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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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Intracellular colocalization between ubiquilin and the presenilins. (A–D) HeLa cells were cotransfected with ubiquilin and either (A) wild-type PS1, (B) wild-type PS2, (C) PS2(ΔC) deletion mutant, or (D) PS2(ΔLC) deletion mutant and costained with appropriate goat anti-presenilin antibodies (left images) and affinity-purified rabbit anti–ubiquilin-C antibody (center images). The green (fluorescein) and red (rhodamine) confocal images in each row were merged and shown on the right, with yellow indicating colocalization of ubiquilin and presenilin proteins. Bar, 10 μm.
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Figure 7: Intracellular colocalization between ubiquilin and the presenilins. (A–D) HeLa cells were cotransfected with ubiquilin and either (A) wild-type PS1, (B) wild-type PS2, (C) PS2(ΔC) deletion mutant, or (D) PS2(ΔLC) deletion mutant and costained with appropriate goat anti-presenilin antibodies (left images) and affinity-purified rabbit anti–ubiquilin-C antibody (center images). The green (fluorescein) and red (rhodamine) confocal images in each row were merged and shown on the right, with yellow indicating colocalization of ubiquilin and presenilin proteins. Bar, 10 μm.

Mentions: As overexpressed presenilin proteins are localized predominantly to the ER, and ubiquilin protein in HeLa cells is found throughout the nucleus and cytoplasm, we determined if ubiquilin colocalized with the presenilins when the latter were overexpressed. Indeed, there was a dramatic change in ubiquilin staining in HeLa cells coexpressing either PS1 or PS2 and ubiquilin, resulting in almost complete colocalization of the presenilin and ubiquilin intracellular staining patterns, as seen by laser confocal immunofluorescence microscopy (Fig. 7A and Fig. B). Careful examination of the colocalized proteins revealed ubiquilin staining to be punctate, whereas presenilin staining could be traced to an ER-like pattern. Particularly striking was colocalization of ubiquilin with irregularly shaped presenilin-enriched structures within the cytoplasm of overexpressing cells. In contrast, there was little colocalization in the nucleus where ubiquilin, but not presenilin, staining could be found.


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

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

Intracellular colocalization between ubiquilin and the presenilins. (A–D) HeLa cells were cotransfected with ubiquilin and either (A) wild-type PS1, (B) wild-type PS2, (C) PS2(ΔC) deletion mutant, or (D) PS2(ΔLC) deletion mutant and costained with appropriate goat anti-presenilin antibodies (left images) and affinity-purified rabbit anti–ubiquilin-C antibody (center images). The green (fluorescein) and red (rhodamine) confocal images in each row were merged and shown on the right, with yellow indicating colocalization of ubiquilin and presenilin proteins. Bar, 10 μm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2169435&req=5

Figure 7: Intracellular colocalization between ubiquilin and the presenilins. (A–D) HeLa cells were cotransfected with ubiquilin and either (A) wild-type PS1, (B) wild-type PS2, (C) PS2(ΔC) deletion mutant, or (D) PS2(ΔLC) deletion mutant and costained with appropriate goat anti-presenilin antibodies (left images) and affinity-purified rabbit anti–ubiquilin-C antibody (center images). The green (fluorescein) and red (rhodamine) confocal images in each row were merged and shown on the right, with yellow indicating colocalization of ubiquilin and presenilin proteins. Bar, 10 μm.
Mentions: As overexpressed presenilin proteins are localized predominantly to the ER, and ubiquilin protein in HeLa cells is found throughout the nucleus and cytoplasm, we determined if ubiquilin colocalized with the presenilins when the latter were overexpressed. Indeed, there was a dramatic change in ubiquilin staining in HeLa cells coexpressing either PS1 or PS2 and ubiquilin, resulting in almost complete colocalization of the presenilin and ubiquilin intracellular staining patterns, as seen by laser confocal immunofluorescence microscopy (Fig. 7A and Fig. B). Careful examination of the colocalized proteins revealed ubiquilin staining to be punctate, whereas presenilin staining could be traced to an ER-like pattern. Particularly striking was colocalization of ubiquilin with irregularly shaped presenilin-enriched structures within the cytoplasm of overexpressing cells. In contrast, there was little colocalization in the nucleus where ubiquilin, but not presenilin, staining could be found.

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