Functional analyses of AtCHR12 and AtCHR23 : plant growth responses upon over-expression of chromatin remodeling ATPase genes

A. Folta

Research output: Thesisinternal PhD, WU

Abstract

Living organisms have to deal with changing environmental conditions during their whole life cycle. In contrast to animals, plants are sessile organisms. Therefore they have evolved multiple regulatory mechanisms that help them to cope with changing conditions. One of the first responses to stress conditions is reduction or arrest of growth. Therefore regulation of growth and development is essential to successfully complete their life cycle. To correctly time their development, plants need to integrate various environmental signals with intrinsic developmental programs. In this integration, regulation of gene expression plays a major role.

The genetic information of an organism is stored in DNA sequence. DNA forms a complex with histones and other proteins, which is called chromatin. Chromatin is a highly dynamic complex and modification of the chromatin structure makes DNA more or less accessible to the transcriptional machinery and other regulatory proteins. The modification of chromatin organization is called chromatin remodeling and it involves both covalent modifications of DNA and histone tails, and non-covalent modification of chromatin structure by ATP-dependent chromatin remodeling complexes. ATP-dependent chromatin remodeling complexes comprise multiple protein subunits with SNF2 ATPase as a catalytic subunit. Depending on the subunit composition, the complexes can perform different tasks at various places of chromatin, and can be active at different developmental stages (Chapter 1).

The SNF2 ATPases are conserved from yeast to plants. In Arabidopsis, 41 SNF2 ATPases have been identified. The focus of this thesis is on two of those ATPases – AtCHR12 and AtCHR23. It has been shown previously that AtCHR12 is involved in growth responses to environmental cues. We have extended these studies to its paralog AtCHR23 (Chapter 2). In contrast to over-expression of AtCHR12, which affects growth only during reproductive stage of development, over-expression of AtCHR23 leads to smaller seedlings and reduced vegetative growth. Upon application of mild abiotic stress, the growth reduction is stronger than in wild-type plants. Moreover, the transgenic plants manifest increased variability of growth. The increased growth variability correlates with increased expression variability of genes associated with stress. The results indicate that accurate and controlled expression of AtCHR23 is required for stability and robustness of growth, as well as gene expression.

Regulation of growth is important not only during vegetative or reproductive stage of development, but also during embryo development. The growth of the embryo is interrupted during the embryo maturation phase and it was suggested that AtCHR12 may be involved in this temporary growth arrest. Here we have shown that both AtCHR12 and AtCHR23 are expressed during embryo development, and that over-expression of AtCHR12 or AtCHR23 affects the embryo maturation phase with consequences on two important developmental transitions in plant life – germination and transition to flowering.

Over-expression of AtCHR12 or AtCHR23 leads to reduced seed germination, which is more pronounced under stress conditions (Chapter 3). The reduced germination of over-expressing lines is associated with increased transcript levels of seed maturation genes and reduced degradation of their mRNAs in germinating seeds. The results indicate that repression of AtCHR12 and AtCHR23 in germinating seeds is required for full germination.

The connection between embryo development and flowering time control was observed in transgenic lines over-expressing AtCHR12 (Chapter 4). Over-expression of AtCHR12 results in early flowering under both long- and short-day conditions. The early flowering phenotype correlates with reduced expression of the main flowering repressor, FLC. The reduced FLC expression correlates with increased levels of repressive histone mark H3K27me3 on the FLC locus. Additionally, FLC expression was affected already during FLC reprogramming, which takes place during embryo development. This leads to reduced FLC expression in mature embryos. The results show that AtCHR12 over-expression affects flowering time by different mechanisms than other Snf2-subfamily ATPases. In contrast to AtCHR12, BRAHMA was shown to regulate flowering time via the photoperiod pathway, while SPLAYED affects flowering time by repressing FT expression.

We have observed that over-expression of AtCHR12 or AtCHR23 affects plant growth in response to stress, and play a role in germination and transition to flowering. These traits are also important for agriculture, and such genes are potentially interesting targets for breeding programs. To test, if such genes have a similar role in crops, we have studied the effect of the tomato ortholog of AtCHR12 and AtCHR23 on tomato growth.

Tomato (Solanum lycopersicum), as well as other crops, have only one ortholog of AtCHR12 and AtCHR23, which was suggested to possess a role of both ATPases. We have successfully cloned the tomato ortholog and over-expressed it in tomato plants (Chapter 5). The transgenic tomato plants have reduced vegetative growth and compacted reproductive structures, resembling the phenotype of AtCHR23 and AtCHR12 over-expression, respectively. However, in contrast to Arabidopsis, the tomato plants responded to abiotic stress similarly as wild-type, and they flowered later than wild-type plants. The results indicate that modification of expression of AtCHR12 and AtCHR23 orthologs could be used to develop novel methods to control plant growth.

Taken together, the research described in this thesis identifies AtCHR12 and AtCHR23 as regulators of plant growth, especially in response to environment, as well as of the seed maturation program with clear effects on seed germination and flowering time, and we show that such genes can be potentially interesting for agriculture and horticulture practice.

 

Original languageEnglish
QualificationDoctor of Philosophy
Awarding Institution
  • Wageningen University
Supervisors/Advisors
  • Bisseling, Ton, Promotor
  • Mlynarova, Ludmila, Co-promotor
Award date19 Nov 2015
Place of PublicationWageningen
Publisher
Print ISBNs9789462575561
Publication statusPublished - 2015

Keywords

  • plants
  • growth
  • gene expression
  • chromatin
  • atp
  • arabidopsis
  • solanum lycopersicum
  • seed germination
  • growth regulators

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