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V Crippa - One of the best experts on this subject based on the ideXlab platform.
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the role of hspb8 a component of the chaperone assisted selective autophagy machinery in cancer
Cells, 2021Co-Authors: R Cristofani, V Crippa, B Tedesco, A. Poletti, M Piccolella, Marina Montagnani Marelli, Roberta M MorettiAbstract:The cellular response to cancer-induced stress is one of the major aspects regulating cancer development and progression. The Heat Shock Protein B8 (HSPB8) is a small chaperone involved in chaperone-assisted selective autophagy (CASA). CASA promotes the selective degradation of proteins to counteract cell stress such as tumor-induced stress. HSPB8 is also involved in (i) the cell division machinery regulating chromosome segregation and cell cycle arrest in the G0/G1 phase and (ii) inflammation regulating dendritic cell maturation and cytokine production. HSPB8 expression and role are tumor-specific, showing a dual and opposite role. Interestingly, HSPB8 may be involved in the acquisition of chemoresistance to drugs. Despite the fact the mechanisms of HSPB8-mediated CASA activation in tumors need further studies, HSPB8 could represent an important factor in cancer induction and progression and it may be a potential target for anticancer treatment in specific types of cancer. In this review, we will discuss the molecular mechanism underlying HSPB8 roles in normal and cancer conditions. The basic mechanisms involved in anti- and pro-tumoral activities of HSPB8 are deeply discussed together with the pathways that modulate HSPB8 expression, in order to outline molecules with a beneficial effect for cancer cell growth, migration, and death.
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Trehalose induces autophagy via lysosomal-mediated TFEB activation in models of motoneuron degeneration
Autophagy, 2018Co-Authors: P Rusmini, Katia Cortese, V Crippa, R Cristofani, M E Cicardi, V Ferrari, G Vezzoli, B Tedesco, M Meroni, Elio MessiAbstract:Macroautophagy/autophagy, a defense mechanism against aberrant stresses, in neurons counteracts aggregate-prone misfolded protein toxicity. Autophagy induction might be beneficial in neurodegenerative diseases (NDs). The natural compound trehalose promotes autophagy via TFEB (transcription factor EB), ameliorating disease phenotype in multiple ND models, but its mechanism is still obscure. We demonstrated that trehalose regulates autophagy by inducing rapid and transient lysosomal enlargement and membrane permeabilization (LMP). This effect correlated with the calcium-dependent phosphatase PPP3/calcineurin activation, TFEB dephosphorylation and nuclear translocation. Trehalose upregulated genes for the TFEB target and regulator Ppargc1a, lysosomal hydrolases and membrane proteins (Ctsb, Gla, Lamp2a, Mcoln1, Tpp1) and several autophagy-related components (Becn1, Atg10, Atg12, Sqstm1/p62, Map1lc3b, Hspb8 and Bag3) mostly in a PPP3- and TFEB-dependent manner. TFEB silencing counteracted the trehalose pro-degradative activity on misfolded protein causative of motoneuron diseases. Similar effects were exerted by trehalase-resistant trehalose analogs, melibiose and lactulose. Thus, limited lysosomal damage might induce autophagy, perhaps as a compensatory mechanism, a process that is beneficial to counteract neurodegeneration. Abbreviations: ALS: amyotrophic lateral sclerosis; AR: androgen receptor; ATG: autophagy related; AV: autophagic vacuole; BAG3: BCL2-associated athanogene 3; BECN1: beclin 1, autophagy related; CASA: chaperone-assisted selective autophagy; CTSB: cathepsin b; DAPI: 4',6-diamidino-2-phenylindole; DMEM: Dulbecco's modified Eagle's medium; EGFP: enhanced green fluorescent protein; fALS, familial amyotrophic lateral sclerosis; FRA: filter retardation assay; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GLA: galactosidase, alpha; HD: Huntington disease; hIPSCs: human induced pluripotent stem cells; HSPA8: heat shock protein A8; HSPB8: heat shock protein B8; IF: immunofluorescence analysis; LAMP1: lysosomal-associated membrane protein 1; LAMP2A: lysosomal-associated membrane protein 2A; LGALS3: lectin, galactose binding, soluble 3; LLOMe: L-leucyl-L-leucine methyl ester; LMP: lysosomal membrane permeabilization; Lys: lysosomes; MAP1LC3B: microtubule-associated protein 1 light chain 3 beta; MCOLN1: mucolipin 1; mRNA: messenger RNA; MTOR: mechanistic target of rapamycin kinase; NDs: neurodegenerative diseases; NSC34: neuroblastoma x spinal cord 34; PBS: phosphate-buffered saline; PD: Parkinson disease; polyQ: polyglutamine; PPARGC1A: peroxisome proliferative activated receptor, gamma, coactivator 1 alpha; PPP3CB: protein phosphatase 3, catalytic subunit, beta isoform; RT-qPCR: real-time quantitative polymerase chain reaction; SBMA: spinal and bulbar muscular atrophy; SCAs: spinocerebellar ataxias; siRNA: small interfering RNA; SLC2A8: solute carrier family 2, (facilitated glucose transporter), member 8; smNPCs: small molecules neural progenitors cells; SOD1: superoxide dismutase 1; SQSTM1/p62: sequestosome 1; STED: stimulated emission depletion; STUB1: STIP1 homology and U-box containing protein 1; TARDBP/TDP-43: TAR DNA binding protein; TFEB: transcription factor EB; TPP1: tripeptidyl peptidase I; TREH: trehalase (brush-border membrane glycoprotein); WB: western blotting; ZKSCAN3: zinc finger with KRAB and SCAN domains 3.
