Pim-1 kinase protects P-glycoprotein from degradation and enables its glycosylation and cell surface expression.
The oncogenic serine/threonine kinase Pim-1 phosphorylates and activates the ATP-binding cassette transporter breast cancer resistance protein (ABCG2). The ABC transporter P-glycoprotein (Pgp; ABCB1) also contains a Pim-1 phosphorylation consensus sequence, and we hypothesized that Pim-1 also regulates Pgp. Pgp is exported from the endoplasmic reticulum (ER) as a 150-kDa species that is glycosylated to 170-kDa Pgp, translocates to the cell surface, and mediates drug efflux; alternatively, 150-kDa Pgp is cleaved to a 130-kDa proteolytic product by ER proteases or undergoes ubiquitination and proteasomal degradation. Pim-1 and Pgp interaction was studied in GST pull-down and phosphorylation in in vitro kinase assays. Pim-1 knockdown and inhibition effects on Pgp expression were studied by immunoblotting and flow cytometry and on Pgp stability by immunoblotting after cycloheximide treatment. Pim-1 directly interacted with and phosphorylated Pgp in intact cells and in vitro. Pim-1 knockdown or inhibition decreased cellular and cell surface 170-kDa Pgp, in association with both transient increase in 130-kDa Pgp and increased Pgp ubiquitination and proteasomal degradation. Pim-1 inhibition also decreased expression of 150-kDa Pgp in the presence of the glycosylation inhibitor 2-deoxy-d-glucose. Finally, Pim-1 inhibition sensitized Pgp-overexpressing cells to doxorubicin. Thus, Pim-1 regulates Pgp expression by protecting 150-kDa Pgp from proteolytic and proteasomal degradation and enabling Pgp glycosylation and cell surface translocation and thus Pgp-mediated drug efflux. Pim-1 inhibitors are entering clinical trials and may provide a novel approach to abrogating drug resistance.
Xie
Yingqiu
Y
University of Maryland Greenebaum Cancer Center, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
Burcu
Mehmet
M
Linn
Douglas E
DE
Qiu
Yun
Y
Baer
Maria R
MR
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D005434:Flow Cytometry; D055785:Gene Knockdown Techniques; D006031:Glycosylation; D018922:HL-60 Cells; D006801:Humans; D006868:Hydrolysis; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D051573:Proto-Oncogene Proteins c-pim-1 / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D003513:Cycloheximide; D051573:Proto-Oncogene Proteins c-pim-1; D046988:Proteasome Endopeptidase Complex
Mol Cancer Ther. 2007 Jan;6(1):163-72
17218638
EMBO J. 1991 Mar;10(3):655-64
1825810
Oncogene. 2008 Aug 14;27(35):4809-19
18438430
Cancer Res. 2007 Jul 15;67(14):6916-24
17638903
Proc Natl Acad Sci U S A. 2002 Feb 19;99(4):2175-80
11854514
J Biol Chem. 1999 Jun 25;274(26):18659-66
10373478
J Biol Chem. 1998 Dec 4;273(49):32373-6
9829963
J Biol Chem. 2005 Dec 16;280(50):41675-82
16227208
Nature. 2001 Aug 23;412(6849):822-6
11518967
Oncogene. 2006 Jan 5;25(1):70-8
16186805
Mol Pharmacol. 2004 Sep;66(3):395-403
15322230
FEBS Lett. 2004 Jul 30;571(1-3):43-9
15280015
Oncogene. 2004 Mar 11;23(10):1838-44
14981536
J Biol Chem. 1997 Jan 10;272(2):709-12
8995353
Blood. 2002 Aug 15;100(4):1224-32
12149202
Bioorg Med Chem Lett. 2007 Mar 15;17(6):1679-83
17251021
J Biol Chem. 1996 Jun 7;271(23):13668-74
8662768
J Biol Chem. 2001 Mar 23;276(12):8657-64
11121420
J Biol Chem. 2008 Feb 8;283(6):3349-3356
18056989
Blood. 2009 Nov 5;114(19):4150-7
19734450
Biochemistry. 2000 Jan 11;39(1):194-204
10625494
Cancer Res. 1999 Mar 1;59(5):1021-8
10070958
J Biol Chem. 1996 Jan 19;271(3):1708-16
8576173
Biochem J. 1994 Apr 1;299 ( Pt 1):309-15
7909431
Proc Natl Acad Sci U S A. 1989 Nov;86(22):8857-61
2682662
Clin Cancer Res. 2004 Mar 1;10(5):1826-34
15014037
Curr Drug Deliv. 2004 Jan;1(1):43-56
16305369
Mol Cancer Res. 2007 Sep;5(9):909-22
17855660
Cancer Lett. 2007 Jan 8;245(1-2):350-2
16504380
Biochem Cell Biol. 1997;75(2):153-62
9250363
Mol Cancer Ther. 2009 Oct;8(10):2882-93
19825806
Br J Haematol. 2006 Sep;134(5):500-9
16869825
J Membr Biol. 2000 Feb 1;173(3):203-14
10667916
Cancer Res. 2008 Jul 1;68(13):5076-85
18593906
J Cell Biochem. 2008 Nov 1;105(4):1048-58
18767071
Curr Cancer Drug Targets. 2005 Sep;5(6):445-55
16178818
Int J Biochem Cell Biol. 2006 Mar;38(3):430-43
16356754
Clin Cancer Res. 2005 Mar 15;11(6):2320-6
15788683
J Clin Oncol. 2005 Oct 10;23(29):7286-95
16145069
Blood. 2005 Oct 15;106(8):2646-54
15994288
J Biol Chem. 1998 Jun 12;273(24):14671-4
9614062
FASEB J. 1999 Oct;13(13):1724-32
10506575