Mustafa Gokhan ERTOSUN, Suray PEHLİVANOGLU, Sayra DİLMAC, Gamze TANRİOVER, Osman Nidai OZES

AKT-mediated phosphorylation of TWIST1 is essential for breast cancer cell metastasis

Previously, it was shown that human TWIST1 (basic helix-loop-helix (b-HLH) is phosphorylated by Akt kinase at S42, T121, and S123. To show in vivo effect of these phosphorylations, we created mouse TWIST1 expression vector and converted the codons of S42, T125, and S127 to unphosphorylatable alanine and phosphorylation mimicking Glutamic acid. We hypothesized that alanine mutants would inhibit the metastatic ability of 4T1 cells while glutamic acid mutants would convert nonmetastatic 67NR cells into metastatic phenotype. To confirm this hypothesis, we created metastatic 4T1 and nonmetastatic 67NR cells expressing alanine mutants and glutamic acid mutants mouse TWIST1, respectively. Then, we injected 1 x 10(6) 67NR and 1 x 10(5) 4T1 cells overexpressing mutants of TWIST1 into the breast tissue of BALB/c mice. At the end of the 4th week, we sacrificed the animals, determined the numbers of tumors at lungs and liver. Although 67NR cells overexpressing wild-type TWIST1 did not show any metastasis, cells overexpressing S42E and T125E mutants showed 15-30 macroscopic metastasis to liver and lungs. Parallel to this, 4T1 cells expressing S42A and T125A mutants of TWIST1 showed no macroscopic metastasis. Our results indicate that phosphorylation of S42 and T125 by AKT is essential for TWIST1-mediated tumor growth and metastasis.

Keywords:

TWIST1, breast cancer, metastasis,

PDF

___

Alexander NR, 2006, CANCER RES, V66, P3365, DOI 10.1158/0008-5472.CAN-05-3401
Bialek P, 2004, DEV CELL, V6, P423, DOI 10.1016/S1534-5807(04)00058-9
CHEN ZF, 1995, GENE DEV, V9, P686, DOI 10.1101/gad.9.6.686
Cheng GZ, 2008, CANCER RES, V68, P957, DOI 10.1158/0008-5472.CAN-07-5067
Cheng GZ, 2007, CANCER RES, V67, P1979, DOI 10.1158/0008-5472.CAN-06-1479
Connerney J, 2006, DEV DYNAM, V235, P1345, DOI 10.1002/dvdy.20717
ElGhouzzi V, 1997, NAT GENET, V15, P42 .
Elias MC, 2005, NEOPLASIA, V7, P824, DOI 10.1593/neo.04352
Funato N, 2001, MOL CELL BIOL, V21, P7416, DOI 10.1128/MCB.21.21.7416-7428.2001
Gong XQ, 2002, J BIOL CHEM, V277, P12310, DOI 10.1074/jbc.M110228200
Howard TD, 1997, NAT GENET, V15, P36, DOI 10.1038/ng0197-36
Kingston RE, 2003, CALCIUM PHOSPHATE TR, V20, P20
Kyo S, 2006, HUM PATHOL, V37, P431, DOI 10.1016/j.humpath.2005.12.021
Lee TK, 2006, CLIN CANCER RES, V12, P5369, DOI 10.1158/1078-0432.CCR-05-2722
Mironchik Y, 2005, CANCER RES, V65, P10801, DOI 10.1158/0008-5472.CAN-05-0712
Qin Q, 2012, CELL RES, V22, P90, DOI 10.1038/cr.2011.144
Rice DPC, 2000, DEVELOPMENT, V127, P1845
Singh S, 2014, BIOMED RES INT, V2014, DOI 10.1155/2014/215748
Wang DW, 2018, ONCOL REP, V39, P31, DOI 10.3892/or.2017.6076
Xu Y, 2017, ONCOGENE, V36, P1157, DOI 10.1038/onc.2016.286
Xu Y, 2013, AM J PATHOL, V183, P1281, DOI 10.1016/j.ajpath.2013.06.021
Xue GD, 2012, CANCER DISCOV, V2, P248, DOI 10.1158/2159-8290.CD-11-0270
Yousfi M, 2002, BIOCHEM BIOPH RES CO, V297, P641, DOI 10.1016/S0006-291X(02)02260-X
Zhang YQ, 2007, PATHOLOGY, V39, P470, DOI 10.1080/00313020701570053