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The small heat shock protein B8 at the interplay between the intracellular degradative pathways in motoneuron disease
2018Co-Authors: A. Poletti, P Rusmini, V Crippa, R Cristofani, M E Cicardi, V Ferrari, B Tedesco, M Meroni, M. Galbiati, S. CerraAbstract:Motor neuron diseases (MNDs) are a large class of neurodegenerative diseases in which upper cortical and lower spinal cord motorneuron are affected. Several MNDs, like spinal and bulbar muscular atrophy (SBMA) or amyotrophic lateral sclerosis (ALS) have been linked to specific gene mutation which results in the production of aberrant proteins prone to misfold and to aggregate. To counteract the accumulation of these misfolded proteins cells, including neurons have developed a finely tuned protein quality control (PQC). The degradative systems are major components of the PQC system and comprise the ubiquitin-proteasome pathway and the autophagic pathways, specifically involved in the maintenance of a normal cell proteostasts. By working in association to salacted molecular chaperones, the degradative systems are essential components of the intracellular protein quality control (PQC) system. Misfolded proteins are neurotoxic and are recognized by chaperones and deleivered to degradative systems for their clearance. A tightly molecularly regulated equilibrium oversees the final fate of aberrant proteins. This equilibrium is crucial to mantain proteostasis in neurons and may chance under various stimuli, and its alteration give rise to a vicious cycle of protein accumulation and to the PQC system damage that might lead to cell death. Among chaperones, the small heat shock protein (HSP) B8, is able to facilitate autophagy and assist the removal of misfolded proteins prone to aggregate in MNDs. HSPB8 associates to BAG3, HSP70 and CHIP (an ubiquitinating enzyme) to deliver misfolded protein to autophagosome and this form of autophagy is called Chaperone-assisted selective autophagy (CASA) and the complex based on HSPB8/BAG3 s named CASA complex. HSPB8 is induced in response to se several neuronal stresses such has proteotoxic and oxidative stresses. We found that HSPB8 is highly induced in two main targets of misfolded protein toxicity in tg mice models of SBMA and ALS, the motorneurons and the muscle. The pharmacological or genetic induction of HSPB8 expression is protective in MNDs, while its siliencing has opposite effects. We also showed that HSPB8 protects from a misfolded protein induced abberant phenotype in fly models of ALS. By increasing HSPB8-mediated selective targeting of misfolded protein to autophagy neurons and muscle reduce their proteasome-mediated clearance limiting its possible overwhelming. Therefore, pharmacological approached which potentiate the HSPB8-BAG3 autophagic pathway could contribute to maintain a correct proteinostasis in motorneuron and muscle cells and might have therapeutic implication in MNDs
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The small heat shock protein B8 (HSPB8) modulates proliferation and migration of breast cancer cells
'Impact Journals LLC', 2017Co-Authors: M Piccolella, P Rusmini, V Crippa, R Cristofani, M E Cicardi, M Meroni, M. Galbiati, N. Ferri, F.f. Morelli, S. CarraAbstract:Breast cancer (BC) is one of the major causes of cancer death in women and is closely related to hormonal dysregulation. Estrogen receptor (ER)-positive BCs are generally treated with anti hormone therapy using antiestrogens or aromatase inhibitors. However, BC cells may become resistant to endocrine therapy, a process facilitated by autophagy, which may either promote or suppress tumor expansion. The autophagy facilitator HSPB8 has been found overexpressed in some BC. Here we found that HSPB8 is highly expressed and differentially modulated by natural or synthetic selective ER modulators (SERMs), in the triple-positive hormone-sensitive BC (MCF-7) cells, but not in triple-negative MDA-MB-231 BC cells. Specific SERMs induced MCF-7 cells proliferation in a HSPB8 dependent manner whereas, did not modify MDA-MB-231 cell growth. ER expression was unaffected in HSPB8-depleted MCF-7 cells. HSPB8 over-expression did not alter the distribution of MCF-7 cells in the various phases of the cell cycle. Conversely and intriguingly, HSPB8 downregulation resulted in an increased number of cells resting in the G0/G1 phase, thus possibly reducing the ability of the cells to pass through the restriction point. In addition, HSPB8 downregulation reduced the migratory ability of MCF-7 cells. None of these modifications were observed, when another small HSP (HSPB1), also expressed in MCF-7 cells, was downregulated. In conclusion, our data suggest that HSPB8 is involved in the mechanisms that regulate cell cycle and cell migration in MCF-7 cells
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transcriptional induction of the heat shock protein b8 mediates the clearance of misfolded proteins responsible for motor neuron diseases
Scientific Reports, 2016Co-Authors: V Crippa, P Rusmini, R Cristofani, M E Cicardi, Elio Messi, M Piccolella, Vito Giuseppe Dagostino, Rosa Loffredo, Michael Pancher, M. GalbiatiAbstract:Neurodegenerative diseases (NDs) are often associated with the presence of misfolded protein inclusions. The chaperone HSPB8 is upregulated in mice, the human brain and muscle structures affected during NDs progression. HSPB8 exerts a potent pro-degradative activity on several misfolded proteins responsible for familial NDs forms. Here, we demonstrated that HSPB8 also counteracts accumulation of aberrantly localized misfolded forms of TDP-43 and its 25 KDa fragment involved in most sporadic cases of Amyotrophic Lateral Sclerosis (sALS) and of Fronto Lateral Temporal Dementia (FLTD). HSPB8 acts with BAG3 and the HSP70/HSC70-CHIP complex enhancing the autophagic removal of misfolded proteins. We performed a high-through put screening (HTS) to find small molecules capable of inducing HSPB8 in neurons for therapeutic purposes. We identified two compounds, colchicine and doxorubicin, that robustly up-regulated HSPB8 expression. Both colchicine and doxorubicin increased the expression of the master regulator of autophagy TFEB, the autophagy linker p62/SQSTM1 and the autophagosome component LC3. In line, both drugs counteracted the accumulation of TDP-43 and TDP-25 misfolded species responsible for motoneuronal death in sALS. Thus, analogs of colchicine and doxorubicin able to induce HSPB8 and with better safety and tolerability may result beneficial in NDs models.
