Life Sciences Commons^™

Csf Protein Changes Associated With Hippocampal Sclerosis Risk Gene Variants Highlight Impact Of Grn/Pgrn, David W. Fardo, Yuriko Katsumata, John S. K. Kauwe, Yuetiva Deming, Oscar Harari, Carlos Cruchaga, Alzheimer’S Disease Neuroimaging Initiative, Peter T. Nelson

Sanders-Brown Center on Aging Faculty Publications

Objective—Hippocampal sclerosis of aging (HS-Aging) is a common cause of dementia in older adults. We tested the variability in cerebrospinal fluid (CSF) proteins associated with previously identified HS-Aging risk single nucleotide polymorphisms (SNPs).

Methods—Alzheimer’s Disease Neuroimaging Initiative cohort (ADNI; n=237) data, combining both multiplexed proteomics CSF and genotype data, were used to assess the association between CSF analytes and risk SNPs in four genes (SNPs): GRN (rs5848), TMEM106B (rs1990622), ABCC9 (rs704180), and KCNMB2 (rs9637454). For controls, non-HS-Aging SNPs in APOE (rs429358/rs7412) and MAPT (rs8070723) were also analyzed against Aβ1-42 and total tau CSF analytes.

Results—The GRN risk …

Stress-Induced Retrotranslocation Of Clusterin/Apoj Into The Cytosol, P Nizard, Suzanne Tetley, Y Le Drean, T Watrin, P Le Goff, Mark R. Wilson, Denis Michel

Mark R Wilson

Clusterin is a usually secreted glycoprotein with chaperone properties. Recently, it has been suggested that clusterin isoforms reside in the nuclear and cytosolic compartments of human cell types, where they can influence various cellular programs including DNA repair, transcription and apoptosis. Several mechanisms have been proposed to explain this atypical location, including alternative transcription initiation and alternative splicing. However none of these have been unequivocally established as occurring in live cells. Here we provide direct experimental evidence that in live intact cells, under certain stress conditions, clusterin can evade the secretion pathway and reach the cytosol. This was demonstrated using …

Clusterin Facilitates In Vivo Clearance Of Extracellular Misfolded Proteins, Amy R. Wyatt, Justin J. Yerbury, Paula Berghofer, I Greguric, Andrew Katsifis, Christopher Dobson, Mark R. Wilson

Mark R Wilson

The extracellular deposition of misfolded proteins is a characteristic of many debilitating age-related disorders. However, little is known about the specific mechanisms that act to suppress this process in vivo. Clusterin (CLU) is an extracellular chaperone that forms stable and soluble complexes with misfolded client proteins. Here we explore the fate of complexes formed between CLU and misfolded proteins both in vitro and in a living organism. We show that proteins injected into rats are cleared more rapidly from circulation when complexed with CLU as a result of their more efficient localisation to the liver and that this clearance is …

Clusterin Interacts With Paclitaxel And Confer Paclitaxel Resistance In Ovarian Cancer, Dong Choon Park, Seung Geun Yeo, Mark R. Wilson, Justin J. Yerbury, Joseph Kwong, William R. Welch, Yang Kyu Choi, Michael J. Birrer, Samuel C. Mok, Kwong-Kwok Wong

Mark R Wilson

Optimal debulking followed by chemotherapy is the standard treatment of managing late-stage ovarian cancer, but chemoresistance is still a major problem. In this study, we compared expression profiles of primary tumor tissue from five long-term (>8 years) and five short-term (years) ovarian cancer survivors and identified clusterin as one of the genes that were significantly up-regulated in short-term survivors. We then evaluated the prognostic significance of clusterin and its possible correlation with chemoresistance in ovarian cancer by immunohistostaining of clusterin in 62 tumor samples from patients with stage III, high-grade serous ovarian cancer. After adjusting for debulking status and …

Structural Characterization Of Clusterin-Chaperone Client Protein Complexes, Amy Wyatt, Justin Yerbury, Mark Wilson

