ABCMdb

ABCG2 p.Thr362Ala

Predicted by SNAP2:	A: N (82%), C: N (82%), D: N (78%), E: N (82%), F: N (87%), G: N (82%), H: N (93%), I: N (78%), K: N (82%), L: N (82%), M: N (82%), N: N (87%), P: N (87%), Q: N (87%), R: N (78%), S: N (97%), V: N (87%), W: N (66%), Y: N (93%),
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: D, Y: N,

[switch to compact view]
Comments [show]

None has been submitted yet.

Publications
[hide] The 44-kDa Pim-1 kinase phosphorylates BCRP/ABCG2 ... J Biol Chem. 2008 Feb 8;283(6):3349-56. Epub 2007 Dec 5. Xie Y, Xu K, Linn DE, Yang X, Guo Z, Shimelis H, Nakanishi T, Ross DD, Chen H, Fazli L, Gleave ME, Qiu Y
The 44-kDa Pim-1 kinase phosphorylates BCRP/ABCG2 and thereby promotes its multimerization and drug-resistant activity in human prostate cancer cells.
J Biol Chem. 2008 Feb 8;283(6):3349-56. Epub 2007 Dec 5., 2008-02-08 [PMID:18056989]

Abstract [show]
We previously showed that the 44-kDa serine/threonine kinase Pim-1 (Pim-1L) can protect prostate cancer cells from apoptosis induced by chemotherapeutic drugs (Xie, Y., Xu, K., Dai, B., Guo, Z., Jiang, T., Chen, H., and Qiu, Y. (2006) Oncogene 25, 70-78). To further explore the mechanisms of Pim-1L-mediated resistance to chemotherapeutic drugs in prostate cancer cells, we employed a yeast two-hybrid screening to identify cellular proteins that were associated with Pim-1L, and we found the ABC transporter BCRP/ABCG2 as one of the potential interacting partners of Pim-1L. We also showed that the expression level of Pim-1L and BCRP was up-regulated in mitoxantrone and docetaxel-resistant prostate cancer cell lines. Pim-1L was co-localized with BCRP on the plasma membrane and induced phosphorylation of BCRP at threonine 362. Knocking-down Pim-1L expression in the drug-resistant prostate cancer cells abolished multimer formation of endogenous BCRP and resensitized the resistant cells to chemotherapeutic drugs suggesting that BCRP phosphorylation induced by Pim-1L was essential for its functionality. This is further corroborated by our finding that the plasma membrane localization and drug-resistant activity of BCRP were compromised by T362A mutation. Our data suggest that Pim-1L may protect prostate cancer cells from apoptosis, at least in part, through regulation of transmembrane drug efflux pump. These findings may provide a potential therapeutic approach by disrupting Pim-1 signaling to reverse BCRP-mediated multidrug resistance.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
3 Knocking-down Pim-1L expression in the drug-resistant prostate cancer cells abolished multimer formation of endogenous BCRP and resensitized the resistant cells to chemotherapeutic drugs suggesting that BCRP phosphorylation induced byPim-1Lwasessentialforitsfunctionality.Thisisfurthercorrob- orated by our finding that the plasma membrane localization and drug-resistant activity of BCRP were compromised by T362A mutation. Login to comment
50 To generate the BCRP T362A or T362D mutant, the threonine residue at position 362 was mutated to alanine or aspartic acid via oligonucleotide-directed mutagenesis with the sense primers 5Ј-GAAGAAGATCGCAGTCTTCAAGG-3Ј or 5Ј-GAAG- AAGATCGACGTCTTCAAGG-3Ј, respectively and their complementary antisense primers by using the QuikChange Mutagenesis kit (Stratagene). Login to comment
99 The stained slides of BCRP mutant T362A were examined by using an inverted fluorescence microscope under a ϫ60 oil-immersion objective, and the co-localization of BCRP with Pim-1L was scanned with a laser confocal system. Login to comment
150 Co-expression of BCRP and Pim-1L in 293T cells induced threonine phosphorylation of BCRP, and the substitution of Thr-362 with alanine (T362A) or the phosphorylation-mimicking aspartate (T362D) completely abolished threonine phosphorylation of BCRP, suggesting that Thr-362 may indeed be phosphorylated by Pim-1L in vivo (Fig. 3C). Login to comment
152 As shown in Fig. 3D, this antibody recognized the wild-type BCRP phosphorylated by Pim-1L but not the unphosphorylytable T362A mutant. Login to comment
157 To test whether phosphorylation of Thr-362 of BCRP is important for BCRP-mediated drug resistance, we infected LNCaP cells with the lentivirus encoding the HA-tagged BCRP or the T362A mutant and then examined whether LNCaP cells were protected by the overexpression of BCRP or its mutant from apoptosis induced by chemotherapeutic drugs MX, TPT, and DTX. Login to comment
