Hong-Yu Hu | Shanghai Institutes for Biological Science (original) (raw)
Papers by Hong-Yu Hu
FEBS letters
TDP-43 (TAR DNA binding protein of 43kDa) and its C-terminal fragments are thought to be linked t... more TDP-43 (TAR DNA binding protein of 43kDa) and its C-terminal fragments are thought to be linked to the pathologies of amyotrophic lateral sclerosis and frontotemporal lobar degeneration. Here, we demonstrate that the aggregates or inclusions formed by its 35-kDa fragment (namely TDP-35) sequester full-length TDP-43 into cytoplasmic inclusions; and this sequestration is mediated by binding with RNA that is enriched in the cytoplasmic inclusions. RNA recognition motif 1 (RRM1) of TDP-43/TDP-35 plays a dominant role in nucleic-acid binding; mutation in this moiety abrogates formation of the TDP-35 inclusions and its RNA-assisted association with TDP-43. Thus, TDP-35 is able to sequester TDP-43 from nuclear localization into cytoplasmic inclusions, and RNA binding plays an essential role in this process. Copyright © 2015. Published by Elsevier B.V.
Scientific Reports
Polyglutamine (polyQ) expansion of proteins can trigger protein misfolding and amyloid-like aggre... more Polyglutamine (polyQ) expansion of proteins can trigger protein misfolding and amyloid-like aggregation, which thus lead to severe cytotoxicities and even the respective neurodegenerative diseases. However, why polyQ aggregation is toxic to cells is not fully elucidated. Here, we took the fragments of polyQ-expanded (PQE) ataxin-7 (Atx7) and huntingtin (Htt) as models to investigate the effect of polyQ aggregates on the cellular proteostasis of endogenous ataxin-3 (Atx3), a protein that frequently appears in diverse inclusion bodies. We found that PQE Atx7 and Htt impair the cellular proteostasis of Atx3 by reducing its soluble as well as total Atx3 level but enhancing formation of the aggregates. Expression of these polyQ proteins promotes proteasomal degradation of endogenous Atx3 and accumulation of its aggregated form. Then we verified that the co-chaperone HSJ1 is an essential factor that orchestrates the balance of cellular proteostasis of Atx3; and further discovered that the...
Biochemical Journal
Ubiquitin-specific protease 19 (USP19) is a member of the deubiquitinating (DUB) enzymes that cat... more Ubiquitin-specific protease 19 (USP19) is a member of the deubiquitinating (DUB) enzymes that catalyze removing the ubiquitin signals from target proteins. Our previous research has demonstrated that USP19 up-regulates the protein level and aggregation of polyQ-expanded huntingtin through the involvement of heat shock protein 90 (HSP90). Here, we present solution structures of the CS1, CS2 and UbL domains of USP19 and structural insights into their domain interactions. We found that the tandem CS domains fold back to interact with the C-terminal USP domain (USPD) intra-molecularly that leads to inhibition of the catalytic core of USP19, especially CS1 interacts with the embedded UbL domain and CS2 does with the CH2 catalytic core. Moreover, CS2 specifically interacts with the NBD domain of HSP90, which can activate the DUB enzyme. A mechanism of auto-inhibition of USP19 and activation by HSP90 is proposed, on which USP19 modulates the protein level of polyQ-expanded huntingtin in ce...
Journal of the American Chemical Society
FASEB journal : official publication of the Federation of American Societies for Experimental Biology, Jan 11, 2018
The components of ubiquitin (Ub)-proteasome system, such as Ub, Ub adaptors, or proteasome subuni... more The components of ubiquitin (Ub)-proteasome system, such as Ub, Ub adaptors, or proteasome subunits, are commonly accumulated with the aggregated proteins in inclusions, but how protein aggregates sequester Ub-related proteins remains elusive. Using N-terminal huntingtin (Htt-N552) and ataxin (Atx)-3 as model proteins, we investigated the molecular mechanism underlying sequestration of Ub adaptors by polyQ-expanded proteins. We found that polyQ-expanded Htt-N552 and Atx-3 sequester endogenous Ub adaptors, human RAD23 homolog B (hHR23B) and ubiquilin (UBQLN)-2, into inclusions. This sequestration effect is dependent on the UBA domains of Ub adaptors and the conjugated Ub of the aggregated proteins. Moreover, polyQ-expanded Htt-N552 and Atx-3 reduce the protein level of xeroderma pigmentosum group C (XPC) by sequestration of hHR23B, suggesting that this process may cut down the available quantity of hHR23B and thus affect its normal function in stabilizing XPC. Our findings demonstrat...
