ABA inhibits myristoylation and induces shuttling of the RGLG1 E3 ligase to promote nuclear degradation of PP2CA

Abstract

Summary Hormone‐ and stress‐induced shuttling of signaling or regulatory proteins is an important cellular mechanism to modulate hormone signaling and cope with abiotic stress. Hormone‐induced ubiquitination plays a crucial role to determine the half‐life of key negative regulators of hormone signaling. For ABA signaling, the degradation of clade‐A PP2Cs, such as PP2CA or ABI1, is a complementary mechanism to PYR/PYL/RCAR‐mediated inhibition of PP2C activity. ABA promotes the degradation of PP2CA through the RGLG1 E3 ligase, although it is not known how ABA enhances the interaction of RGLG1 with PP2CA given that they are predominantly found in the plasma membrane and the nucleus, respectively. We demonstrate that ABA modifies the subcellular localization of RGLG1 and promotes nuclear interaction with PP2CA. We found RGLG1 is myristoylated in vivo, which facilitates its attachment to the plasma membrane. ABA inhibits the myristoylation of RGLG1 through the downregulation of N‐myristoyltransferase 1 (NMT1) and promotes nuclear translocation of RGLG1 in a cycloheximide‐insensitive manner. Enhanced nuclear recruitment of the E3 ligase was also promoted by increasing PP2CA protein levels and the formation of RGLG1–receptor–phosphatase complexes. We show that RGLG1Gly2Ala mutated at the N‐terminal myristoylation site shows constitutive nuclear localization and causes an enhanced response to ABA and salt or osmotic stress. RGLG1/5 can interact with certain monomeric ABA receptors, which facilitates the formation of nuclear complexes such as RGLG1–PP2CA–PYL8. In summary, we provide evidence that an E3 ligase can dynamically relocalize in response to both ABA and increased levels of its target, which reveals a mechanism to explain how ABA enhances RGLG1–PP2CA interaction and hence PP2CA degradation.

The MATH-BTB BPM3 and BPM5 subunits of Cullin3-RING E3 ubiquitin ligases target PP2CA and other clade A PP2Cs for degradation

“…Transient expression experiments in N. benthamiana revealed that ABA enhances BPM3/5-mediated degradation of PP2CA, in agreement with previous results for PUB12-mediated degradation of ABI1 or RGLG1-mediated degradation of PP2CA (19)(20)(21). Therefore, these data suggest that at resting ABA levels, PP2C protein levels remain sufficient to block ABA signaling because interaction with E3 ligases is dependent on ABA concentration.…”

“…Recently, studies that address the turnover of core ABA signaling components have been published (19)(20)(21)(27)(28)(29)(30)(31)(32). Regarding the turnover of clade A PP2Cs, in the case of ABI1, it has been demonstrated that ABA induces degradation of this PP2C through PUB12/13 E3 ligases (19).…”

“…PP2CA is another key negative regulator of ABA signaling (14,(33)(34)(35)(36)(37). Recently, we demonstrated that the RGLG1 and RGLG5 RING-type E3 ubiquitin ligases relieve PP2C blockade of ABA signaling by mediating PP2CA protein degradation, which is enhanced by ABA (20,21). However, experiments with double rglg1 rglg5 mutants indicated that additional unidentified E3s contribute to regulation of PP2CA levels (20).…”

“…Transcription of some PYR/PYL/RCARs is repressed whereas that of PP2Cs is stimulated in response to ABA (5,18), indicating there exists a negative feedback transcriptional mechanism to modulate ABA signaling by controlling transcript levels of core elements. Recently, it has been discovered that degradation of PP2Cs is a complementary mechanism to PYR/PYL/RCAR-mediated inhibition of PP2C activity (19)(20)(21).…”

Abscisic acid dynamics, signaling, and functions in plants

“…In contrast, under stressed conditions or ABA treatment, activities of PP2Cs can be reduced by ABA receptor PYLs and several other regulators, such as the putative leucine‐rich repeat‐RLK, RECEPTOR DEAD KINASE1 (RDK1) that can promote ABA responses by interacting with ABI1 (Kumar et al 2017). PP2Cs are also degraded by the 26S proteasome pathway through the PUB12/13 U‐box, RGLG1/5 RING‐type and multimeric cullinc3 (CUL3)‐RING‐based E3 ligases by interacting with the adaptor BTB/POZ AND MATH DOMAIN proteins (BPMs), which is promoted by ABA (Figure 2) (Kong et al 2015; Wu et al 2016; Belda‐Palazon et al 2019; Julian et al 2019).…”