158 As shown in Fig. 4A, the wild-type BCRP significantly increased the survival of LNCaP cells in response to these drugs, but the T362A mutant failed to do so, suggesting that the integrity of Thr-362 is essential for the functionality of BCRP. Login to comment
163 A, the effects of BCRP phosphorylation site mutant T362A on drug-resistant activity. Login to comment
171 To test whether Thr-362 phosphorylation plays a role for BCRP membrane localization, we infected LNCaP cells by the lentivirus encoding the wild-type BCRP or the T362A mutant. Login to comment
172 Immunofluorescence microscopy revealed that the wild-type BCRP and the phosphorylation- mimmicking mutant T362D were predominantly localized on plasma membrane, whereas the T362A mutant was mainly localized in cytoplasmic compartment (Fig. 5A). Login to comment
173 The retention of BCRP in the cytoplasmic compartment caused by the T362A mutation was further confirmed by fractionation experiments (Fig. 5B). Login to comment
175 We therefore examined the effects of T362A mutation on BCRP dimerization. Login to comment
176 We co-transfected the wild-type HA-tagged BCRP with the Myc-tagged BCRP or its T362A mutant into 293T cells. Login to comment
178 As shown in Fig. 5C, the wild-type Myc-BCRP was efficiently associated with the wild-type HA-BCRP but such interaction was disrupted by the T362A mutation. Login to comment
190 Effects of T362A mutation on the subcellular distribution and multimerization of BCRP in prostate cancer cells. Login to comment
191 A, T362A mutant predominately localizes in the intracellular compartment. Login to comment
192 LNCaP cells were treated with the lentivirus encoding the wild-type BCRP, T362A, or T362D mutant. Login to comment
197 LNCaP cells were treated with the lentivirus encoding the wild-type BCRP or T362A mutant, or the vector control. Login to comment
201 C, T362A mutation disrupts the dimerization of BCRP. Login to comment
100 The stained slides of BCRP mutant T362A were examined by using an inverted fluorescence microscope under a ϫ60 oil-immersion objective, and the co-localization of BCRP with Pim-1L was scanned with a laser confocal system. Login to comment
151 Co-expression of BCRP and Pim-1L in 293T cells induced threonine phosphorylation of BCRP, and the substitution of Thr-362 with alanine (T362A) or the phosphorylation-mimicking aspartate (T362D) completely abolished threonine phosphorylation of BCRP, suggesting that Thr-362 may indeed be phosphorylated by Pim-1L in vivo (Fig. 3C). Login to comment
153 As shown in Fig. 3D, this antibody recognized the wild-type BCRP phosphorylated by Pim-1L but not the unphosphorylytable T362A mutant. Login to comment
159 As shown in Fig. 4A, the wild-type BCRP significantly increased the survival of LNCaP cells in response to these drugs, but the T362A mutant failed to do so, suggesting that the integrity of Thr-362 is essential for the functionality of BCRP. Login to comment
164 A, the effects of BCRP phosphorylation site mutant T362A on drug-resistant activity. Login to comment
174 The retention of BCRP in the cytoplasmic compartment caused by the T362A mutation was further confirmed by fractionation experiments (Fig. 5B). Login to comment
177 We co-transfected the wild-type HA-tagged BCRP with the Myc-tagged BCRP or its T362A mutant into 293T cells. Login to comment
179 As shown in Fig. 5C, the wild-type Myc-BCRP was efficiently associated with the wild-type HA-BCRP but such interaction was disrupted by the T362A mutation. Login to comment
193 LNCaP cells were treated with the lentivirus encoding the wild-type BCRP, T362A, or T362D mutant. Login to comment
198 LNCaP cells were treated with the lentivirus encoding the wild-type BCRP or T362A mutant, or the vector control. Login to comment
202 C, T362A mutation disrupts the dimerization of BCRP. Login to comment

[hide] The Pim kinase inhibitor SGI-1776 decreases cell s... Biochem Pharmacol. 2013 Feb 15;85(4):514-24. doi: 10.1016/j.bcp.2012.12.006. Epub 2012 Dec 19. Natarajan K, Bhullar J, Shukla S, Burcu M, Chen ZS, Ambudkar SV, Baer MR
The Pim kinase inhibitor SGI-1776 decreases cell surface expression of P-glycoprotein (ABCB1) and breast cancer resistance protein (ABCG2) and drug transport by Pim-1-dependent and -independent mechanisms.
Biochem Pharmacol. 2013 Feb 15;85(4):514-24. doi: 10.1016/j.bcp.2012.12.006. Epub 2012 Dec 19., [PMID:23261525]

