Inhibition of proteolytic and ATPase activities of the proteasome by the BTK inhibitor CGI-1746.

Bruton's tyrosine kinase (BTK) inhibitor, ibrutinib, has been shown to synergize with proteasome inhibitors (PIs) in reducing the viability of cells derived from B cell malignancies, but the mechanism is not known. We report here that an off-target effect of ibrutinib causes synergy because not all BTK inhibitors exhibited the synergistic effect, and those that synergized did so even in cells that do not express BTK. The allosteric BTK inhibitor CGI-1746 showed the strongest synergy.

Analysis of Arc/Arg3.1 Oligomerization In Vitro and in Living Cells.

Arc (also known as Arg3.1) is an activity-dependent immediate early gene product enriched in neuronal dendrites. Arc plays essential roles in long-term potentiation, long-term depression, and synaptic scaling.

Differential Interactions of the Proteasome Inhibitor PI31 with Constitutive and Immuno-20S Proteasomes.

PI31 (roteasome nhibitor of ,000 Da) is a 20S proteasome binding protein originally identified as an in vitro inhibitor of 20S proteasome proteolytic activity. Recently reported cryo-electron microscopy structures of 20S-PI31 complexes have revealed that the natively disordered proline-rich C-terminus of PI31 enters the central chamber in the interior of the 20S proteasome and interacts directly with the proteasome's multiple catalytic threonine residues in a manner predicted to inhibit their enzymatic function while evading its own proteolysis. Higher eukaryotes express an alternative form of the 20S proteasome (termed "immuno-proteasome") that features genetically and functionally distinct catalytic subunits.

Ηigh-resolution structure of mammalian PI31-20S proteasome complex reveals mechanism of proteasome inhibition.

Proteasome-catalyzed protein degradation mediates and regulates critical aspects of many cellular functions and is an important element of proteostasis in health and disease. Proteasome function is determined in part by the types of proteasome holoenzymes formed between the 20S core particle that catalyzes peptide bond hydrolysis and any of multiple regulatory proteins to which it binds. One of these regulators, PI31, was previously identified as an in vitro 20S proteasome inhibitor, but neither the molecular mechanism nor the possible physiologic significance of PI31-mediated proteasome inhibition has been clear.

High-resolution structure of mammalian PI31â€"20S proteasome complex reveals mechanism of proteasome inhibition.

Proteasome-catalyzed protein degradation mediates and regulates critical aspects of many cellular functions and is an important element of proteostasis in health and disease. Proteasome function is determined in part by the types of proteasome holoenzymes formed between the 20S core particle that catalyzes peptide bond hydrolysis and any of multiple regulatory proteins to which it binds. One of these regulators, PI31, was previously identified as an 20S proteasome inhibitor, but neither the molecular mechanism nor the possible physiologic significance of PI31-mediated proteasome inhibition has been clear.

Highly conserved shifts in ubiquitin-proteasome system (UPS) activity drive mitochondrial remodeling during quiescence.

Defects in cellular proteostasis and mitochondrial function drive many aspects of infertility, cancer, and other age-related diseases. All of these conditions rely on quiescent cells, such as oocytes and adult stem cells, that reduce their activity and remain dormant as part of their roles in tissue homeostasis, reproduction, and even cancer recurrence. Using a multi-organism approach, we show that dynamic shifts in the ubiquitin proteasome system drive mitochondrial remodeling during cellular quiescence.

The specter of irreparable ignorance: counterfactuals and causality in economics.

Those economists who have emphasized true uncertainty have tended to draw an epistemic distinction between an ascertainable past and an unknowable future. But in one critical respect-in extracting causal relationships-that epistemic distinction is not warranted. Whether they are situated in the past or future, causal arguments in economics depend equally on counterfactual reasoning.

Palmitoylated Proteins in Dendritic Spine Remodeling.

Activity-responsive changes in the actin cytoskeleton are required for the biogenesis, motility, and remodeling of dendritic spines. These changes are governed by proteins that regulate the polymerization, depolymerization, bundling, and branching of actin filaments. Thus, processes that have been extensively characterized in the context of non-neuronal cell shape change and migration are also critical for learning and memory.

Introduction to the Thematic Minireview Series: Autophagy.

Autophagy is a highly conserved, tightly regulated cellular process that degrades intracellular constituents via lysosomes. Autophagy mediates many normal cellular functions and is dysregulated in numerous diseases. This Thematic Series consists of five Minireviews that highlight selected topics of current autophagy research ranging from the molecular mechanisms and regulation of autophagy to the roles of autophagy in health and disease.

The rate of protein synthesis in hematopoietic stem cells is limited partly by 4E-BPs.

Adult stem cells must limit their rate of protein synthesis, but the underlying mechanisms remain largely unexplored. Differences in protein synthesis among hematopoietic stem cells (HSCs) and progenitor cells did not correlate with differences in proteasome activity, total RNA content, mRNA content, or cell division rate. However, adult HSCs had more hypophosphorylated eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) and 4E-BP2 as compared with most other hematopoietic progenitors.

