The Proteasome and Ageing.

The proteasome is a multi-subunit proteolytic complex that functions to degrade normal proteins for physiological regulation and to eliminate abnormal proteins for cellular protection. Generally, the proteasome targets substrate proteins that are marked by attachment of multiple ubiquitin molecules. In various types of cells in an organism, damage to proteins occurs both from internal sources such as reactive oxygen species and from external ones such as UV radiation from the sun. The proteasome functions to protect the cells by degrading damaged proteins. With ageing, however, the capacity of the proteasome to degrade damaged proteins is reduced as indicated by evidence gathered by many studies. Studies on ageing in muscle, skin, and brain show that with age catalytic activity of the proteasome is decreased and the expression of proteasome subunits is altered. Age-related accumulation of damaged or misfolded proteins causes further reduction of proteasome activity. Abnormal proteins also accumulate as a result of age-related neurodegenerative diseases. Deficits in proteasome activity might be responsible for accumulation of protein aggregates and thus contribute to the pathology. Results from several studies suggest a link between the proteasome and longevity. This chapter reviews the various ways in which the proteasome is associated with the ageing process and examines evidence gathered from investigations on cultured cells, model organisms, and humans.

Degradation of Transcriptional Repressor ATF4 during Long-Term Synaptic Plasticity.

Maintenance of long-term synaptic plasticity requires gene expression mediated by cAMP-responsive element binding protein (CREB). Gene expression driven by CREB can commence only if the inhibition by a transcriptional repressor activating transcription factor 4 (ATF4; also known as CREB2) is relieved. Previous research showed that the removal of ATF4 occurs through ubiquitin-proteasome-mediated proteolysis. Using chemically induced hippocampal long-term potentiation (cLTP) as a model system, we investigate the mechanisms that control ATF4 degradation. We observed that ATF4 phosphorylated at serine-219 increases upon induction of cLTP and decreases about 30 min thereafter. Proteasome inhibitor ß-lactone prevents the decrease in ATF4. We found that the phosphorylation of ATF4 is mediated by cAMP-dependent protein kinase. Our initial experiments towards the identification of the ligase that mediates ubiquitination of ATF4 revealed a possible role for ß-transducin repeat containing protein (ß-TrCP). Regulation of ATF4 degradation is likely to be a mechanism for determining the threshold for gene expression underlying maintenance of long-term synaptic plasticity and by extension, long-term memory.

Recent developments in transcriptional and translational regulation underlying long-term synaptic plasticity and memory.

Formation of long-term synaptic plasticity that underlies long-term memory requires new protein synthesis. Years of research has elucidated some of the transcriptional and translational mechanisms that contribute to the production of new proteins. Early research on transcription focused on the transcription factor cAMP-responsive element binding protein. Since then, other transcription factors, such as the Nuclear Receptor 4 family of proteins that play a role in memory formation and maintenance have been identified. In addition, several studies have revealed details of epigenetic mechanisms consisting of new types of chemical alterations of DNA such as hydroxymethylation, and various histone modifications in long-term synaptic plasticity and memory. Our understanding of translational control critical for memory formation began with the identification of molecules that impinge on the 5' and 3' untranslated regions of mRNAs and continued with the appreciation for local translation near synaptic sites. Lately, a role for noncoding RNAs such as microRNAs in regulating translation factors and other molecules critical for memory has been found. This review describes the past research in brief and mainly focuses on the recent work on molecular mechanisms of transcriptional and translational regulation that form the underpinnings of long-term synaptic plasticity and memory.

Proteolysis, synaptic plasticity and memory.

Protein degradation has many critical functions in the nervous system such as refinement of synaptic connections during development and synaptic plasticity and memory in the adult organisms. A major cellular machinery of proteolysis is the ubiquitin-proteasome pathway (UPP). The UPP precisely regulates proteolysis by covalently attaching ubiquitin, a small protein, to substrates through sequential enzymatic reactions and the proteins marked with the ubiquitin tag are degraded by complex containing many subunits called the proteasome. Research over the years has shown a role for the UPP in regulating presynaptic and postsynaptic proteins critical for neurotransmission and synaptic plasticity. Studies have also revealed a role for the UPP in various forms of memory. Mechanistic investigations suggest that the function of the UPP in neurons is not homogenous and is subject to local regulation in different neuronal sub-compartments. In both invertebrate and vertebrate model systems, local roles have been found for enzymes that attach ubiquitin to substrate proteins as well as for enzymes that remove ubiquitin from substrates. The proteasome also has disparate functions in different parts of the neuron. In addition to the UPP, proteolysis by the lysosome and autophagy play a role in synaptic plasticity and memory. This review details the functions of proteolysis in synaptic plasticity and summarizes the findings on the connection between proteolysis and memory mainly focusing on the UPP including its local roles.

