ABCA1 p.Thr1286Ala
| Predicted by SNAP2: | A: N (78%), C: N (82%), D: N (61%), E: N (78%), F: N (66%), G: N (72%), H: N (78%), I: N (72%), K: N (78%), L: N (72%), M: N (78%), N: N (82%), P: N (78%), Q: N (87%), R: N (78%), S: N (87%), V: N (78%), W: N (53%), Y: N (53%), |
| Predicted by PROVEAN: | A: N, C: N, D: N, E: N, F: N, G: N, H: N, I: N, K: N, L: N, M: N, N: N, P: N, Q: N, R: N, S: N, V: N, W: N, Y: N, |
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[hide] Phosphorylation of a pest sequence in ABCA1 promot... J Biol Chem. 2003 Sep 26;278(39):37368-74. Epub 2003 Jul 17. Martinez LO, Agerholm-Larsen B, Wang N, Chen W, Tall AR
Phosphorylation of a pest sequence in ABCA1 promotes calpain degradation and is reversed by ApoA-I.
J Biol Chem. 2003 Sep 26;278(39):37368-74. Epub 2003 Jul 17., [PMID:12869555]
Abstract [show]
ATP-binding cassette transporter A1 (ABCA1), the defective molecule in Tangier disease, mediates the apoAI-dependent efflux of excess cholesterol from cells. We recently showed that ABCA1 proteolysis by calpain was dependent on a PEST sequence in the cytoplasmic region of ABCA1 and was reversed by apoA-I interaction with ABCA1. We show here that phosphorylation of ABCA1 in HEK293 cells was reduced by 63 +/- 2.4% after removal of the PEST sequence (ABCA1delPEST) or by incubation of cells with apoAI (58 +/- 3.3%). By contrast, ABCA1delPEST showed no further decrease of phosphorylation upon apoAI treatment. To assess the hypothesis that PEST sequence phosphorylation could regulate ABCA1 calpain proteolysis, we mutagenized S/T residues in the PEST sequence and identified Thr-1286 and Thr-1305 as constitutively phosphorylated residues. The ABCA1-T1286A/T1305A mutant was not degraded by calpain and was not further stabilized upon apoA-I treatment. The T1286A/T1305A mutant showed a 3.1-fold increase in cell surface expression and a 2.3-fold increase of apoAI-mediated cholesterol efflux compared with wild type ABCA1. In conclusion, we propose a mechanism of regulation of ABCA1 cell surface expression and function in which the interaction with apoA-I results in dephosphorylation of the ABCA1 PEST sequence and thereby inhibits calpain degradation leading to an increase of ABCA1 cell surface expression.
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No. Sentence Comment
5 The ABCA1-T1286A/T1305A mutant was not degraded by calpain and was not further stabilized upon apoA-I treatment.
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ABCA1 p.Thr1286Ala 12869555:5:10
status: NEW6 The T1286A/T1305A mutant showed a 3.1-fold increase in cell surface expression and a 2.3-fold increase of apoAI-mediated cholesterol efflux compared with wild type ABCA1.
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ABCA1 p.Thr1286Ala 12869555:6:4
status: NEW31 mABCA1-FLAG mutation constructs were further called: MutAAAA for mutation to Ala on T1286A/S1296A/S1302A/T1305A, MutTAAT for mutation to Ala on S1296A/S1302A, and MutASSA for mutation to Ala on * This work was supported in part by National Institutes of Health Grant HL22682.
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ABCA1 p.Thr1286Ala 12869555:31:84
status: NEW41 Printed in U.S.A. This paper is available on line at http://www.jbc.org T1286A/T1305A.
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ABCA1 p.Thr1286Ala 12869555:41:74
status: NEW121 Mutation of Thr residues (1286 and 1305) decreases ABCA1 Phosphorylation. WT-mABCA1-FLAG, mABCA1delPEST-FLAG, mABCA1-FLAG mutated to Ala on T1286A/S1296A/S1302A/T1305A (MutAAAA), mutated to Ala on S1296A/S1302A (MutTAAT), or mutated to Ala on T1286A/T1305A (MutASSA), were transiently transfected in 6 wells of HEK-293 cells (2 g of plasmid DNA per well).
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ABCA1 p.Thr1286Ala 12869555:121:140
status: NEWX
ABCA1 p.Thr1286Ala 12869555:121:243
status: NEW150 ABCA1 mutated on Thr-1286 and Thr-1305 is not degraded by calpain and is not stabilized by apoA-I. WT-mABCA1-FLAG or mABCA1-FLAG mutated to Ala on T1286A/T1305A (MutTSSA) were transiently transfected into HEK293 cells.
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ABCA1 p.Thr1286Ala 12869555:150:147
status: NEW189 Effect of apigenin (CK2 inhibitor) and H-89 (PKA inhibitor) on ABCA1 phosphorylation. WT-mABCA1-FLAG or mABCA1-FLAG mutated to Ala on T1286A/T1305A (MutASSA) were transiently transfected in 6 wells of HEK-293 cells (2 g of plasmid DNA per well).
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ABCA1 p.Thr1286Ala 12869555:189:134
status: NEW30 mABCA1-FLAG mutation constructs were further called: MutAAAA for mutation to Ala on T1286A/S1296A/S1302A/T1305A, MutTAAT for mutation to Ala on S1296A/S1302A, and MutASSA for mutation to Ala on * This work was supported in part by National Institutes of Health Grant HL22682.
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ABCA1 p.Thr1286Ala 12869555:30:84
status: NEW38 39, Issue of September 26, pp. 37368-37374, 2003 (c) 2003 by The American Society for Biochemistry and Molecular Biology, Inc. Printed in U.S.A. This paper is available on line at http://www.jbc.org T1286A/T1305A.
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ABCA1 p.Thr1286Ala 12869555:38:201
status: NEW118 Mutation of Thr residues (1286 and 1305) decreases ABCA1 Phosphorylation. WT-mABCA1-FLAG, mABCA1delPEST-FLAG, mABCA1-FLAG mutated to Ala on T1286A/S1296A/S1302A/T1305A (MutAAAA), mutated to Ala on S1296A/S1302A (MutTAAT), or mutated to Ala on T1286A/T1305A (MutASSA), were transiently transfected in 6 wells of HEK-293 cells (2 òe;g of plasmid DNA per well).
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ABCA1 p.Thr1286Ala 12869555:118:140
status: NEWX
ABCA1 p.Thr1286Ala 12869555:118:243
status: NEW147 ABCA1 mutated on Thr-1286 and Thr-1305 is not degraded by calpain and is not stabilized by apoA-I. WT-mABCA1-FLAG or mABCA1-FLAG mutated to Ala on T1286A/T1305A (MutTSSA) were transiently transfected into HEK293 cells.
X
ABCA1 p.Thr1286Ala 12869555:147:147
status: NEW186 Effect of apigenin (CK2 inhibitor) and H-89 (PKA inhibitor) on ABCA1 phosphorylation. WT-mABCA1-FLAG or mABCA1-FLAG mutated to Ala on T1286A/T1305A (MutASSA) were transiently transfected in 6 wells of HEK-293 cells (2 òe;g of plasmid DNA per well).
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ABCA1 p.Thr1286Ala 12869555:186:134
status: NEW