Molecular and cellular roles of PI31 (PSMF1) protein in regulation of proteasome function.

We investigated molecular features and cellular roles of PI31 (PSMF1) on regulation of proteasome function. PI31 has a C-terminal HbYX (where Hb is a hydrophobic amino acid, Y is tyrosine, and X is any amino acid) motif characteristic of several proteasome activators. Peptides corresponding to the PI31 C terminus also bind to and activate the 20 S proteasome in an HbYX-dependent manner, but intact PI31protein inhibits in vitro 20 S activity.

Multiple autism-linked genes mediate synapse elimination via proteasomal degradation of a synaptic scaffold PSD-95.

The activity-dependent transcription factor myocyte enhancer factor 2 (MEF2) induces excitatory synapse elimination in mouse neurons, which requires fragile X mental retardation protein (FMRP), an RNA-binding protein implicated in human cognitive dysfunction and autism. We report here that protocadherin 10 (Pcdh10), an autism-spectrum disorders gene, is necessary for this process. MEF2 and FMRP cooperatively regulate the expression of Pcdh10.

ATP binding by proteasomal ATPases regulates cellular assembly and substrate-induced functions of the 26 S proteasome.

We examined the role of ATP binding by six different ATPase subunits (Rpt1-6) in the cellular assembly and molecular functions of mammalian 26 S proteasome. Four Rpt subunits (Rpt1-4) with ATP binding mutations were incompetent for cellular assembly into 26 S proteasome. In contrast, analogous mutants of Rpt5 and Rpt6 were incorporated normally into 26 S proteasomes in both intact cells and an in vitro assembly assay.

Reconstitution of PA700, the 19S regulatory particle, from purified precursor complexes.

Here, we describe methodology for the in vitro reconstitution of PA700, the 19S regulatory particle of the 26S proteasome, from three purified subcomplexes that closely represent cellular assembly intermediates. These PA700 subcomplexes (denoted PS-1, PS-2, and PS-3) account for all subunits present in purified PA700 but have no overlapping or non-PA700 components. The reconstituted PA700 displays functional features indistinguishable from independently purified PA700, including ATPase activity, deubiquitylating activity, and ATP-dependent binding and activation of the 20S proteasome.

C termini of proteasomal ATPases play nonequivalent roles in cellular assembly of mammalian 26 S proteasome.

The 26 S proteasome comprises two multisubunit subcomplexes as follows: 20 S proteasome and PA700/19 S regulatory particle. The cellular mechanisms by which these subcomplexes assemble into 26 S proteasome and the molecular determinants that govern the assembly process are poorly defined. Here, we demonstrate the nonequivalent roles of the C termini of six AAA subunits (Rpt1-Rpt6) of PA700 in 26 S proteasome assembly in mammalian cells.

PSMB8 encoding the β5i proteasome subunit is mutated in joint contractures, muscle atrophy, microcytic anemia, and panniculitis-induced lipodystrophy syndrome.

We performed homozygosity mapping in two recently reported pedigrees from Portugal and Mexico with an autosomal-recessive autoinflammatory syndrome characterized by joint contractures, muscle atrophy, microcytic anemia, and panniculitis-induced lipodystrophy (JMP). This revealed only one homozygous region spanning 2.4 Mb (5818 SNPs) on chromosome 6p21 shared by all three affected individuals from both families.

The C terminus of Rpt3, an ATPase subunit of PA700 (19 S) regulatory complex, is essential for 26 S proteasome assembly but not for activation.

PA700, the 19 S regulatory subcomplex of the 26 S proteasome, contains a heterohexameric ring of AAA subunits (Rpt1 to -6) that forms the binding interface with a heteroheptameric ring of α subunits (α1 to -7) of the 20 S proteasome. Binding of these subcomplexes is mediated by interactions of C termini of certain Rpt subunits with cognate binding sites on the 20 S proteasome. Binding of two Rpt subunits (Rpt2 and Rpt5) depends on their last three residues, which share an HbYX motif (where Hb is a hydrophobic amino acid) and open substrate access gates in the center of the α ring.

Accelerated formation of alpha-synuclein oligomers by concerted action of the 20S proteasome and familial Parkinson mutations.

A hallmark of Parkinson disease (PD) is the formation of intracellular protein inclusions called Lewy bodies that also contain mitochondria. alpha-Synuclein (alpha Syn) is a major protein component of Lewy bodies, where it is in an amyloid conformation and a significant fraction is truncated by poorly understood proteolytic events. Previously, we demonstrated that the 20S proteasome cleaves alpha Syn in vitro to produce fragments like those observed in Lewy bodies and that the fragments accelerate the formation of amyloid fibrils from full-length alpha Syn.