Understanding the Role of Chloroplast Signalling in Plant Development and Drought Tolerance
Creators: name: Phua, Su Yin
Publisher: name: The Australian National University
Year: 2015
Subjects: SAL1
PAP
transgenics
drought
germination
plant hormones

description: 3’-phosphoadenosine 5’-phosphate (PAP) was recently proposed as a new chloroplast retrograde signal that accumulates during drought stress. PAP is a by-product of secondary sulfur assimilation, generated by SULFOTRANSFERASES (SOTs) upon transferring of the sulfate group from 3’-phosphoadenosine 5’-phosphosulfate (PAPS) to sulfate acceptor molecules. PAP is efficiently degraded by the SAL1 phosphatase into inorganic phosphate and adenosine monophosphate (AMP). Consequently, PAP levels are normally kept at very low during plant development. Constitutive over-accumulation of this retrograde signal resulted in plants with pleiotropic altered phenotypes as demonstrated by the sal1 mutant in Arabidopsis, namely improved drought tolerance, altered rosette morphology and delayed development. These multifaceted altered phenotypes of sal1 correlate well with the many transcriptional changes, which are mainly due to the inhibition of 5’- 3’ EXORIBONUCLEASES (XRNs) by PAP. However, the details of downstream changes that links SAL1-PAP-XRN to the many altered phenotypes of sal1 remains poorly understood. This thesis investigates the possibility of conferring drought tolerance to plants by manipulating the SAL1 gene expression with minimal negative effects on growth, while unraveling the signalling pathway(s) contributing to sal1 altered development and drought tolerance. I hypothesised that PAP accumulation at early developmental stages is detrimental for plant growth and development while its accumulation at later stages of development or prior to drought stress is beneficiary for conferring plant drought tolerance. To test this hypothesis, two different strategies for easy manipulation of SAL1 expression were attempted: inducible silencing of SAL1 in wild-type Arabidopsis and inducible complementation of SAL1 in the sal1 mutant background. Surprisingly, efficient silencing of SAL1 could not be achieved even with a strong constitutive promoter driving the expression of SAL1-hair-pin RNA interference (hpRNAi) or SAL1-artificial microRNA (amiRNA) and inducible-silencing was similarly inefficient. On the other hand, inducible complementation of SAL1 in the sal1 mutant background allowed for better manipulation of SAL1 expression and PAP levels. This provides the platform for exploring the engineering of drought tolerant plants without compromising plant development by manipulating PAP levels via regulating SAL1 expression. Since hormones are key regulators of plant development, the possible interaction between PAP and hormone signalling was investigated to study the basis of sal1 developmental phenotypes. Significantly, a comprehensive hormonal profiling showed reduced gibberellins (GAs) content in sal1 and improved growth rate was achieved when sal1 was treated with GAs and brassinosteroids (BRs), which are interdependent hormones that promote growth. Additionally, sal1 germination was hyper-responsive to treatments with abscisic acid (ABA) and/or GAs biosynthetic inhibitor - paclobutrazol (PAC). Furthermore, this observation can be reproduced by feeding wild-type seeds with PAP in addition to ABA and PAC, suggesting that PAP could act as secondary messenger for both GAs and ABA signalling. Meanwhile, upon taking into consideration both the findings of this thesis and literature available, it is proposed that SAL1-PAP-XRN stabilises DELLA accumulation by reducing transcription of key GA biosynthetic genes, allowing the DELLA protein to interact and affect the key transcription factors of light and other hormonal signalling pathways such as BRs, jasmonic acid (JA) and ethylene. It is likely that the overall interactions and feedback between 1) altered sulfur metabolism, 2) altered hormonal homeostasis and 3) inhibitory effects of PAP on XRNs and other nucleotide-binding proteins eventually culminate in the pleitropic developmental phenotype and drought tolerance of sal1. The ultimate application from this project will be to engineer drought tolerance in Brassicaceae by manipulating the SAL1-PAP pathway. This thesis has generated the tools that will aid in the above engineering and uncovered the potential connections between the sal1 altered phenotypes and SAL1-PAP-XRNs, linking chloroplast signalling to plant hormonal homeostasis in regulating plant development and drought tolerance.