The role of glucocorticoid receptor phosphorylation in Mcl-1 and NOXA gene expression.
BACKGROUND
The cyclin-dependent kinase (CDK) and mitogen-activated protein kinase (MAPK) mediated phosphorylation of glucocorticoid receptor (GR) exerts opposite effects on GR transcriptional activity and affects other posttranslational modifications within this protein. The major phosphorylation site of human GR targeted by MAPK family is the serine 226 and multiple kinase complexes phosphorylate receptor at the serine 211 residue. We hypothesize that GR posttranslational modifications are involved in the determination of the cellular fate in human lymphoblastic leukemia cells. We investigated whether UV signalling through alternative GR phosphorylation determined the cell type specificity of glucocorticoids (GCs) mediated apoptosis.


RESULTS
We have identified putative Glucocorticoid Response Elements (GREs) within the promoter regulatory regions of the Bcl-2 family members NOXA and Mcl-1 indicating that they are direct GR transcriptional targets. These genes were differentially regulated in CEM-C7-14, CEM-C1-15 and A549 cells by glucocorticoids and JNK pathway. In addition, our results revealed that the S211 phosphorylation was dominant in CEM-C7-14, whereas the opposite was the case in CEM-C1-15 where prevalence of S226 GR phosphorylation was observed. Furthermore, multiple GR isoforms with cell line specific patterns were identified in CEM-C7-14 cells compared to CEM-C1-15 and A549 cell lines with the same antibodies.


CONCLUSIONS
GR phosphorylation status kinetics, and site specificity as well as isoform variability differ in CEM-C7-14, CEM-C1-15, and A549 cells. The positive or negative response to GCs induced apoptosis in these cell lines is a consequence of the variable equilibrium of NOXA and Mcl-1 gene expression potentially mediated by alternatively phosphorylated GR, as well as the balance of MAPK/CDK pathways controlling GR phosphorylation pattern. Our results provide molecular base and valuable knowledge for improving the GC based therapies of leukaemia.
School of Pharmacy and Pharmaceutical Sciences, The University of Manchester, Manchester, UK.
James T
JT
Lynch
Ramkumar
R
Rajendran
Georgia
G
Xenaki
Ilhem
I
Berrou
Constantinos
C
Demonacos
Marija
M
Krstic-Demonacos
D017209:Apoptosis / Q000187:drug effects / Q000528:radiation effects; D001483:Base Sequence; D045744:Cell Line, Tumor; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology; D016193:G1 Phase / Q000187:drug effects / Q000528:radiation effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000528:radiation effects; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008969:Molecular Sequence Data; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009928:Organ Specificity / Q000187:drug effects / Q000528:radiation effects; D010766:Phosphorylation / Q000187:drug effects / Q000528:radiation effects; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D014466:Ultraviolet Rays
D064549:Myeloid Cell Leukemia Sequence 1 Protein; C498702:PMAIP1 protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D011965:Receptors, Glucocorticoid; D003907:Dexamethasone; D018844:Cyclin-Dependent Kinases; D048031:JNK Mitogen-Activated Protein Kinases