Phosphorylation of human progesterone receptor by cyclin-dependent kinase 2 on three sites that are authentic basal phosphorylation sites in vivo.

The human progesterone receptor (hPR) in T47D breast cancer cells is phosphorylated on at least nine different serine residues. We have previously reported the identification of five sites; three are hormone inducible (Ser102, Ser294 and Ser345), and their phosphorylation correlates with the timing of the change in receptor mobility on gel electrophoresis in response to hormone treatment. The other two sites, Ser81 and Ser162, along with the remaining sites, are basally phosphorylated and exhibit a general increase in phosphorylation in response to hormone. With the exception of Ser81, all of these sites are in Ser-Pro motifs, suggesting that proline-directed kinases are responsible for their phosphorylation. We now report that cyclin A-cyclin-dependent kinase-2 complexes phosphorylate hPR-B in vitro with a high stoichiometry on three sites that are authentic basal sites in vivo. One of these is Ser162, which has been described previously. The other two sites are identified here as Ser190 and Ser400. The specificity and stoichiometry of the in vitro phosphorylation suggest that hPR phosphorylation may be regulated in a cell cycle-dependent manner in vivo.

[1]  D. Edwards,et al.  Stoichiometry and Site-specific Phosphorylation of Human Progesterone Receptor in Native Target Cells and in the Baculovirus Expression System* , 1996, The Journal of Biological Chemistry.

[2]  K. Horwitz,et al.  Role of Phosphorylation on DNA Binding and Transcriptional Functions of Human Progesterone Receptors* , 1996, The Journal of Biological Chemistry.

[3]  N. Weigel,et al.  Phosphorylation of Ser211 in the Chicken Progesterone Receptor Modulates its Transcriptional Activity* , 1996, The Journal of Biological Chemistry.

[4]  D. Picard,et al.  Activation of the unliganded estrogen receptor by EGF involves the MAP kinase pathway and direct phosphorylation. , 1996, The EMBO journal.

[5]  M. Haussler,et al.  Human vitamin D receptor phosphorylation by casein kinase II at Ser-208 potentiates transcriptional activation. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[6]  D. Edwards,et al.  Two Types of Anti-progestins Have Distinct Effects on Site-specific Phosphorylation of Human Progesterone Receptor (*) , 1996, The Journal of Biological Chemistry.

[7]  D. Pappin,et al.  Identification of phosphorylation sites in the mouse oestrogen receptor , 1995, The Journal of Steroid Biochemistry and Molecular Biology.

[8]  Daniel Metzger,et al.  Activation of the Estrogen Receptor Through Phosphorylation by Mitogen-Activated Protein Kinase , 1995, Science.

[9]  D. Lannigan,et al.  Estradiol and phorbol ester cause phosphorylation of serine 118 in the human estrogen receptor. , 1995, Molecular endocrinology.

[10]  D. Edwards,et al.  Identification of a group of Ser-Pro motif hormone-inducible phosphorylation sites in the human progesterone receptor. , 1995, Molecular endocrinology.

[11]  M. Sliwkowski,et al.  HER-2 tyrosine kinase pathway targets estrogen receptor and promotes hormone-independent growth in human breast cancer cells. , 1995, Oncogene.

[12]  E. Wilson,et al.  Identification of three proline-directed phosphorylation sites in the human androgen receptor. , 1995, Molecular endocrinology.

[13]  S. Elledge,et al.  Inhibition of cyclin-dependent kinases by p21. , 1995, Molecular biology of the cell.

[14]  David O. Morgan,et al.  Principles of CDK regulation , 1995, Nature.

[15]  D. DeFranco,et al.  Selectivity of Cell Cycle Regulation of Glucocorticoid Receptor Function (*) , 1995, The Journal of Biological Chemistry.

[16]  S. F. Arnold,et al.  In vivo and in vitro phosphorylation of the human estrogen receptor , 1995, The Journal of Steroid Biochemistry and Molecular Biology.

[17]  D. Edwards,et al.  Identification of Phosphorylation Sites Unique to the B Form of Human Progesterone Receptor , 1994 .

[18]  A. Munck,et al.  Cell cycle-dependent glucocorticoid receptor phosphorylation and activity. , 1994, Molecular endocrinology.

[19]  N. Weigel,et al.  Phosphorylation of Ser530 facilitates hormone-dependent transcriptional activation of the chicken progesterone receptor. , 1994, Molecular endocrinology.

[20]  D. W. Waring,et al.  Activation of the progesterone receptor by the gonadotropin-releasing hormone self-priming signaling pathway. , 1994, Molecular endocrinology.

[21]  A. Maggi,et al.  Insulin-like growth factors activate estrogen receptor to control the growth and differentiation of the human neuroblastoma cell line SK-ER3. , 1994, Molecular endocrinology.

[22]  D. Edwards,et al.  The DNA-bending protein HMG-1 enhances progesterone receptor binding to its target DNA sequences , 1994, Molecular and cellular biology.