M. Galbiati - One of the best experts on this subject based on the ideXlab platform.
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The small heat shock protein B8 at the interplay between the intracellular degradative pathways in motoneuron disease
2018Co-Authors: A. Poletti, P Rusmini, V Crippa, R Cristofani, M E Cicardi, V Ferrari, B Tedesco, M Meroni, M. Galbiati, S. CerraAbstract:Motor neuron diseases (MNDs) are a large class of neurodegenerative diseases in which upper cortical and lower spinal cord motorneuron are affected. Several MNDs, like spinal and bulbar muscular atrophy (SBMA) or amyotrophic lateral sclerosis (ALS) have been linked to specific gene mutation which results in the production of aberrant proteins prone to misfold and to aggregate. To counteract the accumulation of these misfolded proteins cells, including neurons have developed a finely tuned protein quality control (PQC). The degradative systems are major components of the PQC system and comprise the ubiquitin-proteasome pathway and the autophagic pathways, specifically involved in the maintenance of a normal cell proteostasts. By working in association to salacted molecular chaperones, the degradative systems are essential components of the intracellular protein quality control (PQC) system. Misfolded proteins are neurotoxic and are recognized by chaperones and deleivered to degradative systems for their clearance. A tightly molecularly regulated equilibrium oversees the final fate of aberrant proteins. This equilibrium is crucial to mantain proteostasis in neurons and may chance under various stimuli, and its alteration give rise to a vicious cycle of protein accumulation and to the PQC system damage that might lead to cell death. Among chaperones, the small heat shock protein (HSP) B8, is able to facilitate autophagy and assist the removal of misfolded proteins prone to aggregate in MNDs. HSPB8 associates to BAG3, HSP70 and CHIP (an ubiquitinating enzyme) to deliver misfolded protein to autophagosome and this form of autophagy is called Chaperone-assisted selective autophagy (CASA) and the complex based on HSPB8/BAG3 s named CASA complex. HSPB8 is induced in response to se several neuronal stresses such has proteotoxic and oxidative stresses. We found that HSPB8 is highly induced in two main targets of misfolded protein toxicity in tg mice models of SBMA and ALS, the motorneurons and the muscle. The pharmacological or genetic induction of HSPB8 expression is protective in MNDs, while its siliencing has opposite effects. We also showed that HSPB8 protects from a misfolded protein induced abberant phenotype in fly models of ALS. By increasing HSPB8-mediated selective targeting of misfolded protein to autophagy neurons and muscle reduce their proteasome-mediated clearance limiting its possible overwhelming. Therefore, pharmacological approached which potentiate the HSPB8-BAG3 autophagic pathway could contribute to maintain a correct proteinostasis in motorneuron and muscle cells and might have therapeutic implication in MNDs
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The small heat shock protein B8 (HSPB8) modulates proliferation and migration of breast cancer cells
'Impact Journals LLC', 2017Co-Authors: M Piccolella, P Rusmini, V Crippa, R Cristofani, M E Cicardi, M Meroni, M. Galbiati, N. Ferri, F.f. Morelli, S. CarraAbstract:Breast cancer (BC) is one of the major causes of cancer death in women and is closely related to hormonal dysregulation. Estrogen receptor (ER)-positive BCs are generally treated with anti hormone therapy using antiestrogens or aromatase inhibitors. However, BC cells may become resistant to endocrine therapy, a process facilitated by autophagy, which may either promote or suppress tumor expansion. The autophagy facilitator HSPB8 has been found overexpressed in some BC. Here we found that HSPB8 is highly expressed and differentially modulated by natural or synthetic selective ER modulators (SERMs), in the triple-positive hormone-sensitive BC (MCF-7) cells, but not in triple-negative MDA-MB-231 BC cells. Specific SERMs induced MCF-7 cells proliferation in a HSPB8 dependent manner whereas, did not modify MDA-MB-231 cell growth. ER expression was unaffected in HSPB8-depleted MCF-7 cells. HSPB8 over-expression did not alter the distribution of MCF-7 cells in the various phases of the cell cycle. Conversely and intriguingly, HSPB8 downregulation resulted in an increased number of cells resting in the G0/G1 phase, thus possibly reducing the ability of the cells to pass through the restriction point. In addition, HSPB8 downregulation reduced the migratory ability of MCF-7 cells. None of these modifications were observed, when another small HSP (HSPB1), also expressed in MCF-7 cells, was downregulated. In conclusion, our data suggest that HSPB8 is involved in the mechanisms that regulate cell cycle and cell migration in MCF-7 cells
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transcriptional induction of the heat shock protein b8 mediates the clearance of misfolded proteins responsible for motor neuron diseases
Scientific Reports, 2016Co-Authors: V Crippa, P Rusmini, R Cristofani, M E Cicardi, Elio Messi, M Piccolella, Vito Giuseppe Dagostino, Rosa Loffredo, Michael Pancher, M. GalbiatiAbstract:Neurodegenerative diseases (NDs) are often associated with the presence of misfolded protein inclusions. The chaperone HSPB8 is upregulated in mice, the human brain and muscle structures affected during NDs progression. HSPB8 exerts a potent pro-degradative activity on several misfolded proteins responsible for familial NDs forms. Here, we demonstrated that HSPB8 also counteracts accumulation of aberrantly localized misfolded forms of TDP-43 and its 25 KDa fragment involved in most sporadic cases of Amyotrophic Lateral Sclerosis (sALS) and of Fronto Lateral Temporal Dementia (FLTD). HSPB8 acts with BAG3 and the HSP70/HSC70-CHIP complex enhancing the autophagic removal of misfolded proteins. We performed a high-through put screening (HTS) to find small molecules capable of inducing HSPB8 in neurons for therapeutic purposes. We identified two compounds, colchicine and doxorubicin, that robustly up-regulated HSPB8 expression. Both colchicine and doxorubicin increased the expression of the master regulator of autophagy TFEB, the autophagy linker p62/SQSTM1 and the autophagosome component LC3. In line, both drugs counteracted the accumulation of TDP-43 and TDP-25 misfolded species responsible for motoneuronal death in sALS. Thus, analogs of colchicine and doxorubicin able to induce HSPB8 and with better safety and tolerability may result beneficial in NDs models.