Current protein & peptide science, 2017
Protein aggregation and amyloidogenesis are closely associated with the pathogenesis of neurodege... more Protein aggregation and amyloidogenesis are closely associated with the pathogenesis of neurodegenerative diseases. Elucidating the morphology and structure of the amyloid aggregates or fibrils is important for understanding the molecular mechanisms of these proteinopathies. This review article describes the general principle and establishment of solid-state circular dichroism (ssCD) spectroscopy, and discusses its application for the analysis of secondary structures of proteins or peptides in amyloids and structural transformation of these proteins or peptides during their amyloidogenic aggregation.
Protein & Peptide Letters
Protein aggregation and amyloidogenesis are closely associated with the pathogenesis of neurodege... more Protein aggregation and amyloidogenesis are closely associated with the pathogenesis of neurodegenerative diseases, including Alzheimer's disease (AD), Parkinson's disease (PD) and amyotrophic lateral sclerosis (ALS) [1]. Structural transformation usually occurs during the aggregation process of the disease proteins. Thus, elucidating the morphology and structure of the amyloid aggregates or fibrils is vitally important for understanding the molecular mechanisms of these proteinopathies [2]. From the structure point of view, the "intermediate state" of a protein is the starting point of its aggregation process, especially when a large percent of proteins are intrinsically disordered. Mompeán and Laurents review the intrinsically disordered domains and the liquid phase formation in the RNA-binding proteins TDP-43 and FUS [3]; Song proposes the idea that the acquisition of membrane-interacting capacity is characteristic of protein aggregation [4]; and Yiu and Chen discuss the general experimental investigation of the oligomerization of intrinsically disordered disease proteins [5]. On the structural aspects of protein aggregates or fibrils, Liu et al. comment on the polymorphic structures of αB-crystallin under different conditions and their relationships with the respective diseases [6]; Lu et al. review various fibril structures of amyloid-β peptide solved by state-of-the-art solid-state NMR techniques [7]; and Khan and Kumar summarize different spectroscopic methods used for structural analysis of protein aggregation [8]. And lastly, Redington et al. discuss various ways of aggregation of protein therapeutics and a variety of approaches taken to prevent or minimize this process [9]. Although structural information on protein aggregates or fibrils is essential for us to understand the disease pathogenesis, elucidating the morphological structures of disease proteins at atomic resolution is still in its infancy. In this research field, many new ideas and methodologies are continuously emerging [10]. I believe that the structural knowledge of protein aggregation will be greatly expanded in the next decade.
PLOS ONE
Ubiquitin-specific protease 19 (USP19) is one of the deubiquitinating enzymes (DUBs) involved in ... more Ubiquitin-specific protease 19 (USP19) is one of the deubiquitinating enzymes (DUBs) involved in regulating the ubiquitination status of substrate proteins. There are two major isoforms of USP19 with distinct C-termini; the USP19_a isoform has a transmembrane domain for anchoring to the endoplasmic reticulum, while USP19_b contains an EEVD motif. Here, we report that the cytoplasmic isoform USP19_b up-regulates the protein levels of the polyglutamine (polyQ)-containing proteins, ataxin-3 (Atx3) and huntingtin (Htt), and thus promotes aggregation of their polyQ-expanded species in cell models. Our data demonstrate that USP19_b may orchestrate the stability, aggregation and degradation of the polyQ-expanded proteins through the heat shock protein 90 (HSP90) chaperone system. USP19_b directly interacts with HSP90 through its N-terminal CS (CHORD and SGT1)/P23 domains. In conjunction with HSP90, the cytoplasmic USP19 may play a key role in triage decision for the disease-related polyQ-expanded substrates, suggesting a function of USP19 in quality control of misfolded proteins by regulating their protein levels.
Biophysical journal, Jan 23, 2017
Ubiquitin-specific protease 25 (Usp25) is a deubiquitinase that is involved in multiple biologica... more Ubiquitin-specific protease 25 (Usp25) is a deubiquitinase that is involved in multiple biological processes. The N-terminal ubiquitin-binding region (UBR) of Usp25 contains one ubiquitin-associated domain, one small ubiquitin-like modifier (SUMO)-interacting motif and two ubiquitin-interacting motifs. Previous studies suggest that the covalent sumoylation in the UBR of Usp25 impairs its enzymatic activity. Here, we raise the hypothesis that non-covalent binding of SUMO, a prerequisite for efficient sumoylation, will impair Usp25's catalytic activity as well. To test our hypothesis and elucidate the underlying molecular mechanism, we investigated the structure and function of the Usp25 N-terminal UBR. The solution structure of Usp251-146 is obtained, and the key residues responsible for recognition of ubiquitin and SUMO2 are identified. Our data suggest inhibition of Usp25's catalytic activity upon the non-covalent binding of SUMO2 to the Usp25 SUMO-interacting motif. We als...