[23]  N. Weigel,et al.  Multiple signaling pathways activate the chicken progesterone receptor. , 1994, Molecular endocrinology.

[24]  B. Katzenellenbogen,et al.  Phosphorylation of the human estrogen receptor. Identification of hormone-regulated sites and examination of their influence on transcriptional activity. , 1994, The Journal of biological chemistry.

[25]  S. Elledge,et al.  The p21 Cdk-interacting protein Cip1 is a potent inhibitor of G1 cyclin-dependent kinases , 1993, Cell.

[26]  K. Korach,et al.  Peptide growth factors elicit estrogen receptor-dependent transcriptional activation of an estrogen-responsive element. , 1993, Molecular endocrinology.

[27]  B. Katzenellenbogen,et al.  Stimulation of estrogen receptor-mediated transcription and alteration in the phosphorylation state of the rat uterine estrogen receptor by estrogen, cyclic adenosine monophosphate, and insulin-like growth factor-I. , 1993, Molecular endocrinology.

[28]  D. Edwards,et al.  The progesterone antagonist RU486 acquires agonist activity upon stimulation of cAMP signaling pathways. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[29]  K. Horwitz,et al.  Antagonist-occupied human progesterone receptors bound to DNA are functionally switched to transcriptional agonists by cAMP. , 1993, The Journal of biological chemistry.

[30]  P. Chambon,et al.  Modulation of transcriptional activation by ligand‐dependent phosphorylation of the human oestrogen receptor A/B region. , 1993, The EMBO journal.

[31]  N. Weigel,et al.  Identification of a hormone-dependent phosphorylation site adjacent to the DNA-binding domain of the chicken progesterone receptor. , 1993, Molecular endocrinology.

[32]  P. Chambon,et al.  Cooperation of proto‐signals for nuclear accumulation of estrogen and progesterone receptors. , 1992, The EMBO journal.

[33]  N. Koszewski,et al.  Estrogen receptor phosphorylation. Hormonal dependence and consequence on specific DNA binding. , 1992, The Journal of biological chemistry.

[34]  D. Edwards,et al.  Effects of hormone and cellular modulators of protein phosphorylation on transcriptional activity, DNA binding, and phosphorylation of human progesterone receptors. , 1992, Molecular endocrinology.

[35]  T. Curran,et al.  Pro-Leu-Ser/Thr-Pro is a consensus primary sequence for substrate protein phosphorylation. Characterization of the phosphorylation of c-myc and c-jun proteins by an epidermal growth factor receptor threonine 669 protein kinase. , 1991, The Journal of biological chemistry.

[36]  T. Wong,et al.  A manual sequencing method for identification of phosphorylated amino acids in phosphopeptides. , 1991, Analytical biochemistry.

[37]  S. Pelech,et al.  Definition of a consensus sequence for peptide substrate recognition by p44mpk, the meiosis-activated myelin basic protein kinase. , 1991, The Journal of biological chemistry.

[38]  Rf Power,et al.  Dopamine activation of an orphan of the steroid receptor superfamily , 1991, Science.

[39]  D. Pappin,et al.  Identification of phosphorylated sites in the mouse glucocorticoid receptor. , 1991, The Journal of biological chemistry.

[40]  B. O’Malley,et al.  Regulation of progesterone receptor-mediated transcription by phosphorylation. , 1990, Science.

[41]  B. O’Malley,et al.  Hormonal regulation and identification of chicken progesterone receptor phosphorylation sites. , 1990, The Journal of biological chemistry.

[42]  P Chambon,et al.  Two distinct estrogen‐regulated promoters generate transcripts encoding the two functionally different human progesterone receptor forms A and B. , 1990, The EMBO journal.

[43]  A. Munck,et al.  Agonist-dependent phosphorylation and nuclear dephosphorylation of glucocorticoid receptors in intact cells. , 1989, The Journal of biological chemistry.

[44]  K. Horwitz,et al.  Synthesis of human progesterone receptors in T47D cells. Nascent A- and B-receptors are active without a phosphorylation-dependent post-translational maturation step. , 1989, The Journal of biological chemistry.

[45]  K. Horwitz,et al.  Phosphotryptic peptide analysis of human progesterone receptor. New phosphorylated sites formed in nuclei after hormone treatment. , 1989, The Journal of biological chemistry.

[46]  G. Fasman,et al.  Phosphorylation loops in synthetic peptides of the human neurofilament protein middle-sized subunit , 1988, Journal of protein chemistry.

[47]  R. Evans,et al.  The steroid and thyroid hormone receptor superfamily. , 1988, Science.

[48]  D. Edwards,et al.  Immunologic analysis of human breast cancer progesterone receptors. 1. Immunoaffinity purification of transformed receptors and production of monoclonal antibodies. , 1987, Biochemistry.

[49]  G. Allen,et al.  Sequencing of proteins and peptides , 1981 .

[50]  G. Drapeau,et al.  Substrate specificity of a proteolytic enzyme isolated from a mutant of Pseudomonas fragi. , 1980, The Journal of biological chemistry.