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transcriptional induction of the heat shock protein b8 mediates the clearance of misfolded proteins responsible for motor neuron diseases
Scientific Reports, 2016Co-Authors: V Crippa, P Rusmini, R Cristofani, M E Cicardi, Elio Messi, M Piccolella, Vito Giuseppe Dagostino, Rosa Loffredo, Michael Pancher, M. GalbiatiAbstract:Neurodegenerative diseases (NDs) are often associated with the presence of misfolded protein inclusions. The chaperone HSPB8 is upregulated in mice, the human brain and muscle structures affected during NDs progression. HSPB8 exerts a potent pro-degradative activity on several misfolded proteins responsible for familial NDs forms. Here, we demonstrated that HSPB8 also counteracts accumulation of aberrantly localized misfolded forms of TDP-43 and its 25 KDa fragment involved in most sporadic cases of Amyotrophic Lateral Sclerosis (sALS) and of Fronto Lateral Temporal Dementia (FLTD). HSPB8 acts with BAG3 and the HSP70/HSC70-CHIP complex enhancing the autophagic removal of misfolded proteins. We performed a high-through put screening (HTS) to find small molecules capable of inducing HSPB8 in neurons for therapeutic purposes. We identified two compounds, colchicine and doxorubicin, that robustly up-regulated HSPB8 expression. Both colchicine and doxorubicin increased the expression of the master regulator of autophagy TFEB, the autophagy linker p62/SQSTM1 and the autophagosome component LC3. In line, both drugs counteracted the accumulation of TDP-43 and TDP-25 misfolded species responsible for motoneuronal death in sALS. Thus, analogs of colchicine and doxorubicin able to induce HSPB8 and with better safety and tolerability may result beneficial in NDs models.
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the small heat shock protein b8 hspb8 promotes autophagic removal of misfolded proteins involved in amyotrophic lateral sclerosis als
Human Molecular Genetics, 2010Co-Authors: V Crippa, P Rusmini, M. Galbiati, A. Boncoraglio, D Sau, Elisa Onesto, E Bolzoni, E Fontana, Marianna Marino, Serena CarraAbstract:Several neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), are characterized by the presence of misfolded proteins, thought to trigger neurotoxicity. Some familial forms of ALS (fALS), clinically indistinguishable from sporadic ALS (sALS), are linked to superoxide dismutase 1 (SOD1) gene mutations. It has been shown that the mutant SOD1 misfolds, forms insoluble aggregates and impairs the proteasome. Using transgenic G93A-SOD1 mice, we found that spinal cord motor neurons, accumulating mutant SOD1 also over-express the small heat shock protein HspB8. Using motor neuronal fALS models, we demonstrated that HspB8 decreases aggregation and increases mutant SOD1 solubility and clearance, without affecting wild-type SOD1 turnover. Notably, HspB8 acts on mutant SOD1 even when the proteasome activity is specifically blocked. The pharmacological blockage of autophagy resulted in a dramatic increase of mutant SOD1 aggregates. Immunoprecipitation studies, performed during autophagic flux blockage, demonstrated that mutant SOD1 interacts with the HspB8/Bag3/Hsc70/CHIP multiheteromeric complex, known to selectively activate autophagic removal of misfolded proteins. Thus, HspB8 increases mutant SOD1 clearance via autophagy. Autophagy activation was also observed in lumbar spinal cord of transgenic G93A-SOD1 mice since several autophago-lysosomal structures were present in affected surviving motor neurons. Finally, we extended our observation to a different ALS model and demonstrated that HspB8 exerts similar effects on a truncated version of TDP-43, another protein involved both in fALS and in sALS. Overall, these results indicate that the pharmacological modulation of HspB8 expression in motor neurons may have important implications to unravel the molecular mechanisms involved both in fALS and in sALS.
Ian R Brown - One of the best experts on this subject based on the ideXlab platform.
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differential targeting of hsp70 heat shock proteins hspa6 and hspa1a with components of a protein disaggregation refolding machine in differentiated human neuronal cells following thermal stress
Frontiers in Neuroscience, 2017Co-Authors: Catherine A S Deane, Ian R BrownAbstract:Heat shock proteins (Hsps) co-operate in multi-protein machines that counter protein misfolding and aggregation and involve DNAJ (Hsp40), HSPA (Hsp70), and HSPH (Hsp105α). The HSPA family is a multigene family composed of inducible and constitutively expressed members. Inducible HSPA6 (Hsp70B') is found in the human genome but not in the genomes of mouse and rat. To advance knowledge of this little studied HSPA member, the targeting of HSPA6 to stress-sensitive neuronal sites with components of a disaggregation/refolding machine was investigated following thermal stress. HSPA6 targeted the periphery of nuclear speckles (perispeckles) that have been characterized as sites of transcription. However, HSPA6 did not co-localize at perispeckles with DNAJB1 (Hsp40-1) or HSPH1 (Hsp105α). At 3 h after heat shock, HSPA6 co-localized with these members of the disaggregation/refolding machine at the granular component (GC) of the nucleolus. Inducible HSPA1A (Hsp70-1) and constitutively expressed HSPA8 (Hsc70) co-localized at nuclear speckles with components of the machine immediately after heat shock, and at the GC layer of the nucleolus at 1 h with DNAJA1 and BAG-1. These results suggest that HSPA6 exhibits targeting features that are not apparent for HSPA1A and HSPA8.
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components of a mammalian protein disaggregation refolding machine are targeted to nuclear speckles following thermal stress in differentiated human neuronal cells
Cell Stress & Chaperones, 2017Co-Authors: Catherine A S Deane, Ian R BrownAbstract:Heat shock proteins (Hsps) are a set of highly conserved proteins involved in cellular repair and protective mechanisms. They counter protein misfolding and aggregation that are characteristic features of neurodegenerative diseases. Hsps act co-operatively in disaggregation/refolding machines that assemble at sites of protein misfolding and aggregation. Members of the DNAJ (Hsp40) family act as "holdases" that detect and bind misfolded proteins, while members of the HSPA (Hsp70) family act as "foldases" that refold proteins to biologically active states. HSPH1 (Hsp105α) is an important additional member of the mammalian disaggregation/refolding machine that acts as a disaggregase to promote the dissociation of aggregated proteins. Components of a disaggregation/refolding machine were targeted to nuclear speckles after thermal stress in differentiated human neuronal SH-SY5Y cells, namely: HSPA1A (Hsp70-1), DNAJB1 (Hsp40-1), DNAJA1 (Hsp40-4), and HSPH1 (Hsp105α). Nuclear speckles are rich in RNA splicing factors, and heat shock disrupts RNA splicing which recovers after stressful stimuli. Interestingly, constitutively expressed HSPA8 (Hsc70) was also targeted to nuclear speckles after heat shock with elements of a disaggregation/refolding machine. Hence, neurons have the potential to rapidly assemble a disaggregation/refolding machine after cellular stress using constitutively expressed Hsc70 without the time lag needed for synthesis of stress-inducible Hsp70. Constitutive Hsc70 is abundant in neurons in the mammalian brain and has been proposed to play a role in pre-protecting neurons from cellular stress.