Scientific reports, Jan 31, 2016
TDP-43 is a DNA/RNA binding protein associated with TDP-43 proteinopathies. Many mutations have b... more TDP-43 is a DNA/RNA binding protein associated with TDP-43 proteinopathies. Many mutations have been identified in the flexible C-terminal region, which is implicated in the disease pathology. We investigated four point mutations in the amyloidogenic core region (residues 311-360) of TDP-43 by biochemical and spectroscopic methods. We found that the G335D mutation enhances the aggregation and inclusion formation of TDP-43 and this mutant in TDP-35 (the C-terminal fragment of 35 kDa) exaggerates the antagonist effect on RNA processing by endogenous TDP-43; whereas Q343R gives an opposite effect. As a comparison, M337V and Q331K have very little impact on the aggregation and inclusion formation of TDP-43 or TDP-35. NMR structural analysis showed that the G335D mutant in the core region forms a loop linker between the two α-helices and promotes α-to-β transition, but Q343R loses the second helix and consequently the structural transformation. Thus, the propensity of structural transfor...
The FEBS journal, Oct 26, 2016
Protein misfolding and aggregation are a hallmark of several neurodegenerative diseases (NDs). Ho... more Protein misfolding and aggregation are a hallmark of several neurodegenerative diseases (NDs). However, how protein aggregation leads to cytotoxicity and neurodegeneration is still controversial. Emerging evidence demonstrates that sequestration of cellular interacting partners by protein aggregates contributes to the pathogenesis of these diseases. Here, we review current research on sequestration of cellular proteins by protein aggregates and its relation to proteinopathies. Based on different interaction modes, we classify these protein sequestrations into four types: protein co-aggregation, domain/motif-mediated sequestration, RNA-assisted sequestration, and sequestration of molecular chaperones. Thus, the cellular essential proteins and/or RNAs hijacked by protein aggregates may lose their biological functions, consequently resulting in cytotoxicity and neurodegeneration. We have proposed a hijacking model recapitulating the sequestration process and the loss-of-function pathol...
The Biochemical journal, Jan 12, 2015
Deubiquitinase Usp28 contains a ubiquitin binding region (UBR) composed of one ubiquitin associat... more Deubiquitinase Usp28 contains a ubiquitin binding region (UBR) composed of one ubiquitin associated domain (UBA) and one ubiquitin interacting motif (UIM) at its N-terminus. Interestingly, an additional SUMO interacting motif (SIM) locates next to its UIM. Up to date, the functional role of Usp28 UBR is still not understood. To elucidate the regulatory mechanism of the UBR on the full functional display of Usp28, in this report, NMR and biochemical approaches are applied. The solution structure of Usp28 UBR is obtained, and the key residues responsible for ubiquitin and SUMO1/2 recognition are identified. Besides, we find that the ubiquitin binding ability of Usp28 UBR is required for the full enzymatic activity of Usp28, while binding of SUMO1/2 impairs the catalytic activity of the enzyme by competitively blocking its interactions with ubiquitin substrates. Our findings provide a first insight for understanding how the enzymatic activity of Usp28 is regulated by its non-catalytic ...
PloS one, 2011
Homo sapiens J domain protein (HSJ1) is a J-domain containing co-chaperone that is known to stimu... more Homo sapiens J domain protein (HSJ1) is a J-domain containing co-chaperone that is known to stimulate ATPase activity of HSP70 chaperone, while it also harbors two ubiquitin (Ub)-interacting motifs (UIMs) that may bind with ubiquitinated substrates and potentially function in protein degradation. We studied the effects of HSJ1a on the protein levels of both normal and the disease--related polyQ-expanded forms of ataxin-3 (Atx3) in cells. The results demonstrate that the N-terminal J-domain and the C-terminal UIM domain of HSJ1a exert opposite functions in regulating the protein level of cellular overexpressed Atx3. This dual regulation is dependent on the binding of the J-domain with HSP70, and the UIM domain with polyUb chains. The J-domain down-regulates the protein level of Atx3 through HSP70 mediated proteasomal degradation, while the UIM domain may alleviate this process via maintaining the ubiquitinated Atx3. We propose that co-chaperone HSJ1a orchestrates the balance of subst...
Acta biochimica et biophysica Sinica, 2008
Most neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, Hunti... more Most neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, Huntington's disease and other polyglutamine diseases are associated with degeneration and death of specific neuronal populations due to misfolding or aggregation of certain proteins. These aggregates often contain ubiquitin that is the signal for proteolysis by the ubiquitin-proteasome system, and chaperone proteins that are involved in the assistance of protein folding. Here we review the role of protein quality control systems in the pathogenesis of neurodegenerative diseases, and aim to learn more from the cooperation between molecular chaperones and ubiquitin-proteasome system responding to cellular protein aggregates, in order to find molecular targets for therapeutic intervention.