Proteasome modulates positive and negative translational regulators in long-term synaptic plasticity.

Proteolysis by the ubiquitin-proteasome pathway appears to have a complex role in synaptic plasticity, but its various functions remain to be elucidated. Using late phase long-term potentiation (L-LTP) in the hippocampus of the mouse as a model for long-term synaptic plasticity, we previously showed that inhibition of the proteasome enhances induction but blocks maintenance of L-LTP. In this study, we investigated the possible mechanisms by which proteasome inhibition has opposite effects on L-LTP induction and maintenance. Our results show that inhibiting phosphatidyl inositol-3 kinase or blocking the interaction between eukaryotic initiation factors 4E (eIF4E) and 4G (eIF4G) reduces the enhancement of L-LTP induction brought about by proteasome inhibition suggesting interplay between proteolysis and the signaling pathway mediated by mammalian target of rapamycin (mTOR). Also, proteasome inhibition leads to accumulation of translational activators in the mTOR pathway such as eIF4E and eukaryotic elongation factor 1A (eEF1A) early during L-LTP causing increased induction. Furthermore, inhibition of the proteasome causes a buildup of translational repressors, such as polyadenylate-binding protein interacting protein 2 (Paip2) and eukaryotic initiation factor 4E-binding protein 2 (4E-BP2), during late stages of L-LTP contributing to the blockade of L-LTP maintenance. Thus, the proteasome plays a critical role in regulating protein synthesis during L-LTP by tightly controlling translation. Our results provide novel mechanistic insights into the interplay between protein degradation and protein synthesis in long-term synaptic plasticity.

Role of ubiquitin-proteasome-mediated proteolysis in nervous system disease.

Proteolysis by the ubiquitin-proteasome pathway (UPP) is now widely recognized as a molecular mechanism controlling myriad normal functions in the nervous system. Also, this pathway is intimately linked to many diseases and disorders of the brain. Among the diseases connected to the UPP are neurodegenerative disorders such as Alzheimer's, Parkinson's and Huntington's diseases. Perturbation in the UPP is also believed to play a causative role in mental disorders such as Angelman syndrome. The pathology of neurodegenerative diseases is characterized by abnormal deposition of insoluble protein aggregates or inclusion bodies within neurons. The ubiquitinated protein aggregates are believed to result from dysfunction of the UPP or from structural changes in the protein substrates which prevent their recognition and degradation by the UPP. An early effect of abnormal UPP in diseases of the nervous system is likely to be impairment of synaptic function. Here we discuss the UPP and its physiological roles in the nervous system and how alterations in the UPP relate to development of nervous system diseases. This article is part of a Special Issue entitled The 26S Proteasome: When degradation is just not enough!

The ubiquitin-proteasome pathway and synaptic plasticity.

Proteolysis by the ubiquitin-proteasome pathway (UPP) has emerged as a new molecular mechanism that controls wide-ranging functions in the nervous system, including fine-tuning of synaptic connections during development and synaptic plasticity in the adult organism. In the UPP, attachment of a small protein, ubiquitin, tags the substrates for degradation by a multisubunit complex called the proteasome. Linkage of ubiquitin to protein substrates is highly specific and occurs through a series of well-orchestrated enzymatic steps. The UPP regulates neurotransmitter receptors, protein kinases, synaptic proteins, transcription factors, and other molecules critical for synaptic plasticity. Accumulating evidence indicates that the operation of the UPP in neurons is not homogeneous and is subject to tightly managed local regulation in different neuronal subcompartments. Investigations on both invertebrate and vertebrate model systems have revealed local roles for enzymes that attach ubiquitin to substrate proteins, as well as for enzymes that remove ubiquitin from substrates. The proteasome also has been shown to possess disparate functions in different parts of the neuron. Here I give a broad overview of the role of the UPP in synaptic plasticity and highlight the local roles and regulation of the proteolytic pathway in neurons.

The ubiquitin-proteasome pathway in health and disease of the nervous system.

In recent years, proteolysis by the ubiquitin-proteasome pathway has attained prominence as a new molecular mechanism that regulates many vital functions of the nervous system, including development of synaptic connections and synaptic plasticity. Here, we review the latest findings on the role of proteolysis in sculpting the nervous system through control of axonal growth, axonal and dendritic pruning, and regulation of synaptic size and number. We also discuss how protein degradation functions in synaptic plasticity and the roles of local proteolysis in neuronal compartments. In addition, we describe how proteolysis is associated with Alzheimer's disease and ataxia. Furthermore, we highlight the recent approaches that exploit components of the ubiquitin-proteasome pathway for amelioration of these diseases in animal models.

