Binding of the periplakin linker requires vimentin acidic residues D176 and E187

Abstract

Plakin proteins form connections that link the cell membrane to the intermediate filament cytoskeleton. Their interactions are mediated by a highly conserved linker domain through an unresolved mechanism. Here analysis of the human periplakin linker domain structure reveals a bi-lobed module transected by an electropositive groove. Key basic residues within the periplakin groove are vital for co-localization with vimentin in human cells and compromise direct binding which also requires acidic residues D176 and E187 in vimentin. We propose a model whereby basic periplakin linker domain residues recognize acidic vimentin side chains and form a complementary binding groove. The model is shared amongst diverse linker domains and can be used to investigate the effects of pathogenic mutations in the desmoplakin linker associated with arrhythmogenic right ventricular cardiomyopathy. Linker modules either act solely or collaborate with adjacent plakin repeat domains to create strong and adaptable tethering within epithelia and cardiac muscle.

Missense Mutations in Desmoplakin Plakin Repeat Domains Have Dramatic Effects on Domain Structure and Function

“…The value of Manders’ overlap coefficient (MOC) ranges from 0 to 1 with an MOC of 0.5, implying that DSP C in one channel co-localises with 50% of the vimentin in another channel. The wild-type DSP C exhibited an MOC of 0.52, indicating substantial co-localisation with vimentin, in agreement with our previous results [ 9 ] ( Figure 4 B). The MOC values of DSP C encompassing soluble mutations R2366C, R2639Q and K2689T were not significantly different to that of the wild-type protein.…”

“…Individual PRDs are poorly expressed in transfected HeLa cells, necessitating the use of a longer C-terminal DSP C construct consisting of desmoplakin residues T1960-A2822, encompassing all three PRDs and the linker module (but excluding the glycine-serine-arginine-rich (GSR) region) ( Figure S3A ). This construct was previously shown to co-localise with IFs in transfected HeLa cells [ 8 , 9 ]. A band corresponding to the expected molecular weight of the DSP C construct (~95kDa) was observed using Western blotting ( Figure S3B ).…”

“…In addition to its three PRDs, desmoplakin possesses a linker module, located between PRDs B and C. Linker modules encompass two PR-like motifs and are found in several other plakin proteins [ 5 ]. The crystal structure of periplakin linker shows an elongated bi-lobed domain that is transected by an electropositive groove, and modelling suggests that the desmoplakin linker exhibits a similar overall topology [ 7 , 9 ]. Basic linker domain side chains recognise acidic IF rod residues by electrostatic complementary in a similar mechanism to that demonstrated by PRDs [ 9 ].…”

“…The crystal structure of periplakin linker shows an elongated bi-lobed domain that is transected by an electropositive groove, and modelling suggests that the desmoplakin linker exhibits a similar overall topology [ 7 , 9 ]. Basic linker domain side chains recognise acidic IF rod residues by electrostatic complementary in a similar mechanism to that demonstrated by PRDs [ 9 ]. Crystallography and small angle X-ray scattering analysis suggest that the three desmoplakin PRDs and linker form an elongated ‘beads on a string’ structure [ 7 ].…”

PIP₃ phosphatase inositol polyphosphate 5-phosphatase K (INPP5K) connects the endoplasmic reticulum to microtubules and mediates the regulation of endoplasmic reticulum morphology

“…Desmoplakin, periplakin, and envoplakin are members of the plakin family of intermediate filament-binding proteins. Basic amino acid clusters of these proteins recognize acidic side chains of vimentin intermediate filaments [30,31]. Although further binding assays between INPP5K and intermediate filaments are required, it may be possible that INPP5K interacts with microtubules via intermediate filaments.…”