Mutational analysis of progesterone receptor functional domains in stable cell lines delineates sets of genes regulated by different mechanisms.
Steroid hormone receptors act directly in the nucleus on the chromatin organization and transcriptional activity of several promoters. Furthermore, they have an indirect effect on cytoplasmic signal transduction pathways, including MAPK, impacting ultimately on gene expression. We are interested in distinguishing between the two modes of action of progesterone receptor (PR) on the control of gene expression and cell proliferation. For this, we have stably expressed, in PR-negative breast cancer cells, tagged forms of the PR isoform B mutated at regions involved either in DNA binding (DNA-binding domain) or in its ability to interact with the estrogen receptor and to activate the c-Src/MAPK/Erk/Msk cascade (estrogen receptor-interacting domain). Both mutants impair PR-mediated activation of a well-understood model promoter in response to progestin, as well as hormone-induced cell proliferation. Additional mutants affecting transactivation activity of PR (activation function 2) or a zinc-finger implicated in dimerization (D-box) have also been tested. Microarrays and gene expression experiments on these cell lines define the subsets of hormone-responsive genes regulated by different modes of action of PR isoform B, as well as genes in which the nuclear and nongenomic pathways cooperate. Correlation between CCND1 expression in the different cell lines and their ability to support cell proliferation confirms CCND1 as a key controller gene.
Quiles
Ignacio
I
Centre de Regulació Genòmica, Universitat Pompeu Fabra, Parc de Recerca Biomèdica de Barcelona, Spain.
Millán-Ariño
Lluís
L
Subtil-Rodríguez
Alicia
A
Miñana
Belén
B
Spinedi
Nora
N
Ballaré
Cecilia
C
Beato
Miguel
M
Jordan
Albert
A
D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism; D004252:DNA Mutational Analysis; D004789:Enzyme Activation / Q000187:drug effects; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D008957:Models, Genetic; D050505:Mutant Proteins / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D011372:Progestins / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D011494:Protein Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011980:Receptors, Progesterone / Q000737:chemistry* / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors
D050505:Mutant Proteins; D011372:Progestins; D012333:RNA, Messenger; D011980:Receptors, Progesterone; D019938:Cyclin D1; D011494:Protein Kinases