Proteasome inhibition enhances the induction and impairs the maintenance of late-phase long-term potentiation.

Protein degradation by the ubiquitin-proteasome pathway plays important roles in synaptic plasticity, but the molecular mechanisms by which proteolysis regulates synaptic strength are not well understood. We investigated the role of the proteasome in hippocampal late-phase long-term potentiation (L-LTP), a model for enduring synaptic plasticity. We show here that inhibition of the proteasome enhances the induction of L-LTP, but inhibits its maintenance. Proteasome inhibitor-mediated enhancement of the early part of L-LTP requires activation of NMDA receptors and the cAMP-dependent protein kinase. Augmentation of L-LTP induction by proteasome inhibition is blocked by a protein synthesis inhibitor anisomycin and is sensitive to the drug rapamycin. Our findings indicate that proteasome inhibition increases the induction of L-LTP by stabilizing locally translated proteins in dendrites. In addition, our data show that inhibition of the proteasome blocks transcription of brain-derived neurotrophic factor (BDNF), which is a cAMP-responsive element-binding protein (CREB)-inducible gene. Furthermore, our results demonstrate that the proteasome inhibitors block degradation of ATF4, a CREB repressor. Thus, proteasome inhibition appears to hinder CREB-mediated transcription. Our results indicate that blockade of proteasome activity obstructs the maintenance of L-LTP by interfering with transcription as well as translation required to sustain L-LTP. Thus, proteasome-mediated proteolysis has different roles during the induction and the maintenance of L-LTP.

A potential proteasome-interacting motif within the ubiquitin-like domain of parkin and other proteins.

Parkin and other unrelated proteins contain a ubiquitin-like domain (UbLD). This article describes a motif that might be important in the interaction of UbLD-containing proteins (UbLPs) with the proteasome. The proteasome-interacting motif, which is conserved in a subset of UbLPs, such as parkin, Rad23 and several transcription factors, is likely to enable the UbLPs to form a complex with the proteasome for proteolysis or the recently discovered non-proteolytic functions of the proteasome.

Perturbations of Ubiquitin-Proteasome-Mediated Proteolysis in Aging and Alzheimer's Disease.

The ubiquitin-proteasome pathway (UPP) has multiple roles in the normal nervous system, including the development of synaptic connections and synaptic plasticity. Research over the past several years has indicated a role for the UPP in aging without any overt pathology in the brain. In addition, malfunction of the UPP is implicated in Alzheimer's disease (AD) and dementia associated with it. In this mini review article, we assess the literature on the role of protein degradation by the UPP in aging and in AD with special emphasis on dysregulation of the UPP and its contribution to cognitive decline and impairment.

Age-dependent changes in brain hydration and synaptic plasticity.

Aging in humans and animals is associated with gradual and variable changes in some cognitive functions, but what causes them and explains individual variations remains unclear. Hydration decreases with aging but whether dehydration contributes to cognitive dysfunction is not known. The brain hydration of aging mice was determined by colloidosmotic-pressure titration. Dehydration increased with age from ∼76 mmHg at 6 weeks to ∼105 mmHg at 40 weeks, or a progressive ∼10 percent loss of brain water but seemed to level off afterward. When we adjusted dehydration in hippocampal slices of <8-week-old mice to the levels seen in mice 40 weeks and older, their basal synaptic responses were amplified at all stimulus voltages tested, but induction of late-phase long-term potentiation was impaired. Our results document progressive brain dehydration with age in inbred mice to levels at which in vitro synaptic plasticity appears dysregulated. They also suggest that dehydration contributes to some of the changes in synaptic plasticity observed with aging, possibly due to adjustments in neuronal excitation mechanisms.

Proteasome limits plasticity-related signaling to the nucleus in the hippocampus.

Proteolysis by the ubiquitin-proteasome pathway has pleiotropic effects on both induction and maintenance of long-term synaptic plasticity. In this study, we examined the effect of proteasome inhibition on signaling to the nucleus during late-phase long-term potentiation. When a subthreshold L-LTP induction protocol was used, proteasome inhibition led to a significant increase in phosphorylated CREB (pCREB) in the nucleus. Inhibitors of cAMP-dependent protein kinase/protein kinase A, extracellular signal-regulated kinase and cGMP-dependent protein kinase/protein kinase G all blocked the proteasome-inhibition-mediated increase in nuclear pCREB after subthreshold stimulation. These results lay the groundwork for understanding a novel role for the proteasome in limiting signaling to the nucleus in the absence of adequate synaptic stimulation.

