SKP1-like protein, CrSKP1-e, interacts with pollen-specific F-box proteins and assembles into SCF-type E3 complex in 'Wuzishatangju' (Citrus reticulata Blanco) pollen

Yi Ren; Qingzhu Hua; Jiayan Pan; Zhike Zhang; Jietang Zhao; Xinhua He; Yonghua Qin; Guibing Hu

doi:10.7717/peerj.10578

SKP1-like protein, CrSKP1-e, interacts with pollen-specific F-box proteins and assembles into SCF-type E3 complex in 'Wuzishatangju' (Citrus reticulata Blanco) pollen

PeerJ. 2020 Dec 22:8:e10578. doi: 10.7717/peerj.10578. eCollection 2020.

Authors

Yi Ren¹, Qingzhu Hua¹, Jiayan Pan¹, Zhike Zhang¹, Jietang Zhao¹, Xinhua He², Yonghua Qin¹, Guibing Hu¹

Affiliations

¹ State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/ Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, College of Horticulture, South China Agricultural University, Guangzhou, China.
² State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources, College of Agriculture, Guangxi University, Nanning, China.

Abstract

S-ribonuclease (S-RNase)-based self-incompatibility (SI) mechanisms have been extensively studied in Solanaceae, Rosaceae and Plantaginaceae. S-RNase-based SI is controlled by two closely related genes, S-RNase and S-locus F-box (SLF), located at a polymorphic S-locus. In the SI system, the SCF-type (SKP1-CUL1-F-box-RBX1) complex functions as an E3 ubiquitin ligase complex for ubiquitination of non-self S-RNase. Pummelo (Citrus grandis) and several mandarin cultivars are suggested to utilize an S-RNase-based SI system. However, the molecular mechanism of the non-S-factors involved in the SI reaction is not straightforward in Citrus. To investigate the SCF-type E3 complex responsible for the SI reaction in mandarin, SLF, SKP1-like and CUL1 candidates potentially involved in the SI reaction of 'Wuzishatangju' (Citrus reticulata Blanco) were identified based on the genome-wide identification and expression analyses. Sixteen pollen-specific F-box genes (CrFBX1-CrFBX16), one pollen-specific SKP1-like gene (CrSKP1-e) and two CUL1 genes (CrCUL1A and CrCUL1B) were identified and cloned from 'Wuzishatangju'. Yeast two-hybrid (Y2H) and in vitro binding assays showed that five CrFBX proteins could bind to CrSKP1-e, which is an ortholog of SSK1 (SLF-interacting-SKP1-like), a non-S-factor responsible for the SI reaction. Luciferase complementation imaging (LCI) and in vitro binding assays also showed that CrSKP1-e interacts with the N-terminal region of both CrCUL1A and CrCUL1B. These results indicate that CrSKP1-e may serve as a functional member of the SCF-type E3 ubiquitin ligase complex in 'Wuzishatangju'.

Keywords: Citrus retuculata Blanco; Cullin1; F-box; SCF complex; SKP1-like; Self-incompatibility.

Associated data

figshare/10.6084/m9.figshare.11786199.v1

Grants and funding

This work was supported by the National Natural Science Foundation of China (No. 31471858 and 31000899), the Science and Technology Planning Project of Guangzhou (No. 201807010018, 201607010314 and 2010r1-C771), and the Natural Science Foundation of Guangdong Province (No. S2013020013084 and S2013010011950). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.