Mark R Wilson

Clusterin (CLU) is a potent extracellular chaperone that inhibits protein aggregation and precipitation otherwise caused by physical or chemical stresses (e.g. heat, reduction). This action involves CLU forming soluble high molecular weight (HMW) complexes with the client protein. Other than their unquantified large size, the physical characteristics of these complexes were previously unknown. In this study, HMW CLU-citrate synthase (CS), HMW CLU-fibrinogen (FGN), and HMW CLU-glutathione S-transferase (GST) complexes were generated in vitro, and their structures studied using size exclusion chromatography (SEC), ELISA, SDS-PAGE, dynamic light scattering (DLS), bisANS fluorescence, and circular dichroism spectrophotometry (CD). Densitometry of …

Identification Of Human Plasma Proteins As Major Clients For The Extracellular Chaperone Clusterin, Amy R. Wyatt, Mark R. Wilson

Mark R Wilson

Clusterin (CLU) is an extracellular chaperone that is likely to play an important role in protein folding quality control. This study identified three deposition disease-associated proteins as major plasma clients for clusterin by studying CLU-client complexes formed in response to physiologically relevant stress (shear stress, similar to 36 dynes/cm(2) at 37 degrees C). Analysis of plasma samples by size exclusion chromatography indicated that (i) relative to control plasma, stressed plasma contained proportionally more soluble protein species of high molecular weight, and (ii) high molecular weight species were far more abundant when proteins purified by anti-CLU immunoaffinity chromatography from stressed plasma …

Effects Of Glycosylation On The Structure And Function Of The Extracellular Chaperone Clusterin, Elise Stewart, Andrew Aquilina, Simon B Easterbrook-Smith, D Murphy-Durland, C Jacobsen, S Moestrup, Mark Wilson

J. A. Aquilina

Clusterin is the first well characterized, constitutively secreted extracellular chaperone that binds to exposed regions of hydrophobicity on non-native proteins. It may help control the folding state of extracellular proteins by targeting them for receptor-mediated endocytosis and intracellular lysosomal degradation. A notable feature of secreted clusterin is its heavy glycosylation. Although carbohydrate comprises approximately 20−25% of the total mass of the mature molecule, its function is unknown. Results from the current study demonstrate that deglycosylation of human serum clusterin had little effect on its overall secondary structure content but produced a small increase in solvent-exposed hydrophobicity and enhanced the propensity …

The Extracellular Chaperone Clusterin Sequesters Oligomeric Forms Of The Amyloid-Beta 1-40 Peptide, Priyanka Narayan, Angel Orte, Richard W. Clarke, Benedetta Bolognesi, Sharon Hook, Kristina A. Ganzinger, Sarah Meehan, Mark R. Wilson, Christopher M. Dobson, David Klenerman

Faculty of Science - Papers (Archive)

In recent genome-wide association studies, the extracellular chaperone protein, clusterin, has been identified as a newly-discovered risk factor in Alzheimer's disease. We have examined the interactions between human clusterin and the Alzheimer's disease-associated amyloid-β _1-40 peptide (Aβ _1-40), which is prone to aggregate into an ensemble of oligomeric intermediates implicated in both the proliferation of amyloid fibrils and in neuronal toxicity. Using highly sensitive single-molecule fluorescence methods, we have found that Aβ _1-40 forms a heterogeneous distribution of small oligomers (from dimers to 50-mers), all of which interact with clusterin to form long-lived, stable complexes. Consequently, clusterin is able …

The Extracellular Chaperone Clusterin Sequesters Oligomeric Forms Of The Amyloid-Beta 1-40 Peptide, Priyanka Narayan, Angel Orte, Richard Clarke, Benedetta Bolognesi, Sharon Hook, Kristina Ganzinger, Sarah Meehan, Mark Wilson, Christopher Dobson, David Klenerman

Mark R Wilson

In recent genome-wide association studies, the extracellular chaperone protein, clusterin, has been identified as a newly-discovered risk factor in Alzheimer's disease. We have examined the interactions between human clusterin and the Alzheimer's disease-associated amyloid-β 1-40 peptide (Aβ 1-40), which is prone to aggregate into an ensemble of oligomeric intermediates implicated in both the proliferation of amyloid fibrils and in neuronal toxicity. Using highly sensitive single-molecule fluorescence methods, we have found that Aβ 1-40 forms a heterogeneous distribution of small oligomers (from dimers to 50-mers), all of which interact with clusterin to form long-lived, stable complexes. Consequently, clusterin is able to …

