Cloning, purification, crystallization and preliminary X-ray crystallographic analysis of SET/TAF-Iβ ΔN from<i>Homo sapiens</i>

Xu, Zhen; Yang, Weili; Shi, Nuannuan; Gao, Yongxiang; Teng, Maikun; Niu, Liwen

doi:10.1107/s1744309110021779

Cited by 2 publications

(2 citation statements)

References 20 publications

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“…Recent data revealed that an alternative mechanism of ceramide-induced activation of PP2A occurs through direct binding of ceramide with SET (32). In silico modeling based on partial crystal structures (33) and ceramide transporter-lipid-binding domains (34) suggests that SET possesses a hydrophobic ceramide-binding pocket. This lipid-binding pocket has a higher affinity for D-erythro (e)-C18 ceramide (9,32) compared with other endogenous ceramides or sphingosine.…”

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confidence: 99%

The NMR‐based characterization of the FTY720‐SET complex reveals an alternative mechanism for the attenuation of the inhibitory SET‐PP2A interaction

Palma

Parnham

et al. 2019

FASEB j.

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The su(var)3‐9, enhancer of zeste, trithorax (SET)/inhibitor 2 of protein phosphatase 2A (PP2A) oncoprotein binds and inhibits PP2A, composed of various isoforms of scaffolding, regulatory, and catalytic subunits. Targeting SET with a sphingolipid analog drug fingolimod (FTY720) or ceramide leads to the reactivation of tumor suppressor PP2A. However, molecular details of the SET‐FTY720 or SET‐ceramide, and mechanism of FTY720‐dependent PP2A activation, remain unknown. Here, we report the first in solution examination of the SET‐FTY720 or SET‐ceramide complexes by NMR spectroscopy. FTY720‐ceramide binding resulted in chemical shifts of residues residing at the N terminus of SET, preventing its dimerization or oligomerization. This then released SET from PP2ACα, resulting in PP2A activation, while monomeric SET remained associated with the B56γ. Our data also suggest that the PP2A holoenzyme, composed of PP2A‐Aβ, PP2A‐B56γ, and PP2ACα subunits, is selectively activated in response to the formation of the SET‐FTY720 complex in A549 cells. Various PP2A‐associated downstream effector proteins in the presence or absence of FTY720 were then identified by stable isotope labeling with amino cells in cell culture, including tumor suppressor nonmuscle myosin IIA. Attenuation of FTY720‐SET association by point mutations of residues that are involved in FTY720 binding or dephosphorylation of SET at Serine 171, enhanced SET oligomerization and the formation of the SET‐PP2A inhibitory complex, leading to resistance to FTY720‐dependent PP2A activation.—De Palma, R. M., Parnham, S. R., Li, Y., Oaks, J. J., Peterson, Y. K., Szulc, Z. M., Roth, B. M., Xing, Y., Ogretmen, B. The NMR‐based characterization of the FTY720‐SET complex reveals an alternative mechanism for the attenuation of the inhibitory SET‐PP2A interaction. FASEB J. 33, 7647–7666 (2019). http://www.fasebj.org

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confidence: 99%

The NMR‐based characterization of the FTY720‐SET complex reveals an alternative mechanism for the attenuation of the inhibitory SET‐PP2A interaction

Palma

Parnham

et al. 2019

FASEB j.

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“…It is a multifunctional protein that interacts with other proteins in the regulation of cellular signaling ( 12 ). SET has been described as an oncogene that regulates important signaling pathways, with SET reported to have roles in inhibiting the DNase activity of tumor suppressor NM23-H1, increasing AP-1 activity, activating MAPK signaling, regulating granzyme B ( 13 ) and producing IFN-γ in human NK cells. All these functions are involved in SET-regulated cell apoptosis, cell cycle progression and cell mortality ( 14 ) through cell processes, such as DNA replication, chromatin remodeling, gene transcription, differentiation and migration ( 15 ).…”

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confidence: 99%

Association between SET expression and glioblastoma cell apoptosis and proliferation

Shi

et al. 2016

Oncology Letters

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Glioblastoma multiforme (GBM) was one of the first cancer types systematically studied at a genomic and transcriptomic level due to its high incidence and aggressivity; however, the detailed mechanism remains unclear, even though it is known that numerous cytokines are involved in the occurrence and development of GBM. The present study aimed to determine whether the SET gene has a role in human glioblastoma carcinogenesis. A total of 32 samples, including 18 cases of glioma, 2 cases of meningioma and 12 normal brain tissue samples, were detected using the streptavidin-peroxidase method through immunohistochemistry. To reduce SET gene expression in U251 and U87MG cell lines, the RNA interference technique was used and transfection with small interfering (si)RNA of the SET gene was performed. Cell apoptosis was detected by flow cytometry, cell migration was examined by Transwell migration assay and cell proliferation was determined by Cell Counting Kit-8. SET, Bcl-2, Bax and caspase-3 mRNA and protein expression levels were detected by reverse transcription-quantitative polymerase chain reaction and western blot analysis, respectively. Positive protein expression of SET was observed in the cell nucleus, with the expression level of SET significantly higher in glioma tissues compared with normal brain tissue (P=0.001). Elevated expression of SET was significantly associated with gender (P=0.002), tumors classified as World Health Organization grade II (P=0.031), III (P=0.003) or IV (P=0.001), and moderately (P=0.031) or poorly differentiated (P=0.001) tumors. Compared with the negative and non-treatment (blank) control cells, SET gene expression was significantly inhibited (P=0.006 and P<0.001), cell apoptosis was significantly increased (P=0.001 and P<0.001), cell proliferation was significantly inhibited (P=0.002 and P=0.015), and cell migration was significantly decreased (P=0.001 and P=0.001) in siRNA-transfected U87MG−SET and U251−SET cells, respectively. In addition, mRNA and protein expression levels of Bcl-2 were significantly inhibited in U87MG−SET and U251−SET cells, while mRNA and protein expression levels of Bax and caspase-3 were significantly increased, compared with the two control groups. Thus, the current data suggests that SET may regulate the proliferation and apoptosis of glioblastoma cells by upregulating Bcl-2, and downregulating Bax and caspase-3.

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Cloning, purification, crystallization and preliminary X-ray crystallographic analysis of SET/TAF-Iβ ΔN fromHomo sapiens

Cited by 2 publications

References 20 publications

The NMR‐based characterization of the FTY720‐SET complex reveals an alternative mechanism for the attenuation of the inhibitory SET‐PP2A interaction

The NMR‐based characterization of the FTY720‐SET complex reveals an alternative mechanism for the attenuation of the inhibitory SET‐PP2A interaction

Association between SET expression and glioblastoma cell apoptosis and proliferation

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