The ubiquitin proteasome system functions as an inhibitory constraint on synaptic strengthening.

BACKGROUND: Long-lasting forms of synaptic plasticity have been shown to depend on changes in gene expression. Although many studies have focused on the regulation of transcription and translation during learning-related synaptic plasticity, regulated protein degradation provides another common means of altering the macromolecular composition of cells. RESULTS: We have investigated the role of the ubiquitin proteasome system in long-lasting forms of learning-related plasticity in Aplysia sensory-motor synapses. We find that inhibition of the proteasome produces a long-lasting (24 hr) increase in synaptic strength between sensory and motor neurons and that it dramatically enhances serotonin-induced long-term facilitation. The increase in synaptic strength produced by proteasome inhibitors is dependent on translation but not transcription. In addition to the increase in synaptic strength, proteasome inhibition leads to an increase in the number of synaptic contacts formed between the sensory and motor neurons. Blockade of the proteasome in isolated postsynaptic motor neurons produces an increase in the glutamate-evoked postsynaptic potential, and blockade of the proteasome in the isolated presynaptic sensory cells produces increases in neurite length and branching. CONCLUSIONS: We conclude that both pre- and postsynaptic substrates of the ubiquitin proteasome function constitutively to regulate synaptic strength and growth and that the ubiquitin proteasome pathway functions in mature neurons as an inhibitory constraint on synaptic strengthening.

Full length cloning and expression analysis of splice variants of regulator of G-protein signaling RGS4 in human and murine brain.

RGS4 (regulator of G protein signaling 4) protein is a GTPase-activating protein specific for Gi/o and Gq alpha subunits. It is highly expressed in brain but the mechanisms by which RGS4 expression is regulated remain unknown. RGS4 is associated with schizophrenia either through heritable genetic polymorphisms or as a co-regulated mediator of the pathology, and may play a role in other brain diseases. As a necessary step towards understanding the transcriptional regulation of RGS4, we isolated full-length splice variants of the human RGS4 and mouse Rgs4 gene using bioinformatic predictions, followed by RACE, RT-PCR, and sequencing. In human brain, we found five different isoforms RGS4-1, RGS4-2, RGS4-3, RGS4-4 and RGS4-5 of which RGS4-2, RGS4-3, RGS4-4 and RGS4-5 are novel. RGS4-1 and 2 encode a 205-amino acid protein, while RGS4-3 encodes a 302 aa protein with an N-terminal extension. RGS4-4 and RGS4-5 encode truncated proteins of 93 aa and 187 aa respectively. Our results indicate that RGS4-1, RGS4-2, RGS4-3 and RGS4-4 are translated into proteins. In contrast, the mouse brain has 3 different splice variants, Rgs4-1, Rgs4-2 and Rgs4-3 which encode the same 205 aa protein but vary in their 3'UTRs. Among the mouse isoforms, Rgs4-1 and Rgs4-3 are novel. Human RGS4 has four different transcription start sites and three different stop sites. We found differential expression of the human isoforms in dorsolateral prefrontal and visual cortex. All five RGS4 splice variants are expressed at high levels in human cortical areas although RGS4 isoforms 1, 2, and 3 are not expressed in the cerebellum. RGS4-2 is tissue-specific whereas RGS4-4 and RGS4-5 appear to be ubiquitously expressed. Our results suggest the intriguing possibility that RGS4 gene expression in the human brain is spatially and temporally regulated through differential transcription of isoforms from alternative promoters. This may have implications for the physiological role of RGS4 and in pathologies of the brain.

Role of the ubiquitin proteasome system in Alzheimer's disease.

Though Alzheimer's disease (AD) is a syndrome with well-defined clinical and neuropathological manifestations, an array of molecular defects underlies its pathology. A role for the ubiquitin proteasome system (UPS) was suspected in the pathogenesis of AD since the presence of ubiquitin immunoreactivity in AD-related neuronal inclusions, such as neurofibrillary tangles, is seen in all AD cases. Recent studies have indicated that components of the UPS could be linked to the early phase of AD, which is marked by synaptic dysfunction, as well as to the late stages of the disease, characterized by neurodegeneration. Insoluble protein aggregates in the brain of AD patients could result from malfunction or overload of the UPS, or from structural changes in the protein substrates, which prevent their recognition and degradation by the UPS. Defective proteolysis could cause the synaptic dysfunction observed early in AD since the UPS is known to play a role in the normal functioning of synapses. In this review, we discuss recent observations on possible links between the UPS and AD, and the potential for utilizing UPS components as targets for treatment of this disease. Publication history: Republished from Current BioData's Targeted Proteins database (TPdb; http://www.targetedproteinsdb.com).