P Rusmini - One of the best experts on this subject based on the ideXlab platform.
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Trehalose induces autophagy via lysosomal-mediated TFEB activation in models of motoneuron degeneration
Autophagy, 2018Co-Authors: P Rusmini, Katia Cortese, V Crippa, R Cristofani, M E Cicardi, V Ferrari, G Vezzoli, B Tedesco, M Meroni, Elio MessiAbstract:Macroautophagy/autophagy, a defense mechanism against aberrant stresses, in neurons counteracts aggregate-prone misfolded protein toxicity. Autophagy induction might be beneficial in neurodegenerative diseases (NDs). The natural compound trehalose promotes autophagy via TFEB (transcription factor EB), ameliorating disease phenotype in multiple ND models, but its mechanism is still obscure. We demonstrated that trehalose regulates autophagy by inducing rapid and transient lysosomal enlargement and membrane permeabilization (LMP). This effect correlated with the calcium-dependent phosphatase PPP3/calcineurin activation, TFEB dephosphorylation and nuclear translocation. Trehalose upregulated genes for the TFEB target and regulator Ppargc1a, lysosomal hydrolases and membrane proteins (Ctsb, Gla, Lamp2a, Mcoln1, Tpp1) and several autophagy-related components (Becn1, Atg10, Atg12, Sqstm1/p62, Map1lc3b, Hspb8 and Bag3) mostly in a PPP3- and TFEB-dependent manner. TFEB silencing counteracted the trehalose pro-degradative activity on misfolded protein causative of motoneuron diseases. Similar effects were exerted by trehalase-resistant trehalose analogs, melibiose and lactulose. Thus, limited lysosomal damage might induce autophagy, perhaps as a compensatory mechanism, a process that is beneficial to counteract neurodegeneration. Abbreviations: ALS: amyotrophic lateral sclerosis; AR: androgen receptor; ATG: autophagy related; AV: autophagic vacuole; BAG3: BCL2-associated athanogene 3; BECN1: beclin 1, autophagy related; CASA: chaperone-assisted selective autophagy; CTSB: cathepsin b; DAPI: 4',6-diamidino-2-phenylindole; DMEM: Dulbecco's modified Eagle's medium; EGFP: enhanced green fluorescent protein; fALS, familial amyotrophic lateral sclerosis; FRA: filter retardation assay; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GLA: galactosidase, alpha; HD: Huntington disease; hIPSCs: human induced pluripotent stem cells; HSPA8: heat shock protein A8; HSPB8: heat shock protein B8; IF: immunofluorescence analysis; LAMP1: lysosomal-associated membrane protein 1; LAMP2A: lysosomal-associated membrane protein 2A; LGALS3: lectin, galactose binding, soluble 3; LLOMe: L-leucyl-L-leucine methyl ester; LMP: lysosomal membrane permeabilization; Lys: lysosomes; MAP1LC3B: microtubule-associated protein 1 light chain 3 beta; MCOLN1: mucolipin 1; mRNA: messenger RNA; MTOR: mechanistic target of rapamycin kinase; NDs: neurodegenerative diseases; NSC34: neuroblastoma x spinal cord 34; PBS: phosphate-buffered saline; PD: Parkinson disease; polyQ: polyglutamine; PPARGC1A: peroxisome proliferative activated receptor, gamma, coactivator 1 alpha; PPP3CB: protein phosphatase 3, catalytic subunit, beta isoform; RT-qPCR: real-time quantitative polymerase chain reaction; SBMA: spinal and bulbar muscular atrophy; SCAs: spinocerebellar ataxias; siRNA: small interfering RNA; SLC2A8: solute carrier family 2, (facilitated glucose transporter), member 8; smNPCs: small molecules neural progenitors cells; SOD1: superoxide dismutase 1; SQSTM1/p62: sequestosome 1; STED: stimulated emission depletion; STUB1: STIP1 homology and U-box containing protein 1; TARDBP/TDP-43: TAR DNA binding protein; TFEB: transcription factor EB; TPP1: tripeptidyl peptidase I; TREH: trehalase (brush-border membrane glycoprotein); WB: western blotting; ZKSCAN3: zinc finger with KRAB and SCAN domains 3.