Clusterin Facilitates In Vivo Clearance Of Extracellular Misfolded Proteins, Amy R. Wyatt, Justin J. Yerbury, Paula Berghofer, I Greguric, Andrew Katsifis, Christopher Dobson, Mark R. Wilson

Faculty of Science - Papers (Archive)

The extracellular deposition of misfolded proteins is a characteristic of many debilitating age-related disorders. However, little is known about the specific mechanisms that act to suppress this process in vivo. Clusterin (CLU) is an extracellular chaperone that forms stable and soluble complexes with misfolded client proteins. Here we explore the fate of complexes formed between CLU and misfolded proteins both in vitro and in a living organism. We show that proteins injected into rats are cleared more rapidly from circulation when complexed with CLU as a result of their more efficient localisation to the liver and that this clearance …

Identification Of Human Plasma Proteins As Major Clients For The Extracellular Chaperone Clusterin, Amy R. Wyatt, Mark R. Wilson

Faculty of Science - Papers (Archive)

Clusterin (CLU) is an extracellular chaperone that is likely to play an important role in protein folding quality control. This study identified three deposition disease-associated proteins as major plasma clients for clusterin by studying CLU-client complexes formed in response to physiologically relevant stress (shear stress, similar to 36 dynes/cm(2) at 37 degrees C). Analysis of plasma samples by size exclusion chromatography indicated that (i) relative to control plasma, stressed plasma contained proportionally more soluble protein species of high molecular weight, and (ii) high molecular weight species were far more abundant when proteins purified by anti-CLU immunoaffinity chromatography from stressed plasma …

Structural Characterization Of Clusterin-Chaperone Client Protein Complexes, Amy R. Wyatt, Justin J. Yerbury, Mark R. Wilson

Faculty of Science - Papers (Archive)

Clusterin (CLU) is a potent extracellular chaperone that inhibits protein aggregation and precipitation otherwise caused by physical or chemical stresses (e.g. heat, reduction). This action involves CLU forming soluble high molecular weight (HMW) complexes with the client protein. Other than their unquantified large size, the physical characteristics of these complexes were previously unknown. In this study, HMW CLU-citrate synthase (CS), HMW CLU-fibrinogen (FGN), and HMW CLU-glutathione S-transferase (GST) complexes were generated in vitro, and their structures studied using size exclusion chromatography (SEC), ELISA, SDS-PAGE, dynamic light scattering (DLS), bisANS fluorescence, and circular dichroism spectrophotometry (CD). Densitometry of …

The Chaperone Action Of Clusterin And Its Putative Role In Quality Control Of Extracellular Protein Folding, Amy Wyatt, Justin Yerbury, Stephen Poon, Rebecca Dabbs, Mark Wilson

Mark R Wilson

The function(s) of clusterin may depend upon its topological location. A variety of intracellular "isoforms" of clusterin have been reported but further work is required to better define their identity. The secreted form of clusterin has a potent ability to inhibit both amorphous and amyloid protein aggregation. In the case of amorphous protein aggregation, clusterin forms stable, soluble high-molecular-weight complexes with misfolded client proteins. Clusterin expression is increased during many types of physiological and pathological stresses and is thought to function as an extracellular chaperone (EC). The pathology of a variety of serious human diseases is thought to arise as …

Clusterin Interacts With Paclitaxel And Confer Paclitaxel Resistance In Ovarian Cancer, Dong Choon Park, Seung Geun Yeo, Mark R. Wilson, Justin J. Yerbury, Joseph Kwong, William R. Welch, Yang Kyu Choi, Michael J. Birrer, Samuel C. Mok, Kwong-Kwok Wong

Faculty of Science - Papers (Archive)

Optimal debulking followed by chemotherapy is the standard treatment of managing late-stage ovarian cancer, but chemoresistance is still a major problem. In this study, we compared expression profiles of primary tumor tissue from five long-term (>8 years) and five short-term (years) ovarian cancer survivors and identified clusterin as one of the genes that were significantly up-regulated in short-term survivors. We then evaluated the prognostic significance of clusterin and its possible correlation with chemoresistance in ovarian cancer by immunohistostaining of clusterin in 62 tumor samples from patients with stage III, high-grade serous ovarian cancer. After adjusting for debulking status and …

