A barley SKP1-like protein controls abundance of the susceptibility factor RACB and influences the interaction of barley with the barley powdery mildew fungus

Abstract

In an increasing number of plant-microbe interactions, it has become evident that the abundance of immunity-related proteins is controlled by the ubiquitin-26S proteasome system. In the interaction of barley with the biotrophic barley powdery mildew fungus Blumeria graminis f.sp. hordei (Bgh), the RAC/ROP [RAT SARCOMA-related C3 botulinum toxin substrate/RAT SARCOMA HOMOLOGUE (RHO) of plants] guanosine triphosphatase (GTPase) HvRACB supports the fungus in a compatible interaction. By contrast, barley HvRBK1, a ROP-binding receptor-like cytoplasmic kinase that interacts with and can be activated by constitutively activated HvRACB, limits fungal infection success. We have identified a barley type II S-phase kinase 1-associated (SKP1)-like protein (HvSKP1-like) as a molecular interactor of HvRBK1. SKP1 proteins are subunits of the SKP1-cullin 1-F-box (SCF)-E3 ubiquitin ligase complex that acts in the specific recognition and ubiquitination of protein substrates for subsequent proteasomal degradation. Transient induced gene silencing of either HvSKP1-like or HvRBK1 increased protein abundance of constitutively activated HvRACB in barley epidermal cells, whereas abundance of dominant negative RACB only weakly increased. In addition, silencing of HvSKP1-like enhanced the susceptibility of barley to haustorium establishment by Bgh. In summary, our results suggest that HvSKP1-like, together with HvRBK1, controls the abundance of HvRACB and, at the same time, modulates the outcome of the barley-Bgh interaction. A possible feedback mechanism from RAC/ROP-activated HvRBK1 on the susceptibility factor HvRACB is discussed.