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Identification of ubiquilin, a novel presenilin interactor that increases presenilin protein accumulation.

Mah AL, Perry G, Smith MA, Monteiro MJ - J. Cell Biol. (2000)

Bottom Line: However, apart from a role in early development, neither the normal function of the presenilins nor the mechanisms by which mutant proteins cause AD are well understood.Moreover, the anti-ubiquilin antibodies robustly stained neurofibrillary tangles and Lewy bodies in AD and Parkinson's disease affected brains, respectively.Our results indicate that ubiquilin may be an important modulator of presenilin protein accumulation and that ubiquilin protein is associated with neuropathological neurofibrillary tangles and Lewy body inclusions in diseased brain.

View Article: PubMed Central - PubMed

Affiliation: Medical Biotechnology Center, Department of Neurology, University of Maryland Biotechnology Institute, Baltimore, Maryland 21201, USA.

ABSTRACT
Mutations in the highly homologous presenilin genes encoding presenilin-1 and presenilin-2 (PS1 and PS2) are linked to early-onset Alzheimer's disease (AD). However, apart from a role in early development, neither the normal function of the presenilins nor the mechanisms by which mutant proteins cause AD are well understood. We describe here the properties of a novel human interactor of the presenilins named ubiquilin. Yeast two-hybrid (Y2H) interaction, glutathione S-transferase pull-down experiments, and colocalization of the proteins expressed in vivo, together with coimmunoprecipitation and cell fractionation studies, provide compelling evidence that ubiquilin interacts with both PS1 and PS2. Ubiquilin is noteworthy since it contains multiple ubiquitin-related domains typically thought to be involved in targeting proteins for degradation. However, we show that ubiquilin promotes presenilin protein accumulation. Pulse-labeling experiments indicate that ubiquilin facilitates increased presenilin synthesis without substantially changing presenilin protein half-life. Immunohistochemistry of human brain tissue with ubiquilin-specific antibodies revealed prominent staining of neurons. Moreover, the anti-ubiquilin antibodies robustly stained neurofibrillary tangles and Lewy bodies in AD and Parkinson's disease affected brains, respectively. Our results indicate that ubiquilin may be an important modulator of presenilin protein accumulation and that ubiquilin protein is associated with neuropathological neurofibrillary tangles and Lewy body inclusions in diseased brain.

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Ubiquilin shares significant homology with several other proteins. The inferred amino acid sequence of the human ubiquilin ORF and its homology to several related proteins: Homo sapiens Chap1, a protein involved in binding Hsp70-like Stch protein; Mus musculus PLIC-1 and PLIC-2, proteins involved in linking integrins to vimentin; X. laevis XDRP1, a protein which binds cyclin A; C. elegans F15C11.2 and two A. thaliana proteins, proteins with unknown functions; and S. cerevisiae DSK2, involved in spindle pole body duplication. Identical residues are darkly shaded, whereas similar residues are lightly shaded. Only homology in five or more sequences are shown. All the proteins contain UB domains (square box) and UBA domains (rounded box), which are extremely well conserved in sequence. In addition, the central region between the UB and UBA domains contain several asparagine-proline repeats, which are either regularly spaced apart (closed circles) or located elsewhere throughout the protein (open circles). The high overall homology of the proteins suggests that they belong to the same multigene family.
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Figure 4: Ubiquilin shares significant homology with several other proteins. The inferred amino acid sequence of the human ubiquilin ORF and its homology to several related proteins: Homo sapiens Chap1, a protein involved in binding Hsp70-like Stch protein; Mus musculus PLIC-1 and PLIC-2, proteins involved in linking integrins to vimentin; X. laevis XDRP1, a protein which binds cyclin A; C. elegans F15C11.2 and two A. thaliana proteins, proteins with unknown functions; and S. cerevisiae DSK2, involved in spindle pole body duplication. Identical residues are darkly shaded, whereas similar residues are lightly shaded. Only homology in five or more sequences are shown. All the proteins contain UB domains (square box) and UBA domains (rounded box), which are extremely well conserved in sequence. In addition, the central region between the UB and UBA domains contain several asparagine-proline repeats, which are either regularly spaced apart (closed circles) or located elsewhere throughout the protein (open circles). The high overall homology of the proteins suggests that they belong to the same multigene family.