Abstract [show]
Overexpression of the ATP-binding cassette (ABC) drug efflux proteins P-glycoprotein (ABCB1) and breast cancer resistance protein (ABCG2) on malignant cells is associated with inferior chemotherapy outcomes. Both, ABCB1 and ABCG2, are substrates of the serine/threonine kinase Pim-1; Pim-1 knockdown decreases their cell surface expression, but SGI-1776, the first clinically tested Pim inhibitor, was shown to reverse drug resistance by directly inhibiting ABCB1-mediated transport. We sought to characterize Pim-1-dependent and -independent effects of SGI-1776 on drug resistance. SGI-1776 at the Pim-1-inhibitory and non-cytotoxic concentration of 1 muM decreased the IC(50)s of the ABCG2 and ABCB1 substrate drugs in cytotoxicity assays in resistant cells, with no effect on the IC(50) of non-substrate drug, nor in parental cells. SGI-1776 also increased apoptosis of cells overexpressing ABCG2 or ABCB1 exposed to substrate chemotherapy drugs and decreased their colony formation in the presence of substrate, but not non-substrate, drugs, with no effect on parental cells. SGI-1776 decreased ABCB1 and ABCG2 surface expression on K562/ABCB1 and K562/ABCG2 cells, respectively, with Pim-1 overexpression, but not HL60/VCR and 8226/MR20 cells, with lower-level Pim-1 expression. Finally, SGI-1776 inhibited uptake of ABCG2 and ABCB1 substrates in a concentration-dependent manner irrespective of Pim-1 expression, inhibited ABCB1 and ABCG2 photoaffinity labeling with the transport substrate [(125)I]iodoarylazidoprazosin ([(125)I]IAAP) and stimulated ABCB1 and ABCG2 ATPase activity. Thus SGI-1776 decreases cell surface expression of ABCB1 and ABCG2 and inhibits drug transport by Pim-1-dependent and -independent mechanisms, respectively. Decrease in ABCB1 and ABCG2 cell surface expression mediated by Pim-1 inhibition represents a novel mechanism of chemosensitization.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
22 Additionally, Pim-1 phosphorylates ABCG2 at threonine 362, knocking down Pim-1 abolishes ABCG2 multimer formation and ABCG2 T362A mutation decreases ABCG2 expression on the cell surface [18]. Login to comment

[hide] Insights into Chemoresistance of Prostate Cancer. Int J Biol Sci. 2015 Aug 1;11(10):1160-70. doi: 10.7150/ijbs.11439. eCollection 2015. Zhang W, Meng Y, Liu N, Wen XF, Yang T
Insights into Chemoresistance of Prostate Cancer.
Int J Biol Sci. 2015 Aug 1;11(10):1160-70. doi: 10.7150/ijbs.11439. eCollection 2015., [PMID:26327810]

Abstract [show]
Prostate cancer (PCa) remains the most prevalent malignancy among males in the western world. Though hormonal therapies through chemical or surgical castration have been proposed many years ago, heretofore, such mainstay for the treatment on advanced PCa has not fundamentally changed. These therapeutic responses are temporary and most cases will eventually undergo PCa recurrence and metastasis, or even progress to castration-resistant prostate cancer (CRPC) due to persistent development of drug resistance. Prostate cancer stem cells (PCSCs) are a small population of cells, which possess unlimited self-renewal capacities, and can regenerate tumorigenic progenies, and play an essential role in PCa therapy resistance, metastasis and recurrence. Nowadays advanced progresses have been made in understanding of PCSC properties, roles of androgen receptor signaling and ATP-binding cassette sub-family G member 2 (ABCG2), as well as roles of genomic non-coding microRNAs and key signaling pathways, which have led to the development of novel therapies which are active against chemoresistant PCa and CRPC. Based on these progresses, this review is dedicated to address mechanisms underlying PCa chemoresistance, unveil crosstalks among pivotal signaling pathways, explore novel biotherapeutic agents, and elaborate functional properties and specific roles of chemoresistant PCSCs, which may act as a promising target for novel therapies against chemoresistant PCa.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
108 Moreover, the plasma membrane localization and drug-resistant activity of ABCG2 were compromised by T362A mutation, suggesting ABCG2 phosphorylation induced by Pim-1L is essential for ABCG2 functionality (78). Login to comment