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Transcriptional control of local auxin distribution by the CsDFB1-CsPHB module regulates floral organogenesis in cucumber
Monday, 2021/03/01 | 08:28:19

Jing Nie,  Nan Shan, Huan Liu, Xuehui Yao, Ziwei Wang, Ruoxue Bai, Yicong Guo,  Ying Duan,  Changlin Wang, and  Xiaolei Sui

PNAS February 23, 2021 118 (8) e2023942118

Significance

Auxin is a key phytohormone influencing multiple aspects of plant development, including meristem maintenance, primordia initiation, floral organogenesis, and vascular differentiation. Local auxin biosynthesis and polar auxin transport are essential to establish and maintain auxin gradients that ensure proper plant development. Here, we demonstrate that CsDFB1, a member of the plant cystatin superfamily, which was previously implicated in defense responses, plays a critical role in regulating local auxin distribution and thus influences floral organogenesis in cucumber. Genetic and biochemical assays suggest that CsDFB1 affects local auxin distribution by acting as an attenuator that interacts with CsPHB and modulates CsPHB-mediated transcriptional control of CsYUC2 and CsPIN1. Our results shed light on the fine tuning of local auxin distribution in plants.

Abstract

Plant cystatins are cysteine proteinase inhibitors that play key roles in defense responses. In this work, we describe an unexpected role for the cystatin-like protein DEFORMED FLORAL BUD1 (CsDFB1) as a transcriptional regulator of local auxin distribution in cucumber (Cucumis sativus L.). CsDFB1 was strongly expressed in the floral meristems, floral primordia, and vasculature. RNA interference (RNAi)-mediated silencing of CsDFB1 led to a significantly increased number of floral organs and vascular bundles, together with a pronounced accumulation of auxin. Conversely, accompanied by a decrease of auxin, overexpression of CsDFB1 resulted in a dramatic reduction in floral organ number and an obvious defect in vascular patterning, as well as organ fusion. CsDFB1 physically interacted with the cucumber ortholog of PHABULOSA (CsPHB), an HD-ZIP III transcription factor whose transcripts exhibit the same pattern as CsDFB1. Overexpression of CsPHB increased auxin accumulation in shoot tips and induced a floral phenotype similar to that of CsDFB1-RNAi lines. Furthermore, genetic and biochemical analyses revealed that CsDFB1 impairs CsPHB-mediated transcriptional regulation of the auxin biosynthetic gene YUCCA2 and the auxin efflux carrier PIN-FORMED1, and thus plays a pivotal role in auxin distribution. In summary, we propose that the CsDFB1-CsPHB module represents a regulatory pathway for local auxin distribution that governs floral organogenesis and vascular differentiation in cucumber.

 

See: https://www.pnas.org/content/118/8/e2023942118

 

Figure 2: Phenotypic analysis of CsDFB1 transgenic plants. (A) RT-qPCR analysis of CsDFB1 (Csa7G067350) expression in floral buds from WT plants, RNA interference lines (RNAi#18 and RNAi#33), and overexpression lines (OE#3 and OE#27). Error bars indicate SD of three biological replicates from different plants. (BI) Detailed phenotypes of MFB (BD) and FFB (EI) from transgenic lines (RNAi#18 and OE#3) and WT plants as control. (BCE, and F) Morphological characteristics of petals from MFBs (B) and FFBs (E), stamens (C), and stigma (F). (G) Hand cross-section of ovary/fruit. (H) Paraffin cross-sections of MVBs in the ovary/fruit. Yellow, blue, and red regions indicate the EP, xylem, and IP, respectively. (D and I) Hand cross-sections of pedicels (D) and peduncles (I) stained with phloroglucinol. Red regions indicate the presence of lignified xylem cells. Blue and white numbers indicate the number of petals (B and E), stigma (F), and carpels (G), respectively; red arrows in G indicate the position of MVBs. (Scale bars, 1 cm in B and E; 1 mm in DG, and I; 500 μm in C and F; and 100 μm in H.) (J) Morphology of cucumber fruits after 30 d of development. Red asterisk indicates organ (fruit) fusion in OE lines. (Scale bar, 2 cm.) (K) Summary of floral organ number and MVB number in the WT and transgenic plants. Error bars indicate SD of 50 flowers from around 10 individual plants. Significance analysis was conducted with the two-tailed Student’s t test (***P < 0.001; **P < 0.01). Abbreviations: DEFORMED FLORAL BUD1 (CsDFB1); MFB, male floral bud; FFB, female floral bud; EP, external phloem; X, xylem; IP, internal phloem; and MVB, main vascular bundles.

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