Mentions: The complete ubiquilin ORF consists of 595 residues, with a sequence rich in glutamines and serines, 10.9% and 11.1%, respectively, but lacking any cysteines (Fig. 4). The protein has a calculated isoelectric point (pI) of 4.97 and is predicted to be soluble, based on having a high hydrophilic profile using MacVector 6.5 software (Oxford Molecular Group; data not shown). Currently known protein structural motifs residing within ubiquilin include an NH2-terminal UB domain and a COOH-terminal ubiquitin-associated (UBA) domain, both of which have been implicated in targeting and degradation of proteins by the ubiquitin-proteasome pathway. The predicted ubiquilin protein also contains numerous regularly spaced asparagine-proline (Asn-Pro) repeats of unknown function (Fig. 4). When first sequenced, no apparent mammalian homologue for ubiquilin had been deposited in GenBank/EMBL/DDBJ databases. However, there was significant homology to several full-length genes of unknown function: the C. elegans F15C11.2 gene (36% identical) and two related genes in Arabidopsis thaliana (34 and 35% identical). In addition, the protein had weak homology to the Saccharomyces cerevisiae DSK2 gene (31% identical), which can suppress a KAR1 defect (Biggins et al. 1996). KAR1 is an essential gene involved in the duplication of the yeast microtubule organizing center known as the spindle pole body. Interestingly, ubiquilin homology to DSK2 was primarily confined to the UB and UBA domains. However, since both UB and UBA domains are present in many other proteins, it is uncertain whether ubiquilin and DSK2 are true orthologs.


Identification of ubiquilin, a novel presenilin interactor that increases presenilin protein accumulation.

Mah AL, Perry G, Smith MA, Monteiro MJ - J. Cell Biol. (2000)

Ubiquilin shares significant homology with several other proteins. The inferred amino acid sequence of the human ubiquilin ORF and its homology to several related proteins: Homo sapiens Chap1, a protein involved in binding Hsp70-like Stch protein; Mus musculus PLIC-1 and PLIC-2, proteins involved in linking integrins to vimentin; X. laevis XDRP1, a protein which binds cyclin A; C. elegans F15C11.2 and two A. thaliana proteins, proteins with unknown functions; and S. cerevisiae DSK2, involved in spindle pole body duplication. Identical residues are darkly shaded, whereas similar residues are lightly shaded. Only homology in five or more sequences are shown. All the proteins contain UB domains (square box) and UBA domains (rounded box), which are extremely well conserved in sequence. In addition, the central region between the UB and UBA domains contain several asparagine-proline repeats, which are either regularly spaced apart (closed circles) or located elsewhere throughout the protein (open circles). The high overall homology of the proteins suggests that they belong to the same multigene family.
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Related In: Results  -  Collection