MHC molecules in the vomeronasal organ: contributors to pheromonal discrimination?

The vomeronasal organ of the accessory olfactory system detects pheromones in several vertebrate species. Recent studies of vomeronasal sensory neurons have shown that they express MHC molecules, which in the immune system help to discriminate self antigens from non-self antigens. These new findings, along with past research demonstrating MHC-based olfactory discrimination, suggest the exciting possibility that MHC molecules together with vomeronasal G-protein-coupled receptors play a role in distinguishing related individuals from unrelated ones based on pheromonal cues.

Differential regulation of proteasome activity in the nucleus and the synaptic terminals.

Proteasome is a multi-subunit proteolytic complex that degrades proteins covalently linked to multiple molecules of ubiquitin. Earlier studies showed a role for the ubiquitin-proteasome pathway in several models of long-term memory and other forms of synaptic plasticity. In Aplysia, the ubiquitin-proteasome pathway has been shown to contribute to the induction of long-term facilitation. In other model systems, ubiquitin-proteasome-mediated proteolysis has also been shown to play a role in synapse development. Previous studies of synaptic plasticity focused on changes in components or the substrates of the ubiquitin-proteasome pathway in whole neurons. Modification of specific synapses would require precise spatial and temporal regulation of the components of the ubiquitin-proteasome pathway within the subcellular compartments of neurons during learning. As a first step towards testing the idea of local regulation of the ubiquitin-proteasome pathway in neurons, we investigated proteasome activity in nuclear and synaptosomal fractions. Here we show that proteasome activity in the synaptic terminals is higher compared to the activity in the nucleus in the Aplysia nervous system as well as in the mouse brain. Furthermore, the proteasome activity in the two neuronal compartments is differentially modulated by protein kinases. Differential regulation of proteasome activity in neuronal compartments such as the synaptic terminals is likely to be a key mechanism underlying synapse-specific plasticity.

Ubiquitin-proteasome-mediated local protein degradation and synaptic plasticity.

A proteolytic pathway in which attachment of a small protein, ubiquitin, marks the substrates for degradation by a multi-subunit complex called the proteasome has been shown to function in synaptic plasticity and in several other physiological processes of the nervous system. Attachment of ubiquitin to protein substrates occurs through a series of highly specific and regulated steps. Degradation by the proteasome is subject to multiple levels of regulation as well. How does the ubiquitin-proteasome pathway contribute to synaptic plasticity? Long-lasting, protein synthesis-dependent, changes in the synaptic strength occur through activation of molecular cascades in the nucleus in coordination with signaling events in specific synapses. Available evidence indicates that ubiquitin-proteasome-mediated degradation has a role in the molecular mechanisms underlying synaptic plasticity that operate in the nucleus as well as at the synapse. Since the ubiquitin-proteasome pathway has been shown to be versatile in having roles in addition to proteolysis in several other cellular processes relevant to synaptic plasticity, such as endocytosis and transcription, this pathway is highly suited for a localized role in the neuron. Because of its numerous roles, malfunctioning of this pathway leads to several diseases and disorders of the nervous system. In this review, I examine the ubiquitin-proteasome pathway in detail and describe the role of regulated proteolysis in long-term synaptic plasticity. Also, using synaptic tagging theory of synapse-specific plasticity, I provide a model on the possible roles and regulation of local protein degradation by the ubiquitin-proteasome pathway.

The proteasome and epigenetics: zooming in on histone modifications.

The proteasome is a structural complex of many proteins that degrades substrates marked by covalent linkage to ubiquitin. Many years of research has shown a role for ubiquitin-proteasome-mediated proteolysis in synaptic plasticity and memory mainly in degrading synaptic, cytoplasmic and nuclear proteins. Recent work indicates that the proteasome has wider proteolytic and non-proteolytic roles in processes such as histone modifications that affect synaptic plasticity and memory. In this review, we assess the evidence gathered from neuronal as well as non-neuronal cell types regarding the function of the proteasome in positive or negative regulation of posttranslational modifications of histones, such as acetylation, methylation and ubiquitination. We discuss the critical roles of the proteasome in clearing excess histone proteins in various cellular contexts and the possible non-proteolytic functions in regulating transcription of target genes. In addition, we summarize the current literature on diverse chromatin-remodeling machineries, such as histone acetyltransferases, deacetylates, methyltransferases and demethylases, as targets for proteasomal degradation across experimental models. Lastly, we provide a perspective on how proteasomal regulation of histone modifications may modulate synaptic plasticity in the nervous system.