The Multiple Functions of the PAQosome: An R2TP- and URI1 Prefoldin-Based Chaperone Complex.

The PAQosome (Particle for Arrangement of Quaternary structure) is a large multisubunit chaperone complex that is essential for the assembly and stabilization of other macromolecular complexes. It also interacts with several chaperones including Hsp90, Hsp70, and CCT. The PAQosome is comprised of the R2TP complex, the URI1 prefoldin complex (also known as the non-canonical prefoldin-like complex), the RNA polymerase subunit RPB5, and the WD40 repeat protein WDR92. The R2TP complex is conserved among eukaryotes and has been comprehensively studied over the last 13 years. The R2TP complex is known for its involvement in the assembly and stabilization of L7Ae ribonucleoproteins, U5 small nuclear ribonucleoprotein, RNA polymerase II, phosphatidylinositol-3-kinase-related proteins (PIKKs), and the tuberous sclerosis complex (TSC1-TSC2). By contrast, the URI1 prefoldin complex has evolved exclusively in higher metazoans. Although the URI1 prefoldin complex was initially reported more than 15 years ago, little is known about its function and its role within the PAQosome. Given that URI1 is overexpressed in many types of cancer, it is surprising that the URI1 prefoldin complex has been overlooked. This chapter provides an update on the recent progress uncovering the physiological roles of each PAQosome subunit and provides an overview of the potential functions of the URI1 prefoldin complex.

The Role of Hsp90-R2TP in Macromolecular Complex Assembly and Stabilization.

Hsp90 is a ubiquitous molecular chaperone involved in many cell signaling pathways, and its interactions with specific chaperones and cochaperones determines which client proteins to fold. Hsp90 has been shown to be involved in the promotion and maintenance of proper protein complex assembly either alone or in association with other chaperones such as the R2TP chaperone complex. Hsp90-R2TP acts through several mechanisms, such as by controlling the transcription of protein complex subunits, stabilizing protein subcomplexes before their incorporation into the entire complex, and by recruiting adaptors that facilitate complex assembly. Despite its many roles in protein complex assembly, detailed mechanisms of how Hsp90-R2TP assembles protein complexes have yet to be determined, with most findings restricted to proteomic analyses and in vitro interactions. This review will discuss our current understanding of the function of Hsp90-R2TP in the assembly, stabilization, and activity of the following seven classes of protein complexes: L7Ae snoRNPs, spliceosome snRNPs, RNA polymerases, PIKKs, MRN, TSC, and axonemal dynein arms.

Identification of Allosteric Inhibitors against Active Caspase-6.

Caspase-6 is a cysteine protease that plays essential roles in programmed cell death, axonal degeneration, and development. The excess neuronal activity of Caspase-6 is associated with Alzheimer disease neuropathology and age-dependent cognitive impairment. Caspase-6 inhibition is a promising strategy to stop early stage neurodegenerative events, yet finding potent and selective Caspase-6 inhibitors has been a challenging task due to the overlapping structural and functional similarities between caspase family members. Here, we investigated how four rare non-synonymous missense single-nucleotide polymorphisms (SNPs), resulting in amino acid substitutions outside human Caspase-6 active site, affect enzyme structure and catalytic efficiency. Three investigated SNPs were found to align with a putative allosteric pocket with low sequence conservation among human caspases. Virtual screening of 57,700 compounds against the putative Caspase-6 allosteric pocket, followed by in vitro testing of the best virtual hits in recombinant human Caspase-6 activity assays identified novel allosteric Caspase-6 inhibitors with IC50 and Ki values ranging from ~2 to 13 µM. This report may pave the way towards the development and optimisation of novel small molecule allosteric Caspase-6 inhibitors and illustrates that functional characterisation of rare natural variants holds promise for the identification of allosteric sites on other therapeutic targets in drug discovery.

Caspase-1 inhibition alleviates cognitive impairment and neuropathology in an Alzheimer's disease mouse model.

Alzheimer's disease (AD) is an intractable progressive neurodegenerative disease characterized by cognitive decline and dementia. An inflammatory neurodegenerative pathway, involving Caspase-1 activation, is associated with human age-dependent cognitive impairment and several classical AD brain pathologies. Here, we show that the nontoxic and blood-brain barrier permeable small molecule Caspase-1 inhibitor VX-765 dose-dependently reverses episodic and spatial memory impairment, and hyperactivity in the J20 mouse model of AD. Cessation of VX-765 results in the reappearance of memory deficits in the mice after 1 month and recommencement of treatment re-establishes normal cognition. VX-765 prevents progressive amyloid beta peptide deposition, reverses brain inflammation, and normalizes synaptophysin protein levels in mouse hippocampus. Consistent with these findings, Caspase-1 null J20 mice are protected from episodic and spatial memory deficits, neuroinflammation and Aß accumulation. These results provide in vivo proof of concept for Caspase-1 inhibition against AD cognitive deficits and pathologies.