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Wax Crystal-Sparse Leaf 4, encoding a β-ketoacyl-coenzyme A synthase 6, is involved in rice cuticular wax accumulation

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Abstract

Key message

WSL4 encodes a KCS6 protein which is required for cuticular wax accumulation in rice.

Abstract

Very long chain fatty acids (VLCFAs) are essential precursors for cuticular wax biosynthesis. VLCFA biosynthesis occurs in the endoplasmic reticulum and requires the fatty acid elongase (FAE) complex. The β-ketoacyl-coenzyme A synthase (KCS) catalyzes the first step of FAE-mediated VLCFA elongation. Here we characterized the Wax Crystal-Sparse Leaf 4 (WSL4) gene involved in leaf cuticular wax accumulation in rice. The wsl4 mutant displayed a pleiotropic phenotype including dwarfism, less tiller numbers and reduced surface wax load. Map-based cloning and nucleotide sequencing results revealed that wsl4 carried a single nucleotide substitution in the second exon of a putative KCS6 gene, encoding one subunit of the FAE complex for VLCFAs. Genetic complementation confirmed that the mutation in WSL4 was responsible for the phenotype of wsl4. WSL4 was constitutively expressed in various rice tissues and localized in the endoplasmic reticulum. Both WSL4-RNAi transgenic lines and WSL4 knocked-out mutants exhibited wax-deficient phenotypes similar to the wsl4 mutant. These data indicate that WSL4 is required for cuticular wax accumulation in rice.

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Abbreviations

CER:

Eceriferum

CoA:

Coenzyme A

ECR:

Enoyl-CoA reductase

ER:

Endoplasmic reticulum

FAE:

Fatty acid elongation

GC–MS:

Gas chromatography–mass spectrometry

GFP:

Green fluorescent protein

GUS:

β-Glucuronidase

HCD:

β-Hydroxyacyl-Coa dehydratase

KCR:

β-Ketoacyl-CoA reductase

KCS:

β-Ketoacyl-CoA synthase

RNAi:

RNA interference

SEM:

Scanning electron microscope

TEM:

Transmission electron microscopy

ORF:

Open reading frame

qRT-PCR:

Quantitative RT-PCR

UBQ:

Ubiquitin

VLCFAs:

Very long chain fatty acids

WSL:

Wax Crystal-Sparse Leaf

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Acknowledgements

This work was supported by the National Key R&D Program of China (2016YFD0100600, 2016YFD0200700), and the National Special Project of China (2014ZX08001-006).

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Author notes

  1. Lu Gan, Shanshan Zhu, and Zhichao Zhao contributed equally to this work.

Authors and Affiliations

  1. National Key Facility for Crop Gene Resources and Genetic Improvement, Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing, 100081, China

    Lu Gan, Shanshan Zhu, Zhichao Zhao, Xiaole Wang, Zhe Zhang, Xin Zhang, Jie Wang, Jiulin Wang, Xiuping Guo & Jianmin Wan

  2. National Key Laboratory for Crop Genetics and Germplasm Enhancement, Jiangsu Plant Gene Engineering Research Center, Nanjing Agricultural University, Nanjing, 210095, China

    Linglong Liu & Jianmin Wan

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  1. Lu Gan
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Correspondence to Jianmin Wan.

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Additional information

Communicated by Qiao Zhao.

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299_2017_2181_MOESM1_ESM.pdf

Supplementary Figure 1. SEM analysis of epicuticular wax crystal patterns on the leaf sheaths of WT (A, B, C) and wsl4 (D, E, F), Bar=1μm. (PDF 241 kb)

Supplementary Table 1. Primers used in this paper. (XLSX 11 kb)

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Gan, L., Zhu, S., Zhao, Z. et al. Wax Crystal-Sparse Leaf 4, encoding a β-ketoacyl-coenzyme A synthase 6, is involved in rice cuticular wax accumulation. Plant Cell Rep 36, 1655–1666 (2017). https://doi.org/10.1007/s00299-017-2181-5

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