Yao xue xue bao = Acta pharmaceutica Sinica, 2002
Sheng wu hua xue yu sheng wu wu li xue bao Acta biochimica et biophysica Sinica, 2003
Spin systems for amino acid residues in mHCN1 pore region peptide have been identified through an... more Spin systems for amino acid residues in mHCN1 pore region peptide have been identified through analysis of 2D NMR spectra. The sequence-specific assignment of spin systems was obtained by NOEs correlation in WET-NOESY spectra, and the complete assignment of proton resonances for backbone and side chain has been achieved. CNS software was used to calculate the structure of mHCN1 19 aa peptide. The results show that an alpha-helix from residue 10 to residue 13 is formed within the pore region. The results of NMR study on mHCN1 pore peptide provide the basis for further understanding the mechanism of ion selectivity of channels.
PloS one, 2014
The ubiquitination levels of protein substrates in eukaryotic cells are delicately orchestrated b... more The ubiquitination levels of protein substrates in eukaryotic cells are delicately orchestrated by various protein cofactors and enzymes. Dendritic cell-derived ubiquitin (Ub)-like protein (DC-UbP), also named as Ub domain-containing protein 2 (UBTD2), is a potential Ub shuttle protein comprised of a Ub-like (UbL) domain and a Ub-binding domain (UBD), but its biological function remains largely unknown. We identified two Ub-related enzymes, the deubiquitinating enzyme USP5 and the Ub-activating enzyme UbE1, as interacting partners of DC-UbP from HEK 293T cells. Biochemical studies revealed that the tandem UBA domains of USP5 and the C-terminal Ub-fold domain (UFD) of UbE1 directly interacted with the C-terminal UbL domain of DC-UbP but on the distinct surfaces. Overexpression of DC-UbP in HEK 293T cells enhanced the association of these two enzymes and thus prompted cellular ubiquitination, whereas knockdown of the protein reduced the cellular ubiquitination level. Together, DC-UbP ...
Structure (London, England : 1993), Jan 4, 2014
Huntington's disease (HD) is an autosomally dominant neurodegenerative disorder caused by exp... more Huntington's disease (HD) is an autosomally dominant neurodegenerative disorder caused by expansion of polyglutamine (polyQ) in the huntingtin (Htt) protein. Htt yeast two-hybrid protein B (HYPB/SETD2), a histone methyltransferase, directly interacts with Htt and is involved in HD pathology. Using NMR techniques, we characterized a polyproline (polyP) stretch at the C terminus of HYPB, which directly interacts with the following WW domain and leads this domain predominantly to be in a closed conformational state. The solution structure shows that the polyP stretch extends from the back and binds to the WW core domain in a typical binding mode. This autoinhibitory structure regulates interaction between the WW domain of HYPB and the proline-rich region (PRR) of Htt, as evidenced by NMR and immunofluorescence techniques. This work provides structural and mechanistic insights into the intramolecular regulation of the WW domain in Htt-interacting partners and will be helpful for und...
Scientific reports, Jan 18, 2014
Expansion of polyglutamine (polyQ) tract may cause protein misfolding and aggregation that lead t... more Expansion of polyglutamine (polyQ) tract may cause protein misfolding and aggregation that lead to cytotoxicity and neurodegeneration, but the underlying mechanism remains to be elucidated. We applied ataxin-3 (Atx3), a polyQ tract-containing protein, as a model to study sequestration of normal cellular proteins. We found that the aggregates formed by polyQ-expanded Atx3 sequester its interacting partners, such as P97/VCP and ubiquitin conjugates, into the protein inclusions through specific interactions both in vitro and in cells. Moreover, this specific sequestration impairs the normal cellular function of P97 in down-regulating neddylation. However, expansion of polyQ tract in Atx3 does not alter the conformation of its surrounding regions and the interaction affinities with the interacting partners, although it indeed facilitates misfolding and aggregation of the Atx3 protein. Thus, we propose a loss-of-function pathology for polyQ diseases that sequestration of the cellular ess...
Acta biochimica et biophysica Sinica, 2005
Human alpha-synuclein is a presynaptic terminal protein and can form insoluble fibrils that are b... more Human alpha-synuclein is a presynaptic terminal protein and can form insoluble fibrils that are believed to play an important role in the pathogenesis of several neurodegenerative diseases such as Parkinson's disease, dementia with Lewy bodies and Lewy body variant of Alzheimer's disease. In this paper, in situ atomic force microscopy has been used to study the structural properties of alpha-synuclein fibrils in solution using two different atomic force microscopy imaging modes: tapping mode and contact mode. In the in situ contact mode atomic force microscopy experiments alpha-synuclein fibrils quickly broke into fragments, and a similar phenomenon was found using tapping mode atomic force microscopy in which alpha-synuclein fibrils were incubated with guanidine hydrochloride (0.6 M). The alpha-synuclein fibrils kept their original filamentous topography for over 1 h in the in situ tapping mode atomic force microscopy experiments. The present results provide indirect eviden...
