The RhoGAP SPIN6 Associates with SPL11 and OsRac1 and Negatively Regulates Programmed Cell Death and Innate Immunity in Rice

Abstract

The ubiquitin proteasome system in plants plays important roles in plant-microbe interactions and in immune responses to pathogens. We previously demonstrated that the rice U-box E3 ligase SPL11 and its Arabidopsis ortholog PUB13 negatively regulate programmed cell death (PCD) and defense response. However, the components involved in the SPL11/PUB13-mediated PCD and immune signaling pathway remain unknown. In this study, we report that SPL11-interacting Protein 6 (SPIN6) is a Rho GTPase-activating protein (RhoGAP) that interacts with SPL11 in vitro and in vivo. SPL11 ubiquitinates SPIN6 in vitro and degrades SPIN6 in vivo via the 26S proteasome-dependent pathway. Both RNAi silencing in transgenic rice and knockout of Spin6 in a T-DNA insertion mutant lead to PCD and increased resistance to the rice blast pathogen Magnaporthe oryzae and the bacterial blight pathogen Xanthomonas oryzae pv. oryzae. The levels of reactive oxygen species and defense-related gene expression are significantly elevated in both the Spin6 RNAi and mutant plants. Strikingly, SPIN6 interacts with the small GTPase OsRac1, catalyze the GTP-bound OsRac1 into the GDP-bound state in vitro and has GAP activity towards OsRac1 in rice cells. Together, our results demonstrate that the RhoGAP SPIN6 acts as a linkage between a U-box E3 ligase-mediated ubiquitination pathway and a small GTPase-associated defensome system for plant immunity.

The Monocot-Specific Receptor-like Kinase SDS2 Controls Cell Death and Immunity in Rice

“…OsRLCK185 phosphorylates MAPKKKε and MAPKKK18 to initiate rice immunity (Wang et al, 2017; Yamada et al, 2017). In our previous study, we reported that SPL11 interacts with SPIN6, which negatively regulates the activity of OsRac1 (Liu et al, 2015). In this study, we found that SDS2 interacts with and phosphorylates SPL11, which in turn ubiquitinates SDS2, leading to its degradation.…”

“…SPL11 , encoding a plant U-box (PUB) type E3 ubiquitin ligase, negatively regulates PCD and innate immunity, and the rice spl11 mutant is lesion mimic (Zeng et al, 2004). SPL11 interacts with the Rho GTPase-activating protein (RhoGAP) SPL11-INTERACTING PROTEIN 6 (SPIN6), which negatively regulates the Rho GTPase OsRac1, a central regulator of rice PTI and ETI signaling (Kawano and Shimamoto, 2013; Liu et al, 2015). Many RLKs are substrates of PUB proteins, and their interactions have been well studied.…”

Chitin and chitin-related compounds in plant–fungal interactions

“…In the situation of defence responses to pathogenic fungi, LysM RLK1 (Wan et al 2008) or CERK1 (Miya et al 2007)/CERK1 (as in Arabidopsis ) or CERK1/CEBiP receptor dimers (as in rice, Shimizu et al 2010) needs longer chito-oligomers, like octamers to activate its response through heterodimerisation (Hayafune et al 2014; Liu et al 2015). The perception of shorter oligosaccharides such as chitotetraose could instead rely on monomeric receptors (Miyata et al 2014; Shinya et al 2015) as into each receptor’s LysM domain, half of this oligomer molecule (C4) fits (Liu et al 2012; Shinya et al 2015).…”

“…The interaction between SPIN6 and OsRac1 may lead to the change of OsRac1 from the GTP state to the GDP state, which reduces the active form of OsRac1 in rice cells. Mutation in the Spin6 gene may cause accumulation of ROS and PR proteins, such as PR1a, PR5 and PBZ1, that results in plant cell death and immunity (Liu et al 2015). …”

Homologous U-box E3 Ubiquitin Ligases OsPUB2 and OsPUB3 Are Involved in the Positive Regulation of Low Temperature Stress Response in Rice (Oryza sativa L.)

“…For example, SPL11 is known to ubiquitinate Rho GTPase-activating protein (RhoGAP) SPIN6 and negatively regulate innate immunity in rice (Zeng et al, 2004; Liu et al, 2015). OsPUB15 is involved in reducing cellular oxidative stress during seedling establishment (Park et al, 2011).…”