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. Author manuscript; available in PMC: 2015 Mar 13.
Published in final edited form as: Cell. 2014 Mar 13;156(6):1193–1206. doi: 10.1016/j.cell.2014.02.008

Figure 5. Structure-based Mutations Disrupts ASCPYD Filament Formation, AIM2PYD/ASCPYD Interaction and NLRP3PYD/ASCPYD Interaction in Vitro and in Cells.

Figure 5

A. Size-exclusion chromatography of WT and mutant ASCPYD showing both filamentous (void) and monomeric fractions from a Superdex 200 column. Hyphen denotes ASCPYD and asterisk denotes a contaminant.

B. Morphology of transfected WT and mutant eGFP-tagged ASCPYD constructs visualized by confocal laser scanning microscopy. The arrowhead depicts filaments. n: nucleus; scale bars = 10µm.

C.Morphology of transfected eGFP-tagged ASCPYD visualized by confocal laser scanning microscopy. Top: ASCPYD with charge reversal mutation on a residue outside the filament interface. Bottom: ASCPYD with triple charge reversal mutation that rescued the defectiveness of the single mutants. Arrow heads depict filaments.

D.A schematic model of AIM2PYD/ASCPYD or NLRP3PYD/ASCPYD filaments composed of a top AIM2PYD or NLRP3PYD layer extended by ASCPYD filament body.

E,F. Mutations of conserved interfacial residues on AIM2PYD (E) and NLRP3PYD-NBD (F) reduced or abolished their ability to nucleate ASCPYD filaments.

See also Figure S5.