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The small heat shock protein B8 at the interplay between the intracellular degradative pathways in motoneuron disease
2018Co-Authors: A. Poletti, P Rusmini, V Crippa, R Cristofani, M E Cicardi, V Ferrari, B Tedesco, M Meroni, M. Galbiati, S. CerraAbstract:Motor neuron diseases (MNDs) are a large class of neurodegenerative diseases in which upper cortical and lower spinal cord motorneuron are affected. Several MNDs, like spinal and bulbar muscular atrophy (SBMA) or amyotrophic lateral sclerosis (ALS) have been linked to specific gene mutation which results in the production of aberrant proteins prone to misfold and to aggregate. To counteract the accumulation of these misfolded proteins cells, including neurons have developed a finely tuned protein quality control (PQC). The degradative systems are major components of the PQC system and comprise the ubiquitin-proteasome pathway and the autophagic pathways, specifically involved in the maintenance of a normal cell proteostasts. By working in association to salacted molecular chaperones, the degradative systems are essential components of the intracellular protein quality control (PQC) system. Misfolded proteins are neurotoxic and are recognized by chaperones and deleivered to degradative systems for their clearance. A tightly molecularly regulated equilibrium oversees the final fate of aberrant proteins. This equilibrium is crucial to mantain proteostasis in neurons and may chance under various stimuli, and its alteration give rise to a vicious cycle of protein accumulation and to the PQC system damage that might lead to cell death. Among chaperones, the small heat shock protein (HSP) B8, is able to facilitate autophagy and assist the removal of misfolded proteins prone to aggregate in MNDs. HSPB8 associates to BAG3, HSP70 and CHIP (an ubiquitinating enzyme) to deliver misfolded protein to autophagosome and this form of autophagy is called Chaperone-assisted selective autophagy (CASA) and the complex based on HSPB8/BAG3 s named CASA complex. HSPB8 is induced in response to se several neuronal stresses such has proteotoxic and oxidative stresses. We found that HSPB8 is highly induced in two main targets of misfolded protein toxicity in tg mice models of SBMA and ALS, the motorneurons and the muscle. The pharmacological or genetic induction of HSPB8 expression is protective in MNDs, while its siliencing has opposite effects. We also showed that HSPB8 protects from a misfolded protein induced abberant phenotype in fly models of ALS. By increasing HSPB8-mediated selective targeting of misfolded protein to autophagy neurons and muscle reduce their proteasome-mediated clearance limiting its possible overwhelming. Therefore, pharmacological approached which potentiate the HSPB8-BAG3 autophagic pathway could contribute to maintain a correct proteinostasis in motorneuron and muscle cells and might have therapeutic implication in MNDs
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The small heat shock protein B8 (HSPB8) modulates proliferation and migration of breast cancer cells
'Impact Journals LLC', 2017Co-Authors: M Piccolella, P Rusmini, V Crippa, R Cristofani, M E Cicardi, M Meroni, M. Galbiati, N. Ferri, F.f. Morelli, S. CarraAbstract:Breast cancer (BC) is one of the major causes of cancer death in women and is closely related to hormonal dysregulation. Estrogen receptor (ER)-positive BCs are generally treated with anti hormone therapy using antiestrogens or aromatase inhibitors. However, BC cells may become resistant to endocrine therapy, a process facilitated by autophagy, which may either promote or suppress tumor expansion. The autophagy facilitator HSPB8 has been found overexpressed in some BC. Here we found that HSPB8 is highly expressed and differentially modulated by natural or synthetic selective ER modulators (SERMs), in the triple-positive hormone-sensitive BC (MCF-7) cells, but not in triple-negative MDA-MB-231 BC cells. Specific SERMs induced MCF-7 cells proliferation in a HSPB8 dependent manner whereas, did not modify MDA-MB-231 cell growth. ER expression was unaffected in HSPB8-depleted MCF-7 cells. HSPB8 over-expression did not alter the distribution of MCF-7 cells in the various phases of the cell cycle. Conversely and intriguingly, HSPB8 downregulation resulted in an increased number of cells resting in the G0/G1 phase, thus possibly reducing the ability of the cells to pass through the restriction point. In addition, HSPB8 downregulation reduced the migratory ability of MCF-7 cells. None of these modifications were observed, when another small HSP (HSPB1), also expressed in MCF-7 cells, was downregulated. In conclusion, our data suggest that HSPB8 is involved in the mechanisms that regulate cell cycle and cell migration in MCF-7 cells
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transcriptional induction of the heat shock protein b8 mediates the clearance of misfolded proteins responsible for motor neuron diseases
Scientific Reports, 2016Co-Authors: V Crippa, P Rusmini, R Cristofani, M E Cicardi, Elio Messi, M Piccolella, Vito Giuseppe Dagostino, Rosa Loffredo, Michael Pancher, M. GalbiatiAbstract:Neurodegenerative diseases (NDs) are often associated with the presence of misfolded protein inclusions. The chaperone HSPB8 is upregulated in mice, the human brain and muscle structures affected during NDs progression. HSPB8 exerts a potent pro-degradative activity on several misfolded proteins responsible for familial NDs forms. Here, we demonstrated that HSPB8 also counteracts accumulation of aberrantly localized misfolded forms of TDP-43 and its 25 KDa fragment involved in most sporadic cases of Amyotrophic Lateral Sclerosis (sALS) and of Fronto Lateral Temporal Dementia (FLTD). HSPB8 acts with BAG3 and the HSP70/HSC70-CHIP complex enhancing the autophagic removal of misfolded proteins. We performed a high-through put screening (HTS) to find small molecules capable of inducing HSPB8 in neurons for therapeutic purposes. We identified two compounds, colchicine and doxorubicin, that robustly up-regulated HSPB8 expression. Both colchicine and doxorubicin increased the expression of the master regulator of autophagy TFEB, the autophagy linker p62/SQSTM1 and the autophagosome component LC3. In line, both drugs counteracted the accumulation of TDP-43 and TDP-25 misfolded species responsible for motoneuronal death in sALS. Thus, analogs of colchicine and doxorubicin able to induce HSPB8 and with better safety and tolerability may result beneficial in NDs models.
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transcriptional induction of the heat shock protein b8 mediates the clearance of misfolded proteins responsible for motor neuron diseases
Scientific Reports, 2016Co-Authors: V Crippa, P Rusmini, R Cristofani, M E Cicardi, Elio Messi, M Piccolella, Vito Giuseppe Dagostino, Rosa Loffredo, Michael Pancher, M. GalbiatiAbstract:Neurodegenerative diseases (NDs) are often associated with the presence of misfolded protein inclusions. The chaperone HSPB8 is upregulated in mice, the human brain and muscle structures affected during NDs progression. HSPB8 exerts a potent pro-degradative activity on several misfolded proteins responsible for familial NDs forms. Here, we demonstrated that HSPB8 also counteracts accumulation of aberrantly localized misfolded forms of TDP-43 and its 25 KDa fragment involved in most sporadic cases of Amyotrophic Lateral Sclerosis (sALS) and of Fronto Lateral Temporal Dementia (FLTD). HSPB8 acts with BAG3 and the HSP70/HSC70-CHIP complex enhancing the autophagic removal of misfolded proteins. We performed a high-through put screening (HTS) to find small molecules capable of inducing HSPB8 in neurons for therapeutic purposes. We identified two compounds, colchicine and doxorubicin, that robustly up-regulated HSPB8 expression. Both colchicine and doxorubicin increased the expression of the master regulator of autophagy TFEB, the autophagy linker p62/SQSTM1 and the autophagosome component LC3. In line, both drugs counteracted the accumulation of TDP-43 and TDP-25 misfolded species responsible for motoneuronal death in sALS. Thus, analogs of colchicine and doxorubicin able to induce HSPB8 and with better safety and tolerability may result beneficial in NDs models.