FEBS letters
TDP-43 (TAR DNA binding protein of 43kDa) and its C-terminal fragments are thought to be linked t... more TDP-43 (TAR DNA binding protein of 43kDa) and its C-terminal fragments are thought to be linked to the pathologies of amyotrophic lateral sclerosis and frontotemporal lobar degeneration. Here, we demonstrate that the aggregates or inclusions formed by its 35-kDa fragment (namely TDP-35) sequester full-length TDP-43 into cytoplasmic inclusions; and this sequestration is mediated by binding with RNA that is enriched in the cytoplasmic inclusions. RNA recognition motif 1 (RRM1) of TDP-43/TDP-35 plays a dominant role in nucleic-acid binding; mutation in this moiety abrogates formation of the TDP-35 inclusions and its RNA-assisted association with TDP-43. Thus, TDP-35 is able to sequester TDP-43 from nuclear localization into cytoplasmic inclusions, and RNA binding plays an essential role in this process. Copyright © 2015. Published by Elsevier B.V.
Scientific Reports
Polyglutamine (polyQ) expansion of proteins can trigger protein misfolding and amyloid-like aggre... more Polyglutamine (polyQ) expansion of proteins can trigger protein misfolding and amyloid-like aggregation, which thus lead to severe cytotoxicities and even the respective neurodegenerative diseases. However, why polyQ aggregation is toxic to cells is not fully elucidated. Here, we took the fragments of polyQ-expanded (PQE) ataxin-7 (Atx7) and huntingtin (Htt) as models to investigate the effect of polyQ aggregates on the cellular proteostasis of endogenous ataxin-3 (Atx3), a protein that frequently appears in diverse inclusion bodies. We found that PQE Atx7 and Htt impair the cellular proteostasis of Atx3 by reducing its soluble as well as total Atx3 level but enhancing formation of the aggregates. Expression of these polyQ proteins promotes proteasomal degradation of endogenous Atx3 and accumulation of its aggregated form. Then we verified that the co-chaperone HSJ1 is an essential factor that orchestrates the balance of cellular proteostasis of Atx3; and further discovered that the...
Biochemical Journal
Ubiquitin-specific protease 19 (USP19) is a member of the deubiquitinating (DUB) enzymes that cat... more Ubiquitin-specific protease 19 (USP19) is a member of the deubiquitinating (DUB) enzymes that catalyze removing the ubiquitin signals from target proteins. Our previous research has demonstrated that USP19 up-regulates the protein level and aggregation of polyQ-expanded huntingtin through the involvement of heat shock protein 90 (HSP90). Here, we present solution structures of the CS1, CS2 and UbL domains of USP19 and structural insights into their domain interactions. We found that the tandem CS domains fold back to interact with the C-terminal USP domain (USPD) intra-molecularly that leads to inhibition of the catalytic core of USP19, especially CS1 interacts with the embedded UbL domain and CS2 does with the CH2 catalytic core. Moreover, CS2 specifically interacts with the NBD domain of HSP90, which can activate the DUB enzyme. A mechanism of auto-inhibition of USP19 and activation by HSP90 is proposed, on which USP19 modulates the protein level of polyQ-expanded huntingtin in ce...
Journal of the American Chemical Society
FASEB journal : official publication of the Federation of American Societies for Experimental Biology, Jan 11, 2018
The components of ubiquitin (Ub)-proteasome system, such as Ub, Ub adaptors, or proteasome subuni... more The components of ubiquitin (Ub)-proteasome system, such as Ub, Ub adaptors, or proteasome subunits, are commonly accumulated with the aggregated proteins in inclusions, but how protein aggregates sequester Ub-related proteins remains elusive. Using N-terminal huntingtin (Htt-N552) and ataxin (Atx)-3 as model proteins, we investigated the molecular mechanism underlying sequestration of Ub adaptors by polyQ-expanded proteins. We found that polyQ-expanded Htt-N552 and Atx-3 sequester endogenous Ub adaptors, human RAD23 homolog B (hHR23B) and ubiquilin (UBQLN)-2, into inclusions. This sequestration effect is dependent on the UBA domains of Ub adaptors and the conjugated Ub of the aggregated proteins. Moreover, polyQ-expanded Htt-N552 and Atx-3 reduce the protein level of xeroderma pigmentosum group C (XPC) by sequestration of hHR23B, suggesting that this process may cut down the available quantity of hHR23B and thus affect its normal function in stabilizing XPC. Our findings demonstrat...
