SPX-domain-containing proteins (SPXs) play an important role in inorganic phosphate (Pi) sensing, signaling, and transport in eukaryotes. In plants, SPXs are known to integrate cellular Pi status and negatively regulate the activity of Pi central regulators, the PHOSPATE STARVATION RESPONSE proteins (PHRs). The stability of SPXs, such as SPX4, is reduced under Pi-deficient conditions. However, the mechanisms by which SPXs are degraded remain unclear. In this study, using a yeast-two-hybrid screen we identified two RING-finger ubiquitin E3 ligases regulating SPX4 degradation, designated SDEL1 and SDEL2, which were post-transcriptionally induced by Pi starvation. We found that both SDELs were located in the nucleus and cytoplasm, had ubiquitin E3 ligase activity, and directly ubiquitinated the K²¹³ and K²⁹⁹ lysine residues in SPX4 to regulate its stability. Furthermore, we found that PHR2, a Pi central regulator in rice, could compete with SDELs by interacting with SPX4 under Pi-sufficient conditions, which protected SPX4 from ubiquitination and degradation. Consistent with the biochemical function of SDEL1 and SDEL2, overexpression of SDEL1 or SDEL2 resulted in Pi overaccumulation and induced Pi-starvation signaling even under Pi-sufficient conditions. Conversely, their loss-of-function mutants displayed decreased Pi accumulation and reduced Pi-starvation signaling. Collectively, our study revealed that SDEL1 and SDEL2 facilitate the degradation of SPX4 to modulate PHR2 activity and regulate Pi homeostasis and Pi signaling in response to external Pi availability in rice.

含SPX域的蛋白质（SPX）在真核生物中的无机磷酸盐（Pi）感测，信号传导和运输中起重要作用。在植物中，已知SPX整合了细胞Pi的状态，并负调控Pi中央调节剂PHOSPATE STARVATION RESPONSE蛋白（PHRs）的活性。在Pi不足的情况下，SPX（例如SPX4）的稳定性会降低。但是，SPX降级的机制仍不清楚。在这项研究中，使用酵母双杂交筛选，我们确定了两个由SP饥饿引起的转录后诱导的调节SPX4降解的RING手指泛素E3连接酶。我们发现两个SDEL都位于细胞核和细胞质中，具有泛素E3连接酶活性，并直接泛素化了K ²¹³和KSPX4中的^299个赖氨酸残基可调节其稳定性。此外，我们发现水稻中的Pi中央调节剂PHR2通过在Pi充足的条件下与SPX4相互作用，可以与SDEL竞争，从而保护SPX4免受泛素化和降解。与SDEL1和SDEL2的生化功能一致，SDEL1或SDEL2的过表达即使在Pi充足的条件下，也会导致Pi过度积累并诱导Pi饥饿信号。相反，其功能丧失的突变体显示出减少的Pi积累和减少的Pi饥饿信号传导。总体而言，我们的研究表明，SDEL1和SDEL2促进SPX4的降解，以调节PHR2活性并调节Pi稳态和Pi信号传导，从而响应水稻中外部Pi的有效性。