R Cristofani - One of the best experts on this subject based on the ideXlab platform.
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the role of hspb8 a component of the chaperone assisted selective autophagy machinery in cancer
Cells, 2021Co-Authors: R Cristofani, V Crippa, B Tedesco, A. Poletti, M Piccolella, Marina Montagnani Marelli, Roberta M MorettiAbstract:The cellular response to cancer-induced stress is one of the major aspects regulating cancer development and progression. The Heat Shock Protein B8 (HSPB8) is a small chaperone involved in chaperone-assisted selective autophagy (CASA). CASA promotes the selective degradation of proteins to counteract cell stress such as tumor-induced stress. HSPB8 is also involved in (i) the cell division machinery regulating chromosome segregation and cell cycle arrest in the G0/G1 phase and (ii) inflammation regulating dendritic cell maturation and cytokine production. HSPB8 expression and role are tumor-specific, showing a dual and opposite role. Interestingly, HSPB8 may be involved in the acquisition of chemoresistance to drugs. Despite the fact the mechanisms of HSPB8-mediated CASA activation in tumors need further studies, HSPB8 could represent an important factor in cancer induction and progression and it may be a potential target for anticancer treatment in specific types of cancer. In this review, we will discuss the molecular mechanism underlying HSPB8 roles in normal and cancer conditions. The basic mechanisms involved in anti- and pro-tumoral activities of HSPB8 are deeply discussed together with the pathways that modulate HSPB8 expression, in order to outline molecules with a beneficial effect for cancer cell growth, migration, and death.
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Trehalose induces autophagy via lysosomal-mediated TFEB activation in models of motoneuron degeneration
Autophagy, 2018Co-Authors: P Rusmini, Katia Cortese, V Crippa, R Cristofani, M E Cicardi, V Ferrari, G Vezzoli, B Tedesco, M Meroni, Elio MessiAbstract:Macroautophagy/autophagy, a defense mechanism against aberrant stresses, in neurons counteracts aggregate-prone misfolded protein toxicity. Autophagy induction might be beneficial in neurodegenerative diseases (NDs). The natural compound trehalose promotes autophagy via TFEB (transcription factor EB), ameliorating disease phenotype in multiple ND models, but its mechanism is still obscure. We demonstrated that trehalose regulates autophagy by inducing rapid and transient lysosomal enlargement and membrane permeabilization (LMP). This effect correlated with the calcium-dependent phosphatase PPP3/calcineurin activation, TFEB dephosphorylation and nuclear translocation. Trehalose upregulated genes for the TFEB target and regulator Ppargc1a, lysosomal hydrolases and membrane proteins (Ctsb, Gla, Lamp2a, Mcoln1, Tpp1) and several autophagy-related components (Becn1, Atg10, Atg12, Sqstm1/p62, Map1lc3b, Hspb8 and Bag3) mostly in a PPP3- and TFEB-dependent manner. TFEB silencing counteracted the trehalose pro-degradative activity on misfolded protein causative of motoneuron diseases. Similar effects were exerted by trehalase-resistant trehalose analogs, melibiose and lactulose. Thus, limited lysosomal damage might induce autophagy, perhaps as a compensatory mechanism, a process that is beneficial to counteract neurodegeneration. Abbreviations: ALS: amyotrophic lateral sclerosis; AR: androgen receptor; ATG: autophagy related; AV: autophagic vacuole; BAG3: BCL2-associated athanogene 3; BECN1: beclin 1, autophagy related; CASA: chaperone-assisted selective autophagy; CTSB: cathepsin b; DAPI: 4',6-diamidino-2-phenylindole; DMEM: Dulbecco's modified Eagle's medium; EGFP: enhanced green fluorescent protein; fALS, familial amyotrophic lateral sclerosis; FRA: filter retardation assay; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GLA: galactosidase, alpha; HD: Huntington disease; hIPSCs: human induced pluripotent stem cells; HSPA8: heat shock protein A8; HSPB8: heat shock protein B8; IF: immunofluorescence analysis; LAMP1: lysosomal-associated membrane protein 1; LAMP2A: lysosomal-associated membrane protein 2A; LGALS3: lectin, galactose binding, soluble 3; LLOMe: L-leucyl-L-leucine methyl ester; LMP: lysosomal membrane permeabilization; Lys: lysosomes; MAP1LC3B: microtubule-associated protein 1 light chain 3 beta; MCOLN1: mucolipin 1; mRNA: messenger RNA; MTOR: mechanistic target of rapamycin kinase; NDs: neurodegenerative diseases; NSC34: neuroblastoma x spinal cord 34; PBS: phosphate-buffered saline; PD: Parkinson disease; polyQ: polyglutamine; PPARGC1A: peroxisome proliferative activated receptor, gamma, coactivator 1 alpha; PPP3CB: protein phosphatase 3, catalytic subunit, beta isoform; RT-qPCR: real-time quantitative polymerase chain reaction; SBMA: spinal and bulbar muscular atrophy; SCAs: spinocerebellar ataxias; siRNA: small interfering RNA; SLC2A8: solute carrier family 2, (facilitated glucose transporter), member 8; smNPCs: small molecules neural progenitors cells; SOD1: superoxide dismutase 1; SQSTM1/p62: sequestosome 1; STED: stimulated emission depletion; STUB1: STIP1 homology and U-box containing protein 1; TARDBP/TDP-43: TAR DNA binding protein; TFEB: transcription factor EB; TPP1: tripeptidyl peptidase I; TREH: trehalase (brush-border membrane glycoprotein); WB: western blotting; ZKSCAN3: zinc finger with KRAB and SCAN domains 3.