Stress-Induced Retrotranslocation Of Clusterin/Apoj Into The Cytosol, P Nizard, Suzanne Tetley, Y Le Drean, T Watrin, P Le Goff, Mark R. Wilson, Denis Michel

Faculty of Science - Papers (Archive)

Clusterin is a usually secreted glycoprotein with chaperone properties. Recently, it has been suggested that clusterin isoforms reside in the nuclear and cytosolic compartments of human cell types, where they can influence various cellular programs including DNA repair, transcription and apoptosis. Several mechanisms have been proposed to explain this atypical location, including alternative transcription initiation and alternative splicing. However none of these have been unequivocally established as occurring in live cells. Here we provide direct experimental evidence that in live intact cells, under certain stress conditions, clusterin can evade the secretion pathway and reach the cytosol. This was demonstrated using …

The Extracellular Chaperone Clusterin Influences Amyloid Formation And Toxicity By Interacting With Pre-Fibrillar Structures, Justin Yerbury, Stephen Poon, Sarah Meehan, Brianna Thompson, Janet Kumita, Christopher Dobson, Mark Wilson

Mark R Wilson

Clusterin is an extracellular chaperone present in all disease-associated extracellular amyloid deposits, however, its roles in amyloid formation and protein deposition in vivo are poorly understood. The current study initially aimed to characterise the effects of clusterin on amyloid formation in vitro by a panel of eight protein substrates. Two of the substrates (Alzheimer's beta peptide and a PI3-SH3 domain) were then used in further experiments to examine the effects of clusterin on amyloid cytotoxicity and to probe the mechanism of clusterin action. We show that clusterin exerts potent effects on amyloid formation, the nature and extent of which vary …

The Extracellular Chaperone Clusterin Potently Inhibits Human Lysozyme Amyloid Formation By Interacting With Prefibrillar Species, Mark Wilson, Justin Yerbury, Stephen Poon, Christopher Dobson, C V Robinson, Elise Stewart, Janet Kumita, Mireille Dumoulin, Gemma Caddy, Christine Hagan

Mark R Wilson

We have studied the effects of the extracellular molecular chaperone, clusterin, on the in vitro aggregation of mutational variants of human lysozyme, including one associated with familial amyloid disease. The aggregation of the amyloidogenic variant I56T is inhibited significantly at clusterin-to-lysozyme ratios as low as 1:80 (i.e. one clusterin molecule per 80 lysozyme molecules). Experiments indicate that under the conditions where inhibition of aggregation occurs, clusterin does not bind detectably to the native or fibrillar states, or to the monomeric transient intermediate known to be a key species in the aggregation reaction. Rather, it seems to interact with oligomeric species …

Effects Of Glycosylation On The Structure And Function Of The Extracellular Chaperone Clusterin, Elise Stewart, Andrew Aquilina, Simon B Easterbrook-Smith, D Murphy-Durland, C Jacobsen, S Moestrup, Mark Wilson

Mark R Wilson

Clusterin is the first well characterized, constitutively secreted extracellular chaperone that binds to exposed regions of hydrophobicity on non-native proteins. It may help control the folding state of extracellular proteins by targeting them for receptor-mediated endocytosis and intracellular lysosomal degradation. A notable feature of secreted clusterin is its heavy glycosylation. Although carbohydrate comprises approximately 20−25% of the total mass of the mature molecule, its function is unknown. Results from the current study demonstrate that deglycosylation of human serum clusterin had little effect on its overall secondary structure content but produced a small increase in solvent-exposed hydrophobicity and enhanced the propensity …

The Acute Phase Protein Haptoglobin Is A Mammalian Extracellular Chaperone With An Action Similar To Clusterin, Justin Yerbury, Mark S Rybchyn, Simon B Easterbrook-Smith, C. Henriques, Mark Wilson

Mark R Wilson

Haptoglobin (Hp) is an acidic glycoprotein present in most body fluids of humans and other mammals. Although the functions of Hp are not yet fully understood, the available evidence indicates that it is likely to play an important role in suppressing inflammatory responses. Some earlier work suggested that Hp might be a newly identified member of a small group of extracellular chaperones found at significant levels in human body fluids. Previously, the only well-characterized member of this group was clusterin, which shares functional similarities with the small heat-shock proteins. We report here that Hp specifically inhibited the precipitation of a …