Steroid hormone regulation of the human kallikrein 10 (KLK10) gene in cancer cell lines and functional characterization of the KLK10 gene promoter.
BACKGROUND
The human kallikrein 10 (KLK10) gene is a new member of the human tissue kallikrein gene family. It encodes for a secreted serine protease (hK10) with predicted trypsin-like enzymatic activity. KLK10 is highly expressed in the sex organs and its expression level changes in malignancy.


METHODS
To determine the role of steroid hormones in KLK10 gene expression, we investigated its modulation by 17beta-estradiol, 5alpha-dihydrotestosterone, norgestrel, dexamethasone and aldosterone, at both the transcription and translation level, in a panel of cancer cell lines. After steroid hormone stimulation, the change of KLK10 mRNA was monitored with reverse transcriptase polymerase chain reaction and hK10 protein levels in the culture supernatant were quantified with an hK10-specific immunoassay. The presence of hormone response elements in the KLK10 gene promoter was examined with the chloramphenicol acetyltransferase reporter gene system.


RESULTS
The KLK10 expression was mainly up-regulated by estrogens, androgens and progestins, and to a lesser extent by dexamethasone and aldosterone in the breast cancer cell lines BT-474, MCF-7 and T-47D, both at the mRNA and protein levels. The effect of stimulation of these steroids on KLK10 expression varied among the cell lines. Estrogens, androgens and progestins were most potent in the BT-474, T-47D and MCF-7 cells, respectively. The up-regulation effect of estrogens, androgens, and progestins on KLK10 expression can be blocked by their antagonists ICI-182, 780, RU-56,187, and mifepristone, respectively. Time course studies showed that hK10 protein started to increase 1 day after steroid hormone stimulation and this increase persisted for 7 days. These data suggest that steroid hormones up-regulate KLK10 gene expression through direct interaction between hormone-receptor complexes and their cognate hormone response elements. To search for hormone response elements, we functionally characterized the KLK10 promoter by placing it upstream of the chloramphenicol acetyltransferase reporter gene. We found that KLK10 promoter activity did not rely on the presence of functional estrogen and androgen receptors. Also, the presence of functional estrogen and androgen receptors did not increase its constitutive activity. We suggest that the hormone response elements that mediate the transcriptional regulation of KLK10 are unlikely to locate in the KLK10 promoter.


CONCLUSIONS
Estrogens, androgens and progestins modulate KLK10 expression through their own receptors but this regulation is not mediated by steroid hormone response elements in the promoter of the KLK10 gene.
Department of Pathology, Mount Sinai Hospital, Toronto, Ontario, Canada M5G 1X5.
Liu-Ying
LY
Luo
Linda
L
Grass
Eleftherios P
EP
Diamandis
D000450:Aldosterone / Q000494:pharmacology; D000726:Androgen Antagonists / Q000494:pharmacology; D002460:Cell Line; D045744:Cell Line, Tumor; D015500:Chloramphenicol O-Acetyltransferase / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D003907:Dexamethasone / Q000494:pharmacology; D013196:Dihydrotestosterone / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D004958:Estradiol / Q000031:analogs & derivatives* / Q000494:pharmacology; D004965:Estrogen Antagonists / Q000494:pharmacology; D005260:Female; D000077267:Fulvestrant; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D012739:Gonadal Steroid Hormones / Q000494:pharmacology*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007610:Kallikreins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D007700:Kinetics; D015735:Mifepristone / Q000494:pharmacology; D009570:Nitriles / Q000494:pharmacology; D009644:Norgestrel / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D015854:Up-Regulation
D000726:Androgen Antagonists; D004965:Estrogen Antagonists; D012739:Gonadal Steroid Hormones; D007093:Imidazoles; D009570:Nitriles; D011993:Recombinant Fusion Proteins; D013196:Dihydrotestosterone; C090226:4-(3,4,4-trimethyl-5-oxo-2-thioxo-1-imidazolidinyl)-2-(trifluoromethyl)benzonitrile; D000077267:Fulvestrant; D015735:Mifepristone; D009644:Norgestrel; D000450:Aldosterone; D004958:Estradiol; D003907:Dexamethasone; D015500:Chloramphenicol O-Acetyltransferase; C100936:KLK10 protein, human; D007610:Kallikreins