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The small heat shock protein B8 at the interplay between the intracellular degradative pathways in motoneuron disease
2018Co-Authors: A. Poletti, P Rusmini, V Crippa, R Cristofani, M E Cicardi, V Ferrari, B Tedesco, M Meroni, M. Galbiati, S. CerraAbstract:Motor neuron diseases (MNDs) are a large class of neurodegenerative diseases in which upper cortical and lower spinal cord motorneuron are affected. Several MNDs, like spinal and bulbar muscular atrophy (SBMA) or amyotrophic lateral sclerosis (ALS) have been linked to specific gene mutation which results in the production of aberrant proteins prone to misfold and to aggregate. To counteract the accumulation of these misfolded proteins cells, including neurons have developed a finely tuned protein quality control (PQC). The degradative systems are major components of the PQC system and comprise the ubiquitin-proteasome pathway and the autophagic pathways, specifically involved in the maintenance of a normal cell proteostasts. By working in association to salacted molecular chaperones, the degradative systems are essential components of the intracellular protein quality control (PQC) system. Misfolded proteins are neurotoxic and are recognized by chaperones and deleivered to degradative systems for their clearance. A tightly molecularly regulated equilibrium oversees the final fate of aberrant proteins. This equilibrium is crucial to mantain proteostasis in neurons and may chance under various stimuli, and its alteration give rise to a vicious cycle of protein accumulation and to the PQC system damage that might lead to cell death. Among chaperones, the small heat shock protein (HSP) B8, is able to facilitate autophagy and assist the removal of misfolded proteins prone to aggregate in MNDs. HSPB8 associates to BAG3, HSP70 and CHIP (an ubiquitinating enzyme) to deliver misfolded protein to autophagosome and this form of autophagy is called Chaperone-assisted selective autophagy (CASA) and the complex based on HSPB8/BAG3 s named CASA complex. HSPB8 is induced in response to se several neuronal stresses such has proteotoxic and oxidative stresses. We found that HSPB8 is highly induced in two main targets of misfolded protein toxicity in tg mice models of SBMA and ALS, the motorneurons and the muscle. The pharmacological or genetic induction of HSPB8 expression is protective in MNDs, while its siliencing has opposite effects. We also showed that HSPB8 protects from a misfolded protein induced abberant phenotype in fly models of ALS. By increasing HSPB8-mediated selective targeting of misfolded protein to autophagy neurons and muscle reduce their proteasome-mediated clearance limiting its possible overwhelming. Therefore, pharmacological approached which potentiate the HSPB8-BAG3 autophagic pathway could contribute to maintain a correct proteinostasis in motorneuron and muscle cells and might have therapeutic implication in MNDs
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The small heat shock protein B8 (HSPB8) modulates proliferation and migration of breast cancer cells
'Impact Journals LLC', 2017Co-Authors: M Piccolella, P Rusmini, V Crippa, R Cristofani, M E Cicardi, M Meroni, M. Galbiati, N. Ferri, F.f. Morelli, S. CarraAbstract:Breast cancer (BC) is one of the major causes of cancer death in women and is closely related to hormonal dysregulation. Estrogen receptor (ER)-positive BCs are generally treated with anti hormone therapy using antiestrogens or aromatase inhibitors. However, BC cells may become resistant to endocrine therapy, a process facilitated by autophagy, which may either promote or suppress tumor expansion. The autophagy facilitator HSPB8 has been found overexpressed in some BC. Here we found that HSPB8 is highly expressed and differentially modulated by natural or synthetic selective ER modulators (SERMs), in the triple-positive hormone-sensitive BC (MCF-7) cells, but not in triple-negative MDA-MB-231 BC cells. Specific SERMs induced MCF-7 cells proliferation in a HSPB8 dependent manner whereas, did not modify MDA-MB-231 cell growth. ER expression was unaffected in HSPB8-depleted MCF-7 cells. HSPB8 over-expression did not alter the distribution of MCF-7 cells in the various phases of the cell cycle. Conversely and intriguingly, HSPB8 downregulation resulted in an increased number of cells resting in the G0/G1 phase, thus possibly reducing the ability of the cells to pass through the restriction point. In addition, HSPB8 downregulation reduced the migratory ability of MCF-7 cells. None of these modifications were observed, when another small HSP (HSPB1), also expressed in MCF-7 cells, was downregulated. In conclusion, our data suggest that HSPB8 is involved in the mechanisms that regulate cell cycle and cell migration in MCF-7 cells
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transcriptional induction of the heat shock protein b8 mediates the clearance of misfolded proteins responsible for motor neuron diseases
Scientific Reports, 2016Co-Authors: V Crippa, P Rusmini, R Cristofani, M E Cicardi, Elio Messi, M Piccolella, Vito Giuseppe Dagostino, Rosa Loffredo, Michael Pancher, M. GalbiatiAbstract:Neurodegenerative diseases (NDs) are often associated with the presence of misfolded protein inclusions. The chaperone HSPB8 is upregulated in mice, the human brain and muscle structures affected during NDs progression. HSPB8 exerts a potent pro-degradative activity on several misfolded proteins responsible for familial NDs forms. Here, we demonstrated that HSPB8 also counteracts accumulation of aberrantly localized misfolded forms of TDP-43 and its 25 KDa fragment involved in most sporadic cases of Amyotrophic Lateral Sclerosis (sALS) and of Fronto Lateral Temporal Dementia (FLTD). HSPB8 acts with BAG3 and the HSP70/HSC70-CHIP complex enhancing the autophagic removal of misfolded proteins. We performed a high-through put screening (HTS) to find small molecules capable of inducing HSPB8 in neurons for therapeutic purposes. We identified two compounds, colchicine and doxorubicin, that robustly up-regulated HSPB8 expression. Both colchicine and doxorubicin increased the expression of the master regulator of autophagy TFEB, the autophagy linker p62/SQSTM1 and the autophagosome component LC3. In line, both drugs counteracted the accumulation of TDP-43 and TDP-25 misfolded species responsible for motoneuronal death in sALS. Thus, analogs of colchicine and doxorubicin able to induce HSPB8 and with better safety and tolerability may result beneficial in NDs models.