Current protein & peptide science, 2017
Protein aggregation and amyloidogenesis are closely associated with the pathogenesis of neurodege... more Protein aggregation and amyloidogenesis are closely associated with the pathogenesis of neurodegenerative diseases. Elucidating the morphology and structure of the amyloid aggregates or fibrils is important for understanding the molecular mechanisms of these proteinopathies. This review article describes the general principle and establishment of solid-state circular dichroism (ssCD) spectroscopy, and discusses its application for the analysis of secondary structures of proteins or peptides in amyloids and structural transformation of these proteins or peptides during their amyloidogenic aggregation.
Protein & Peptide Letters
Protein aggregation and amyloidogenesis are closely associated with the pathogenesis of neurodege... more Protein aggregation and amyloidogenesis are closely associated with the pathogenesis of neurodegenerative diseases, including Alzheimer's disease (AD), Parkinson's disease (PD) and amyotrophic lateral sclerosis (ALS) [1]. Structural transformation usually occurs during the aggregation process of the disease proteins. Thus, elucidating the morphology and structure of the amyloid aggregates or fibrils is vitally important for understanding the molecular mechanisms of these proteinopathies [2]. From the structure point of view, the "intermediate state" of a protein is the starting point of its aggregation process, especially when a large percent of proteins are intrinsically disordered. Mompeán and Laurents review the intrinsically disordered domains and the liquid phase formation in the RNA-binding proteins TDP-43 and FUS [3]; Song proposes the idea that the acquisition of membrane-interacting capacity is characteristic of protein aggregation [4]; and Yiu and Chen discuss the general experimental investigation of the oligomerization of intrinsically disordered disease proteins [5]. On the structural aspects of protein aggregates or fibrils, Liu et al. comment on the polymorphic structures of αB-crystallin under different conditions and their relationships with the respective diseases [6]; Lu et al. review various fibril structures of amyloid-β peptide solved by state-of-the-art solid-state NMR techniques [7]; and Khan and Kumar summarize different spectroscopic methods used for structural analysis of protein aggregation [8]. And lastly, Redington et al. discuss various ways of aggregation of protein therapeutics and a variety of approaches taken to prevent or minimize this process [9]. Although structural information on protein aggregates or fibrils is essential for us to understand the disease pathogenesis, elucidating the morphological structures of disease proteins at atomic resolution is still in its infancy. In this research field, many new ideas and methodologies are continuously emerging [10]. I believe that the structural knowledge of protein aggregation will be greatly expanded in the next decade.
PLOS ONE
Ubiquitin-specific protease 19 (USP19) is one of the deubiquitinating enzymes (DUBs) involved in ... more Ubiquitin-specific protease 19 (USP19) is one of the deubiquitinating enzymes (DUBs) involved in regulating the ubiquitination status of substrate proteins. There are two major isoforms of USP19 with distinct C-termini; the USP19_a isoform has a transmembrane domain for anchoring to the endoplasmic reticulum, while USP19_b contains an EEVD motif. Here, we report that the cytoplasmic isoform USP19_b up-regulates the protein levels of the polyglutamine (polyQ)-containing proteins, ataxin-3 (Atx3) and huntingtin (Htt), and thus promotes aggregation of their polyQ-expanded species in cell models. Our data demonstrate that USP19_b may orchestrate the stability, aggregation and degradation of the polyQ-expanded proteins through the heat shock protein 90 (HSP90) chaperone system. USP19_b directly interacts with HSP90 through its N-terminal CS (CHORD and SGT1)/P23 domains. In conjunction with HSP90, the cytoplasmic USP19 may play a key role in triage decision for the disease-related polyQ-expanded substrates, suggesting a function of USP19 in quality control of misfolded proteins by regulating their protein levels.
Biophysical journal, Jan 23, 2017
Ubiquitin-specific protease 25 (Usp25) is a deubiquitinase that is involved in multiple biologica... more Ubiquitin-specific protease 25 (Usp25) is a deubiquitinase that is involved in multiple biological processes. The N-terminal ubiquitin-binding region (UBR) of Usp25 contains one ubiquitin-associated domain, one small ubiquitin-like modifier (SUMO)-interacting motif and two ubiquitin-interacting motifs. Previous studies suggest that the covalent sumoylation in the UBR of Usp25 impairs its enzymatic activity. Here, we raise the hypothesis that non-covalent binding of SUMO, a prerequisite for efficient sumoylation, will impair Usp25's catalytic activity as well. To test our hypothesis and elucidate the underlying molecular mechanism, we investigated the structure and function of the Usp25 N-terminal UBR. The solution structure of Usp251-146 is obtained, and the key residues responsible for recognition of ubiquitin and SUMO2 are identified. Our data suggest inhibition of Usp25's catalytic activity upon the non-covalent binding of SUMO2 to the Usp25 SUMO-interacting motif. We als...
