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FLA, which encodes a homolog of UBP, is required for chlorophyll accumulation and development of lemma and palea in rice

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Abstract

Key message

FLA, the homolog of ubiquitin-specific protease does not have deubiquitination activity, but it is essential for flower and chloroplast development in rice.

Abstract

Ubiquitin-specific proteases (UBPs) are widely distributed and highly conserved proteins and are also members of the most important family of deubiquitination enzymes. Although the functions and phylogenies of UBPs from yeast, mammals and Arabidopsis have been widely reported, the functions and evolutionary relationships of UBPs in rice remain unclear. In this study, we characterized the rice flower and leaf color aberrant mutant (fla), which exhibited a variety of developmental defects, including abnormal floral organs and pollen development, and leaf bleaching. We isolated FLA by positional cloning and found that it encodes a homolog of ubiquitin-specific protease. FLA is a ubiquitously expressed gene with the highest expression in floral organs. Subcellular localization analysis indicated that FLA is a cell membrane protein. Through searches of the rice genome database (http://rice.plantbiology.msu.edu), we identified 35 UBP family members in the rice genome. These proteins were grouped into 16 subfamilies based on phylogenetic analysis, and FLA was found to belong to the G8 subfamily. In vitro activity assays revealed that FLA does not have deubiquitination activity. Our data suggest that FLA plays an important role in the development of floral organs and chloroplast in rice, but that this role probably does not involve deubiquitination activity, because FLA does not have an active site and deubiquitination activity.

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Abbreviations

UBP/USP:

Ubiquitin-specific protease

DUBs:

Deubiquitination enzymes

MATH:

Meprin and TRAF homology

UBQ:

Ubiquitin homolog

ZnF-UBP:

Zinc-finger ubiquitin-specific protease

UBA:

Ubiquitin-associated

ZnF-MYND:

Myeloid, Nervy and DEAF1-type zinc finger

DUSP:

Domain in USPs

I2–KI:

Iodine–potassium iodide

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Acknowledgements

This work was supported by National Natural Science Foundation of China (No. 31501287), National Key Research and Development Program of China (2016YFD0100101, 2016YFD0100301), National Science and Technology Support Program of China (2015BAD01B01-1), CAAS Science and Technology Innovation Program, National Infrastructure for Crop Germplasm Resources (NICGR2017-01), Protective Program of Crop Germplasm of China (2017NWB 036-01, 2017NWB036-12-2).

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  1. Xiaoding Ma and Jiani Zhang contributed equally to this work.

Authors and Affiliations

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

    Xiaoding Ma, Bing Han, Di Cui & Longzhi Han

  2. Chongqing Normal University, Chongqing, 401331, China

    Jiani Zhang & Jianghong Tang

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  1. Xiaoding Ma
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Correspondence to Longzhi Han.

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Supplementary Fig. S1

Comparison of plant agronomic traits between WT and fla. a–c Quantification of the seed set of the main panicle (a), tiller number (b) and height (c) of WT and fla plants. The percentage of the seed set in the main panicle of the fla mutant includes normal seed and seed produced by three types of mutant flowers (types 2, 3, and 5; Fig.1a). Values are the means ± SD (n=20). *P<0.01 (t test). d WT and fla panicles. Bar 5 cm (d) (JPG 450 KB)

Supplementary Fig. S2

Phylogenetic analysis of the UBP gene family in Arabidopsis and rice. Bootstrap values are shown at each node (JPG 2062 KB)

Supplementary Fig. S3

Alignment of the 35 rice UBP proteins. Black and gray boxes denote identical and similar amino acids, respectively (PDF 1570 KB)

Supplementary Fig. S4

Analysis of deubiquitination activity in vitro with rice UBQ10 as the substrate. UBP6 was used as a positive control. Anti-ubiquitin antibody was used to detect ubiquitin. Cleaved products are indicated by black arrowheads. IB, immunoblot (JPG 118 KB)

Supplementary Fig. S5

Analysis of soluble and insoluble pUb protein accumulation in WT and fla. A section of a representative Coomassie brilliant blue-stained SDS-PAGE gel was used as a loading control (bottom). SP, soluble pUb proteins; ISP, insoluble pUb proteins (JPG 23 KB)

Supplementary material 6 (DOCX 18 KB)

Supplementary material 7 (DOCX 17 KB)

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Ma, X., Zhang, J., Han, B. et al. FLA, which encodes a homolog of UBP, is required for chlorophyll accumulation and development of lemma and palea in rice. Plant Cell Rep 38, 321–331 (2019). https://doi.org/10.1007/s00299-018-2368-4

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  • DOI: https://doi.org/10.1007/s00299-018-2368-4

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