The ARP2/3 complex, acting cooperatively with Class I formins, modulates penetration resistance in Arabidopsis against powdery mildew invasion

Abstract

The actin cytoskeleton regulates an array of diverse cellular activities that support the establishment of plant-microbe interactions and plays a critical role in the execution of plant immunity. However, molecular and cellular mechanisms regulating the assembly and rearrangement of actin filaments at plant-pathogen interaction sites remain largely elusive. Here, using live-cell imaging, we show that one of the earliest cellular responses in Arabidopsis thaliana upon powdery mildew attack is the formation of patch-like actin filament structures beneath fungal invasion sites. The actin filaments constituting actin patches undergo rapid turnover, which is regulated by the actin-related protein (ARP)2/3 complex and its activator, the WAVE/SCAR regulatory complex (W/SRC). The focal accumulation of phosphatidylinositol-4,5-bisphosphate at fungal penetration sites appears to be a crucial upstream modulator of the W/SRC-ARP2/3 pathway-mediated actin patch formation. Knockout of W/SRC-ARP2/3 pathway subunits partially compromised penetration resistance with impaired endocytic recycling of the defense-associated t-SNARE protein PEN1 and its deposition into apoplastic papillae. Simultaneously knocking out ARP3 and knocking down the Class I formin (AtFH1) abolished actin patch formation, severely impaired the deposition of cell wall appositions, and promoted powdery mildew entry into host cells. Our results demonstrate that the ARP2/3 complex and formins, two actin-nucleating systems, act cooperatively and contribute to Arabidopsis penetration resistance to fungal invasion.

Controlling the Gate: The Functions of the Cytoskeleton in Stomatal Movement

“…PI3P binding inhibits SCAB1 oligomerization, which further impairs AF destabilization and reorganization during ABA-induced stomatal closure ( Yang et al, 2021 ). Several members of other ABP families, such as Villins and ADFs, as well as the upstream regulator of ARP2/3 complex, the WAVE/SCAR complex ( Xiang et al, 2007 ; Zhao et al, 2010 ; Qin et al, 2021 ), bind to and are regulated by phospholipids. These interactions pose the possibility that phospholipids may regulate stomatal movement via multiple mechanism.…”

Lipid Signaling Requires ROS Production to Elicit Actin Cytoskeleton Remodeling during Plant Innate Immunity

“…A key question in this field is why host-cell actin filaments assemble quickly following perception of microbial invaders. One proposed model is that a rearranged actin cytoskeletal network provides tracks for the delivery of defense materials; this is supported by observations of an actin patch as well as actin cables focusing on the penetration site during pathogenic and non-pathogenic fungal and oomycete interactions [ 46 , 50 , 51 , 52 ]. These actin-based structures facilitate the deposition of callose, which fortifies the plant cell wall and helps abrogate pathogen invasion [ 19 , 20 , 53 ].…”

“…Using a powerful combination of high spatiotemporal resolution imaging and genetic tools, we and others have demonstrated that turnover of single actin filaments is precisely regulated through nucleation, severing, and availability of actin filament plus ends [ 22 , 25 , 58 , 60 , 61 ]. The accumulation of actin filaments during PTI could be contributed through increased actin nucleation by the Arp2/3 complex or formins [ 52 , 62 ] or through inhibition of filament severing by ADF4 [ 19 ]. Heterodimeric capping protein (CP) limits actin filament assembly by preventing addition of monomers to plus ends and preventing filament–filament annealing [ 24 , 25 ] and has been implicated in multiple signaling pathways [ 20 , 21 , 25 , 37 ].…”

Cooperative actin filament nucleation by the Arp2/3 complex and formins maintains the homeostatic cortical array in Arabidopsis epidermal cells

“…The cortical cytoplasm of epidermal cells contains a dynamic actin array that is rather disordered and comprises comingled actin filament bundles and individual actin filaments (Staiger et al, 2009; Smertenko et al, 2010). Evidence for actin patches or dense dendritic actin networks is limited to certain cell types, such as guard mother cells (Facette et al, 2015), the apex of trichomes (Yanagisawa et al, 2015), or at focal sites elicited by pathogen attack (Hardham et al, 2007; Qin et al, 2021). Our results revealed that the density of actin filaments in the cortical array, extent of filament bundling, as well as filament nucleation frequency all decreased when either the Arp2/3 complex or formins were inhibited, suggesting that formins were not able to maintain the actin abundance at normal levels when the Arp2/3 complex was genetically- or chemically-inhibited, and the Arp2/3 complex also failed to do so when formins were inhibited.…”

“…In growing trichome branches, for example, the Arp2/3 complex generates a tip-localized actin filament array constrained by cortical microtubules that coordinates growth (Yanagisawa et al, 2018; Yanagisawa et al, 2015). During fungal invasion, the Arp2/3 complex localizes to the site of penetration peg attack in epidermal pavement cells and generates a large cortical actin filament patch necessary to suppress invasion (Qin et al, 2021). However, the molecular mechanisms by which the Arp2/3 complex coordinates the organization of cortical actin filament arrays in unstimulated cells or how it generates unique arrays in response to biotic and abiotic stress are poorly understood.…”