Scientific reports, Jan 31, 2016
TDP-43 is a DNA/RNA binding protein associated with TDP-43 proteinopathies. Many mutations have b... more TDP-43 is a DNA/RNA binding protein associated with TDP-43 proteinopathies. Many mutations have been identified in the flexible C-terminal region, which is implicated in the disease pathology. We investigated four point mutations in the amyloidogenic core region (residues 311-360) of TDP-43 by biochemical and spectroscopic methods. We found that the G335D mutation enhances the aggregation and inclusion formation of TDP-43 and this mutant in TDP-35 (the C-terminal fragment of 35 kDa) exaggerates the antagonist effect on RNA processing by endogenous TDP-43; whereas Q343R gives an opposite effect. As a comparison, M337V and Q331K have very little impact on the aggregation and inclusion formation of TDP-43 or TDP-35. NMR structural analysis showed that the G335D mutant in the core region forms a loop linker between the two α-helices and promotes α-to-β transition, but Q343R loses the second helix and consequently the structural transformation. Thus, the propensity of structural transfor...
The FEBS journal, Oct 26, 2016
Protein misfolding and aggregation are a hallmark of several neurodegenerative diseases (NDs). Ho... more Protein misfolding and aggregation are a hallmark of several neurodegenerative diseases (NDs). However, how protein aggregation leads to cytotoxicity and neurodegeneration is still controversial. Emerging evidence demonstrates that sequestration of cellular interacting partners by protein aggregates contributes to the pathogenesis of these diseases. Here, we review current research on sequestration of cellular proteins by protein aggregates and its relation to proteinopathies. Based on different interaction modes, we classify these protein sequestrations into four types: protein co-aggregation, domain/motif-mediated sequestration, RNA-assisted sequestration, and sequestration of molecular chaperones. Thus, the cellular essential proteins and/or RNAs hijacked by protein aggregates may lose their biological functions, consequently resulting in cytotoxicity and neurodegeneration. We have proposed a hijacking model recapitulating the sequestration process and the loss-of-function pathol...
The Biochemical journal, Jan 12, 2015
Deubiquitinase Usp28 contains a ubiquitin binding region (UBR) composed of one ubiquitin associat... more Deubiquitinase Usp28 contains a ubiquitin binding region (UBR) composed of one ubiquitin associated domain (UBA) and one ubiquitin interacting motif (UIM) at its N-terminus. Interestingly, an additional SUMO interacting motif (SIM) locates next to its UIM. Up to date, the functional role of Usp28 UBR is still not understood. To elucidate the regulatory mechanism of the UBR on the full functional display of Usp28, in this report, NMR and biochemical approaches are applied. The solution structure of Usp28 UBR is obtained, and the key residues responsible for ubiquitin and SUMO1/2 recognition are identified. Besides, we find that the ubiquitin binding ability of Usp28 UBR is required for the full enzymatic activity of Usp28, while binding of SUMO1/2 impairs the catalytic activity of the enzyme by competitively blocking its interactions with ubiquitin substrates. Our findings provide a first insight for understanding how the enzymatic activity of Usp28 is regulated by its non-catalytic ...
PloS one, 2011
Homo sapiens J domain protein (HSJ1) is a J-domain containing co-chaperone that is known to stimu... more Homo sapiens J domain protein (HSJ1) is a J-domain containing co-chaperone that is known to stimulate ATPase activity of HSP70 chaperone, while it also harbors two ubiquitin (Ub)-interacting motifs (UIMs) that may bind with ubiquitinated substrates and potentially function in protein degradation. We studied the effects of HSJ1a on the protein levels of both normal and the disease--related polyQ-expanded forms of ataxin-3 (Atx3) in cells. The results demonstrate that the N-terminal J-domain and the C-terminal UIM domain of HSJ1a exert opposite functions in regulating the protein level of cellular overexpressed Atx3. This dual regulation is dependent on the binding of the J-domain with HSP70, and the UIM domain with polyUb chains. The J-domain down-regulates the protein level of Atx3 through HSP70 mediated proteasomal degradation, while the UIM domain may alleviate this process via maintaining the ubiquitinated Atx3. We propose that co-chaperone HSJ1a orchestrates the balance of subst...
Acta biochimica et biophysica Sinica, 2008
Most neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, Hunti... more Most neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, Huntington's disease and other polyglutamine diseases are associated with degeneration and death of specific neuronal populations due to misfolding or aggregation of certain proteins. These aggregates often contain ubiquitin that is the signal for proteolysis by the ubiquitin-proteasome system, and chaperone proteins that are involved in the assistance of protein folding. Here we review the role of protein quality control systems in the pathogenesis of neurodegenerative diseases, and aim to learn more from the cooperation between molecular chaperones and ubiquitin-proteasome system responding to cellular protein aggregates, in order to find molecular targets for therapeutic intervention.
