Arabidopsis SH3P2 is an ubiquitin-binding protein that functions together with ESCRT-I and the deubiquitylating enzyme AMSH3

Abstract

Clathrin-mediated endocytosis of plasma membrane proteins is an essential regulatory process that controls plasma membrane protein abundance and is therefore important for many signaling pathways, such as hormone signaling and biotic and abiotic stress responses. On endosomal sorting, plasma membrane proteins maybe recycled or targeted for vacuolar degradation, which is dependent on ubiquitin modification of the cargos and is driven by the endosomal sorting complexes required for transport (ESCRTs). Components of the ESCRT machinery are highly conserved among eukaryotes, but homologs of ESCRT-0 that are responsible for recognition and concentration of ubiquitylated proteins are absent in plants. Recently several ubiquitin-binding proteins have been identified that serve in place of ESCRT-0; however, their function in ubiquitin recognition and endosomal trafficking is not well understood yet. In this study, we identified Src homology-3 (SH3) domain-containing protein 2 (SH3P2) as a ubiquitin- and ESCRT-I-binding protein that functions in intracellular trafficking. SH3P2 colocalized with clathrin light chain-labeled punctate structures and interacted with clathrin heavy chain , indicating a role for SH3P2 in clathrin-mediated endocytosis. Furthermore, SH3P2 cofractionates with clathrin-coated vesicles (CCVs), suggesting that it associates with CCVs Mutants of and genetically interact, suggesting that they could function in the same pathway. Based on these results, we suggest a role of SH3P2 as an ubiquitin-binding protein that binds and transfers ubiquitylated proteins to the ESCRT machinery.

Meeting report – Cellular gateways: expanding the role of endocytosis in plant development

“…Four complexes are found in mammals and some fungi, ESCRT-0, I, II and III; in plants, ESCRT-0 is missing. Erika Isono (University of Konstanz, Germany) reported that the SH3 domaincontaining protein 2 (SH3P2) that is present in CCVs is an ubiquitinbinding protein that can interact with both the deubiquitylating enzyme associated molecule with the SH3 domain of STAM3 (AMSH3) and ESCRT-I (Nagel et al, 2017). In addition, AMSH3 function is regulated by apoptosis-linked gene-2 interacting protein X (ALIX) (Kalinowska et al, 2015), which suggests that SH3P2, AMSH3, ESCRT-I and ALIX function together to send ubiquitylated cargos for degradation through the ESCRT pathway.…”

Proteasome‐independent functions of lysine‐63 polyubiquitination in plants

“…The substrate preference of TOL proteins is not known, but the presence of tandem UBDs in TOLs probably provides a high avidity for K63-linked Ub chains. The AtSH3P2 protein binds K63 polyubiquitinated proteins and is localized at the plasma membrane, in clathrin-coated vesicles and in endosomes (Nagel et al, 2017). AtSH3P2 associates with clathrin and some of the plant ESCRT-I subunits to probably funnel Box 1 Dual role of K63 polyubiquitination in endocytosis of yeast and mammals…”

“…AtFYVE1 is involved in autophagy via a direct association with the ATG8/LC3-interacting AtSH3P2 protein, which participates in autophagosome formation (Zhuang et al, 2013). Interestingly, both AtFYVE1 and AtSH3P2 bind Ub, although the linkage preference has not been determined for AtFYVE1 (Gao et al, 2014;Nagel et al, 2017). The precise role of AtAMSH1 and AtFYVE1 and, more specifically, of their ability to bind to or hydrolyze K63-linked Ub chains, in autophagy, is still unclear.…”

TOLs Function as Ubiquitin Receptors in the Early Steps of the ESCRT Pathway in Higher Plants

“…The plantspecific FYVE1/FREE1 localizes to MVBs and binds ubiquitin as well as PtdIns(3)P, and furthermore interacts with the Src homology-3 (SH3) domain-containing protein 2 (SH3P2), VPS23A/B, and the ESCRT-III complex (Barberon et al, 2014;Gao et al, 2014;Kolb et al, 2015;Belda-Palazon et al, 2016). At the PM, the ubiquitin-binding protein SH3P2 has been implicated in vacuolar trafficking of ubiquitinated cargoes, as it binds K63-linked ubiquitin and VPS23A and localizes to clathrin-coated vesicles (Kolb et al, 2015;Nagel et al, 2017). Target of Myb 1 (TOM1) proteins are evolutionarily ancient ubiquitin receptors, which function in the capturing of ubiquitinated cargo and are thought to act together with or replace the ESCRT-0 machinery (Wang et al, 2010).…”