FYVE1/FREE1 Interacts with the PYL4 ABA Receptor and Mediates Its Delivery to the Vacuolar Degradation Pathway

Abstract

Recently, we described the ubiquitylation of PYL4 and PYR1 by the RING E3 ubiquitin ligase RSL1 at the plasma membrane of This suggested that ubiquitylated abscisic acid (ABA) receptors might be targeted to the vacuolar degradation pathway because such ubiquitylation is usually an internalization signal for the endocytic route. Here, we show that FYVE1 (previously termed FREE1), a recently described component of the endosomal sorting complex required for transport (ESCRT) machinery, interacted with RSL1-receptor complexes and recruited PYL4 to endosomal compartments. Although the ESCRT pathway has been assumed to be reserved for integral membrane proteins, we show the involvement of this pathway in the degradation of ABA receptors, which can be associated with membranes but are not integral membrane proteins. Knockdown alleles are hypersensitive to ABA, illustrating the biological relevance of the ESCRT pathway for the modulation of ABA signaling. In addition, mutants are impaired in the targeting of ABA receptors for vacuolar degradation, leading to increased accumulation of PYL4 and an enhanced response to ABA Pharmacological and genetic approaches revealed a dynamic turnover of ABA receptors from the plasma membrane to the endosomal/vacuolar degradation pathway, which was mediated by FYVE1 and was dependent on RSL1. This process involves clathrin-mediated endocytosis and trafficking of PYL4 through the ESCRT pathway, which helps to regulate the turnover of ABA receptors and attenuate ABA signaling.

Proteasome‐independent functions of lysine‐63 polyubiquitination in plants

“…The ELC/AtVPS23A ESCRT-I component also binds Ub in vitro (Spitzer et al, 2006). Interestingly, ELC/ AtVPS23A and AtFYVE1 directly bind to the AtPYL4 abscisic acid (ABA) receptor and AtIRT1, two ubiquitinated endocytosed cargos (Barberon et al, 2014;Belda-Palazon et al, 2016;Yu et al, 2016). Consistently, genetic interference with ELC/AtVPS23A or AtFYVE1 expression yields ubiquitinated protein accumulation in (Zhou et al, 2013), sequestration of anthocyanins in the vacuole (Pourcel et al, 2010), chlorophagy of physiological and damaged chloroplasts (Izumi et al, 2017), reticulophagy (Liu et al, 2012), pexophagy (Shibata et al, 2013), mitophagy (Li et al, 2014) and xenophagy (Hafren et al, 2017).…”

“…The BR receptor AtBRI1 is indeed decorated with K63-linked chains in vivo, undergoes K63 polyUb-dependent New Phytologist endocytosis and the expression of a ubiquitination-defective BRI1 variant leads to BR hypersensitivity (Martins et al, 2015). Similarly, the plasma membrane-associated pool of AtPYR/AtPYL ABA receptors is subjected to ubiquitination at the plasma membrane by the AtRSL1 RING E3 ligase and ESCRT-dependent endocytic sorting (Bueso et al, 2014;Belda-Palazon et al, 2016;Yu et al, 2016). Consistently, Arabidopsis fyve1 and vps23 mutants show stabilization of the AtPYL4 ABA receptor and are hypersensitive to ABA.…”

A novel family of transcription factors conserved in angiosperms is required for ABA signalling

“…So far, two main types of negative feedback regulatory loops of ABA signalling have been identified: the ubiquitin‐mediated proteasomal degradation of ABA key central regulators and the expressional regulation of the key central regulators genes. E3 ligases such as KEG, DWA1, and DWA2 target ABF/AREB/ABI5‐type bZIP transcription factors for proteasomal degradation (Chen, Liu, Stone, & Callis, ; Lee, Kang, et al, ; Liu & Stone, , ; Stone, Williams, Farmer, Vierstra, & Callis, ), whereas E2 ligase VPS23A and E3 ligases CUL4 and RSL1 promote the proteasomal degradation of PYR/PYL/RCABR receptors (Belda‐Palazon et al, ; Bueso et al, ; Seo et al, ; Yu et al, ). E3 ligases PUB12 and PUB13 target PP2C proteins, but only when they interact with ABA receptors (Kong et al, ).…”

ESCRT‐dependent vacuolar sorting and degradation of the auxin biosynthetic enzyme YUC1 flavin monooxygenase

“…Membrane‐localized auxin efflux carrier PIN2 (Spitzer et al ) and Brassinosteroid receptor BRI1 (Martins et al ) have been reported to be PVC cargo proteins, and are sorted for vacuolar degradation by the ESCRT system. Although the ESCRT pathway has long been considered specifically to be targeting integral membrane proteins, recent studies have demonstrated that non‐integral membrane proteins, such as the Arabidopsis ABA receptor PYR1‐LIKE 4 (PYL4), also employ ESCRT machinery components FYVE1 and VPS23A for endosomal sorting (Belda‐Palazon et al ; Yu et al ).…”