Yao xue xue bao = Acta pharmaceutica Sinica, 2002
Sheng wu hua xue yu sheng wu wu li xue bao Acta biochimica et biophysica Sinica, 2003
Spin systems for amino acid residues in mHCN1 pore region peptide have been identified through an... more Spin systems for amino acid residues in mHCN1 pore region peptide have been identified through analysis of 2D NMR spectra. The sequence-specific assignment of spin systems was obtained by NOEs correlation in WET-NOESY spectra, and the complete assignment of proton resonances for backbone and side chain has been achieved. CNS software was used to calculate the structure of mHCN1 19 aa peptide. The results show that an alpha-helix from residue 10 to residue 13 is formed within the pore region. The results of NMR study on mHCN1 pore peptide provide the basis for further understanding the mechanism of ion selectivity of channels.
PloS one, 2014
The ubiquitination levels of protein substrates in eukaryotic cells are delicately orchestrated b... more The ubiquitination levels of protein substrates in eukaryotic cells are delicately orchestrated by various protein cofactors and enzymes. Dendritic cell-derived ubiquitin (Ub)-like protein (DC-UbP), also named as Ub domain-containing protein 2 (UBTD2), is a potential Ub shuttle protein comprised of a Ub-like (UbL) domain and a Ub-binding domain (UBD), but its biological function remains largely unknown. We identified two Ub-related enzymes, the deubiquitinating enzyme USP5 and the Ub-activating enzyme UbE1, as interacting partners of DC-UbP from HEK 293T cells. Biochemical studies revealed that the tandem UBA domains of USP5 and the C-terminal Ub-fold domain (UFD) of UbE1 directly interacted with the C-terminal UbL domain of DC-UbP but on the distinct surfaces. Overexpression of DC-UbP in HEK 293T cells enhanced the association of these two enzymes and thus prompted cellular ubiquitination, whereas knockdown of the protein reduced the cellular ubiquitination level. Together, DC-UbP ...
Structure (London, England : 1993), Jan 4, 2014
Huntington's disease (HD) is an autosomally dominant neurodegenerative disorder caused by exp... more Huntington's disease (HD) is an autosomally dominant neurodegenerative disorder caused by expansion of polyglutamine (polyQ) in the huntingtin (Htt) protein. Htt yeast two-hybrid protein B (HYPB/SETD2), a histone methyltransferase, directly interacts with Htt and is involved in HD pathology. Using NMR techniques, we characterized a polyproline (polyP) stretch at the C terminus of HYPB, which directly interacts with the following WW domain and leads this domain predominantly to be in a closed conformational state. The solution structure shows that the polyP stretch extends from the back and binds to the WW core domain in a typical binding mode. This autoinhibitory structure regulates interaction between the WW domain of HYPB and the proline-rich region (PRR) of Htt, as evidenced by NMR and immunofluorescence techniques. This work provides structural and mechanistic insights into the intramolecular regulation of the WW domain in Htt-interacting partners and will be helpful for und...
Scientific reports, Jan 18, 2014
Expansion of polyglutamine (polyQ) tract may cause protein misfolding and aggregation that lead t... more Expansion of polyglutamine (polyQ) tract may cause protein misfolding and aggregation that lead to cytotoxicity and neurodegeneration, but the underlying mechanism remains to be elucidated. We applied ataxin-3 (Atx3), a polyQ tract-containing protein, as a model to study sequestration of normal cellular proteins. We found that the aggregates formed by polyQ-expanded Atx3 sequester its interacting partners, such as P97/VCP and ubiquitin conjugates, into the protein inclusions through specific interactions both in vitro and in cells. Moreover, this specific sequestration impairs the normal cellular function of P97 in down-regulating neddylation. However, expansion of polyQ tract in Atx3 does not alter the conformation of its surrounding regions and the interaction affinities with the interacting partners, although it indeed facilitates misfolding and aggregation of the Atx3 protein. Thus, we propose a loss-of-function pathology for polyQ diseases that sequestration of the cellular ess...
Acta biochimica et biophysica Sinica, 2005
Human alpha-synuclein is a presynaptic terminal protein and can form insoluble fibrils that are b... more Human alpha-synuclein is a presynaptic terminal protein and can form insoluble fibrils that are believed to play an important role in the pathogenesis of several neurodegenerative diseases such as Parkinson's disease, dementia with Lewy bodies and Lewy body variant of Alzheimer's disease. In this paper, in situ atomic force microscopy has been used to study the structural properties of alpha-synuclein fibrils in solution using two different atomic force microscopy imaging modes: tapping mode and contact mode. In the in situ contact mode atomic force microscopy experiments alpha-synuclein fibrils quickly broke into fragments, and a similar phenomenon was found using tapping mode atomic force microscopy in which alpha-synuclein fibrils were incubated with guanidine hydrochloride (0.6 M). The alpha-synuclein fibrils kept their original filamentous topography for over 1 h in the in situ tapping mode atomic force microscopy experiments. The present results provide indirect eviden...