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Characterization of SET/I2PP2A isoforms in dogs.

Yabe R, Fujiwara N, Mizuno T, Usui T, Ohama T, Sato K - J. Vet. Med. Sci. (2014)

Bottom Line: SET is an endogenous protein phosphatase 2A (PP2A) inhibitor and is associated with a poor prognosis in human leukemia.Here, we found that canine cells express several isoforms of the SET protein.We confirmed that SETα and β possess the ability to associate with PP2A.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Veterinary Pharmacology, Joint Faculty of Veterinary Medicine, Yamaguchi University, Yamaguchi 753-8515, Japan.

ABSTRACT
SET is an endogenous protein phosphatase 2A (PP2A) inhibitor and is associated with a poor prognosis in human leukemia. Previously, we reported increased SET protein levels in canine lymphoma cell lines and the potential therapeutic application of SET antagonists in canine lymphoma. Here, we found that canine cells express several isoforms of the SET protein. We cloned 4 isoforms of SET, named SETα, β, γ and δ. Genomic BLAST showed that the SET genes are located on chromosomes X, 7, 1 and 8, respectively. An immunofluorescent study showed nuclear localization of SETα and β, and nuclear and cytosolic localization of SETγ and δ. We confirmed that SETα and β possess the ability to associate with PP2A. Our data reveal the existence of unique SET isoforms that should be taken into account in SET-targeting drug development studies in dogs.

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Related in: MedlinePlus

Expression and localization of canine SET isoforms. (A) HEK293T cells weretransfected to express FLAGx3-tagged canine SET isoforms, and protein expression wasanalyzed by immunoblotting. VCP was used as a loading control. (B) The localization ofendogenous SET in CMeC1 cells was examined by immunofluorescent staining usinganti-SET (N-term) and anti-SET (Middle) (Green). Phalloidin (Red) and Hoechst (Blue)were used to stain F-actin and the nucleus. (C) HeLa cells were transfected to expressFLAGx3-tagged canine SET isoforms and ZsGreen, and the localization of SET isoformswas determined by immunofluorescent staining using an anti-FLAG antibody (Red).Hoechst (Blue) was used to stain the nucleus.
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fig_004: Expression and localization of canine SET isoforms. (A) HEK293T cells weretransfected to express FLAGx3-tagged canine SET isoforms, and protein expression wasanalyzed by immunoblotting. VCP was used as a loading control. (B) The localization ofendogenous SET in CMeC1 cells was examined by immunofluorescent staining usinganti-SET (N-term) and anti-SET (Middle) (Green). Phalloidin (Red) and Hoechst (Blue)were used to stain F-actin and the nucleus. (C) HeLa cells were transfected to expressFLAGx3-tagged canine SET isoforms and ZsGreen, and the localization of SET isoformswas determined by immunofluorescent staining using an anti-FLAG antibody (Red).Hoechst (Blue) was used to stain the nucleus.

Mentions: To examine whether canine SET isoforms are conformationally stable, HEK293T cells weretransfected to express the canine SET isoforms, and protein levels were determined byimmunoblotting (Fig. 4AFig. 4.


Characterization of SET/I2PP2A isoforms in dogs.

Yabe R, Fujiwara N, Mizuno T, Usui T, Ohama T, Sato K - J. Vet. Med. Sci. (2014)

Expression and localization of canine SET isoforms. (A) HEK293T cells weretransfected to express FLAGx3-tagged canine SET isoforms, and protein expression wasanalyzed by immunoblotting. VCP was used as a loading control. (B) The localization ofendogenous SET in CMeC1 cells was examined by immunofluorescent staining usinganti-SET (N-term) and anti-SET (Middle) (Green). Phalloidin (Red) and Hoechst (Blue)were used to stain F-actin and the nucleus. (C) HeLa cells were transfected to expressFLAGx3-tagged canine SET isoforms and ZsGreen, and the localization of SET isoformswas determined by immunofluorescent staining using an anti-FLAG antibody (Red).Hoechst (Blue) was used to stain the nucleus.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4197151&req=5

fig_004: Expression and localization of canine SET isoforms. (A) HEK293T cells weretransfected to express FLAGx3-tagged canine SET isoforms, and protein expression wasanalyzed by immunoblotting. VCP was used as a loading control. (B) The localization ofendogenous SET in CMeC1 cells was examined by immunofluorescent staining usinganti-SET (N-term) and anti-SET (Middle) (Green). Phalloidin (Red) and Hoechst (Blue)were used to stain F-actin and the nucleus. (C) HeLa cells were transfected to expressFLAGx3-tagged canine SET isoforms and ZsGreen, and the localization of SET isoformswas determined by immunofluorescent staining using an anti-FLAG antibody (Red).Hoechst (Blue) was used to stain the nucleus.
Mentions: To examine whether canine SET isoforms are conformationally stable, HEK293T cells weretransfected to express the canine SET isoforms, and protein levels were determined byimmunoblotting (Fig. 4AFig. 4.

Bottom Line: SET is an endogenous protein phosphatase 2A (PP2A) inhibitor and is associated with a poor prognosis in human leukemia.Here, we found that canine cells express several isoforms of the SET protein.We confirmed that SETα and β possess the ability to associate with PP2A.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Veterinary Pharmacology, Joint Faculty of Veterinary Medicine, Yamaguchi University, Yamaguchi 753-8515, Japan.

ABSTRACT
SET is an endogenous protein phosphatase 2A (PP2A) inhibitor and is associated with a poor prognosis in human leukemia. Previously, we reported increased SET protein levels in canine lymphoma cell lines and the potential therapeutic application of SET antagonists in canine lymphoma. Here, we found that canine cells express several isoforms of the SET protein. We cloned 4 isoforms of SET, named SETα, β, γ and δ. Genomic BLAST showed that the SET genes are located on chromosomes X, 7, 1 and 8, respectively. An immunofluorescent study showed nuclear localization of SETα and β, and nuclear and cytosolic localization of SETγ and δ. We confirmed that SETα and β possess the ability to associate with PP2A. Our data reveal the existence of unique SET isoforms that should be taken into account in SET-targeting drug development studies in dogs.

Show MeSH
Related in: MedlinePlus