Arabidopsis PIZZA Has the Capacity to Acylate Brassinosteroids

Katja Schneider,Yuji Kamiya,Youichi Kondou,Yusuke Jikumaru,Takanari Ichikawa,Atsushi Hanada,Shozo Fujioka,Shinjiro Yamaguchi,Ayako Kawamura,Christian Breuer,Keiko Sugimoto,Minami Matsui

doi:10.1371/journal.pone.0046805

Katja Schneider, Yuji Kamiya + Show 10 more

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https://doi.org/10.1371/journal.pone.0046805

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Abstract

Brassinosteroids (BRs) affect a wide range of developmental processes in plants and compromised production or signalling of BRs causes severe growth defects. To identify new regulators of plant organ growth, we searched the Arabidopsis FOX (Full-length cDNA Over-eXpressor gene) collection for mutants with altered organ size and isolated two overexpression lines that display typical BR deficient dwarf phenotypes. The phenotype of these lines, caused by an overexpression of a putative acyltransferase gene PIZZA (PIZ), was partly rescued by supplying exogenous brassinolide (BL) and castasterone (CS), indicating that endogenous BR levels are rate-limiting for the growth of PIZ overexpression lines. Our transcript analysis further showed that PIZ overexpression leads to an elevated expression of genes involved in BR biosynthesis and a reduced expression of BR inactivating hydroxylases, a transcriptional response typical to low BR levels. Taking the advantage of relatively high endogenous BR accumulation in a mild bri1-301 background, we found that overexpression of PIZ results in moderately reduced levels of BL and CS and a strong reduction of typhasterol (TY) and 6-deoxocastasterone (6-deoxoCS), suggesting a role of PIZ in BR metabolism. We tested a set of potential substrates in vitro for heterologously expressed PIZ and confirmed its acyltransferase activity with BL, CS and TY. The PIZ gene is expressed in various tissues but as reported for other genes involved in BR metabolism, the loss-of-function mutants did not display obvious growth phenotypes under standard growth conditions. Together, our data suggest that PIZ can modify BRs by acylation and that these properties might help modulating endogenous BR levels in Arabidopsis.

Highlights

How characteristic size of organs is controlled in multicellular organisms is a fascinating and important question in biology
We recovered the cDNA inserted in both lines by PCR using primers specific for the FOX vector and found by sequencing analysis that both cDNAs encode the same gene (At4g31910) which we named PIZZA (PIZ) for the round leaf phenotypes of its overexpressor
The transgenic lines expressing PIZ cDNA driven by the CaMV 35S (35S) promoter in wild-type background reproduced the dwarf phenotype identical to F23131 and F28215 plants (Figure 2), confirming that the phenotype of the FOX lines is caused by overexpression of the PIZ gene

Summary

Introduction

How characteristic size of organs is controlled in multicellular organisms is a fascinating and important question in biology. The final size of plant organs is determined by the balance between cell proliferation and cell differentiation. In the subsequent postmitotic phase, cells differentiate and expand their volume through water uptake into the vacuole and cell wall biogenesis. This step is often associated with an alternative cell cycle called endoreduplication cycle or endocycle in which cells amplify the nuclear DNA content to increase their ploidy level [1]. Controlling the duration of cell proliferation and cell expansion or the transition from the proliferative phase to the expansion phase is crucial for the determination of final organ size (reviewed in [2]). Accumulating evidence suggest that these processes are controlled both transcriptionally and post-translationally, and several transcriptional regulators or enzymes involved in the post-translational protein modification have been shown to act on cell proliferation or cell expansion [3,4]

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