gene provides a unique opportunity to study the intersection of the diverse areas of GA signal transduction, developmental identity, and chromatin structure. Based on the phenotype of the pkl mutant and the function of proteins that are similar to PKL, our working model is that PKL regulates the transcription of genes in response to GA. Specifically, we propose that PKL establishes transcriptional repression of embryonic genes during germination by altering the structure of chromatin. This model will be tested by the following experimental strategies: examination of the expression of PKL, characterization of the DNA-binding site of PKL, identification of genes that exhibit PKL-dependent transcription, identification of proteins that interact with PKL, and genetic screens for mutations that affect the phenotype of pkl plants.
The intent of the proposed experiments is to elucidate the role of PKL and to identify factors and mechanisms that govern GA signal transduction and developmental transitions in plants. Insight gained from these studies may be useful in increasing agricultural productivity and/or in the generation of new crops. The observation that CHD proteins are conserved in eukaryotes suggests that the results obtained in our studies of PKL function in Arabidopsis may be generally applicable to eukaryotes. Perhaps plants and animals utilize common regulatory machinery to govern developmental transitions. If so, the proposed experiments with PKL in Arabidopsis may shed light on one of the factors involved in such transitions.
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