A direct role for SNX9 in the biogenesis of filopodia

Abstract

Filopodia are finger-like actin-rich protrusions that extend from the cell surface and are important for cell–cell communication and pathogen internalization. The small size and transient nature of filopodia combined with shared usage of actin regulators within cells confounds attempts to identify filopodial proteins. Here, we used phage display phenotypic screening to isolate antibodies that alter the actin morphology of filopodia-like structures (FLS) in vitro. We found that all of the antibodies that cause shorter FLS interact with SNX9, an actin regulator that binds phosphoinositides during endocytosis and at invadopodia. In cells, we discover SNX9 at specialized filopodia in Xenopus development and that SNX9 is an endogenous component of filopodia that are hijacked by Chlamydia entry. We show the use of antibody technology to identify proteins used in filopodia-like structures, and a role for SNX9 in filopodia.

Chlamydia trachomatis TmeA Directly Activates N-WASP To Promote Actin Polymerization and Functions Synergistically with TarP during Invasion

“…Based on electron microscopy data, Faris et al (48) indicated a requirement of TmeA in filopodia formation. This observation would be consistent with robust induction of surface structures induced by high-multiplicity of infection (MOI) chlamydial infection ( 9), but it is unclear how this fits with data indicating that Chlamydia hijacks existing macropinocytosis filopodia instead of inducing the de novo assembly of the structures (57). Perhaps functionally distinct protrusions are being manifested when Chlamydia associates with host cells.…”

Surfing on Membrane Waves: Microvilli, Curved Membranes, and Immune Signaling

“…In a cell-free system for reconstituting actin bundles of filopodia (80), immunodepletion of SNX9 resulted in shorter actin bundles. Moreover, SNX9 localizes to the filopodial tip and shaft in RPE-1 cells (79). The positive role of SNX9 in filopodia formation is dependent on its activity in stimulating N-WASp and Arp2/3 (81), which probably overrides the curvature-sensing function of the N-BAR domain.…”

“…Interestingly, IRSp53 is also expressed in T cells (78), raising its possible role in regulating microvilli formation. Surprisingly, the N-BAR protein sorting nexin 9 (SNX9), which is expected to recognize positive curvatures, is involved the biogenesis of filopodia (79). In a cell-free system for reconstituting actin bundles of filopodia (80), immunodepletion of SNX9 resulted in shorter actin bundles.…”

Adding SNX to the mix: SNX9 drives filopodia biogenesis

“…At steady-state, the actin within filopodia remains dynamic, as filaments undergo treadmilling, and filopodial lengths are modulated by the balance between actin assembly at the tip and disassembly at the base. Building upon this basic understanding of filopodial initiation, growth, and maintenance, Jarsch et l. (3) probed deeper into the mechanisms of filopodia biogenesis by integrating a powerful in vitro filopodial assembly platform with a phage display-based functional screening system.…”

“…This biomimetic system allows for actin polymerization into structures that reflect the organization of filopodia and that contain the core molecular constituents responsible for their assembly in cells (5). In the current paper (3), Jarsch et al sought to functionally dissect the requirements for FLS biogenesis through the addition of a library of phage displaying single-chain antibodies (6). After initially subtracting phage that recognized several known filopodial components, they selected for phage that bound to FLSs, and used a subset of the subsequently cloned/purified antibodies to screen for alterations in FLS formation.…”