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Figure 4: Ubiquilin shares significant homology with several other proteins. The inferred amino acid sequence of the human ubiquilin ORF and its homology to several related proteins: Homo sapiens Chap1, a protein involved in binding Hsp70-like Stch protein; Mus musculus PLIC-1 and PLIC-2, proteins involved in linking integrins to vimentin; X. laevis XDRP1, a protein which binds cyclin A; C. elegans F15C11.2 and two A. thaliana proteins, proteins with unknown functions; and S. cerevisiae DSK2, involved in spindle pole body duplication. Identical residues are darkly shaded, whereas similar residues are lightly shaded. Only homology in five or more sequences are shown. All the proteins contain UB domains (square box) and UBA domains (rounded box), which are extremely well conserved in sequence. In addition, the central region between the UB and UBA domains contain several asparagine-proline repeats, which are either regularly spaced apart (closed circles) or located elsewhere throughout the protein (open circles). The high overall homology of the proteins suggests that they belong to the same multigene family.
Mentions: The complete ubiquilin ORF consists of 595 residues, with a sequence rich in glutamines and serines, 10.9% and 11.1%, respectively, but lacking any cysteines (Fig. 4). The protein has a calculated isoelectric point (pI) of 4.97 and is predicted to be soluble, based on having a high hydrophilic profile using MacVector 6.5 software (Oxford Molecular Group; data not shown). Currently known protein structural motifs residing within ubiquilin include an NH2-terminal UB domain and a COOH-terminal ubiquitin-associated (UBA) domain, both of which have been implicated in targeting and degradation of proteins by the ubiquitin-proteasome pathway. The predicted ubiquilin protein also contains numerous regularly spaced asparagine-proline (Asn-Pro) repeats of unknown function (Fig. 4). When first sequenced, no apparent mammalian homologue for ubiquilin had been deposited in GenBank/EMBL/DDBJ databases. However, there was significant homology to several full-length genes of unknown function: the C. elegans F15C11.2 gene (36% identical) and two related genes in Arabidopsis thaliana (34 and 35% identical). In addition, the protein had weak homology to the Saccharomyces cerevisiae DSK2 gene (31% identical), which can suppress a KAR1 defect (Biggins et al. 1996). KAR1 is an essential gene involved in the duplication of the yeast microtubule organizing center known as the spindle pole body. Interestingly, ubiquilin homology to DSK2 was primarily confined to the UB and UBA domains. However, since both UB and UBA domains are present in many other proteins, it is uncertain whether ubiquilin and DSK2 are true orthologs.

Bottom Line: However, apart from a role in early development, neither the normal function of the presenilins nor the mechanisms by which mutant proteins cause AD are well understood.Moreover, the anti-ubiquilin antibodies robustly stained neurofibrillary tangles and Lewy bodies in AD and Parkinson's disease affected brains, respectively.Our results indicate that ubiquilin may be an important modulator of presenilin protein accumulation and that ubiquilin protein is associated with neuropathological neurofibrillary tangles and Lewy body inclusions in diseased brain.

View Article: PubMed Central - PubMed

Affiliation: Medical Biotechnology Center, Department of Neurology, University of Maryland Biotechnology Institute, Baltimore, Maryland 21201, USA.

ABSTRACT
Mutations in the highly homologous presenilin genes encoding presenilin-1 and presenilin-2 (PS1 and PS2) are linked to early-onset Alzheimer's disease (AD). However, apart from a role in early development, neither the normal function of the presenilins nor the mechanisms by which mutant proteins cause AD are well understood. We describe here the properties of a novel human interactor of the presenilins named ubiquilin. Yeast two-hybrid (Y2H) interaction, glutathione S-transferase pull-down experiments, and colocalization of the proteins expressed in vivo, together with coimmunoprecipitation and cell fractionation studies, provide compelling evidence that ubiquilin interacts with both PS1 and PS2. Ubiquilin is noteworthy since it contains multiple ubiquitin-related domains typically thought to be involved in targeting proteins for degradation. However, we show that ubiquilin promotes presenilin protein accumulation. Pulse-labeling experiments indicate that ubiquilin facilitates increased presenilin synthesis without substantially changing presenilin protein half-life. Immunohistochemistry of human brain tissue with ubiquilin-specific antibodies revealed prominent staining of neurons. Moreover, the anti-ubiquilin antibodies robustly stained neurofibrillary tangles and Lewy bodies in AD and Parkinson's disease affected brains, respectively. Our results indicate that ubiquilin may be an important modulator of presenilin protein accumulation and that ubiquilin protein is associated with neuropathological neurofibrillary tangles and Lewy body inclusions in diseased brain.

Show MeSH
Related in: MedlinePlus