A Novel Estradiol/Estrogen Receptor α-dependent Transcriptional Mechanism Controls Expression of the Human Prolactin Receptor

Abstract

2 -activated transcriptional expression of the hPRLR through PIII. Estradiol produced in breast stroma and adipose tissue, which are major sources of estrogen in post-menopausal women, could up-regulate hPRLR gene expression and stimulate breast tumor growth. Prolactin (PRL),2 a hormone of the lactogen/cytokine family, is produced in the anterior pituitary gland and exerts diverse cellular actions through its transmembrane receptors (PRLR) in several target tissues. Prolactin is a major factor in the proliferation and differentiation of breast epithelium and is the primary hormone in the stimulation and maintenance of lactation. Prolactin is a tumor promoter in rodents (1, 2) and has been implicated in the development of breast cancer. The hPRLR has several forms, including long form (stimulatory) and short form(s) (inhibitory) (3, 4), which are expressed in normal and tumoral breast tissue and in most human breast cancer cells (5-7), and prolactin exhibits synergistic actions with estrogens on the proliferation of these cells (8). There is local production of PRL in mammary epithelial cells, and increased expression of the PRLR long form occurs in a significant number of human mammary tumors (9 -12). A lower ratio of short (inhibitory forms)/long (activating form) receptors reported in breast tumor tissues could cause unopposed prolactin-mediated stimulatory actions of the long form and may contribute to breast tumor development and progression (7). Moreover, the PRL antagonist G129R was reported to cause apoptosis in breast cancer cells (8, 13). These findings, and the correlation between serum PRL and the incidence and progression of breast tumors (11,12,14), indicate that PRL has a role in human breast cancer. Stromal and adipose tissue are the major sources of estrogen in post-menopausal women, and could exert paracrine control of prolactin and prolactin receptor expression in adjacent mammary epithelial cells.Our previous studies on the hPRLR gene revealed its complex 5Ј genomic structure, with multiple (six) alternative non-coding exons 1 and promoter utilization (15,16). These include the preferentially utilized, generic promoter 1/exon-1 (PIII/hE1 3 ), which is also present in rat and mouse, and five human specific exon-1/promoters (hE1 N1-5 ) (15-17). These forms were found to be expressed in breast cancer cells, and variably in other tissues (16). Quantitative competitive reverse transcriptase-PCR analysis showed that E 2 induced increases of PRLR non-coding exon-1 hE1 3 (generic) mRNA transcripts directed by promoter III (hPIII) in breast cancer cells. Also, in transfection studies E 2 activated the hPIII promoter (18). This promoter contains functional Sp1 and C/EBP sites that bind Sp1/Sp3 and C/EBP␤, respectively (16,19). The lack of a formal ERE in the hPIII promoter suggested that the effect of estradiol is mediated through association of the activated ER with relevant DNA binding transfactors. Thus, although there is not a classical ERE within the hPRLR promoter/5Ј-flanking region of hPIII, our ...

Progesterone induces expression of the prolactin receptor gene through cooperative action of Sp1 and C/EBP

“…A study of the rat PrlR promoter region III showed that it has a C/EBPβ and a nearly adjacent Sp1 site in its proximal region . The proximal human promoter retains these response elements and is, in fact, 81 % similar in sequence to the rat promoter (Dong et al, 2006), as is the mouse PIII promoter (Ormandy et al, 1998). The rat generic PrlR PIII promoter is constitutively activated by C/EBP and Sp1 in liver and gonadal cells (Hu et al, 1996;Moldrup et al, 1996); the human PrlR promoter in MCF7 breast cancer cells is activated by E acting through its receptor which tethers to these same transcription factors (Dong et al, 2006).…”

“…The mouse PrlR promoter III (824-PIII; GeneBank accession #AC074172; a kind gift from Dr. Robert Callahan, NCI) including the 5' flanking region/exon 1 (Dong et al, 2006) and contained within a pGL3 basic vector (Promega, Madison, WI) was PCR cloned with 5 Prime (Gaithersburg, MD) using XhoI/HindIII primers to make the −439/−179-PIII (-439-PIII) construct. Primers (Table 1A) were used at a concentration of 6 μM each with 50 ng of 824-PIII.…”

“…Additionally, the cooperative effects of these three hormones may occur in part due to mutual induction of each other's receptors. While Prl upregulates PR mRNA in the mammary gland (Ormandy et al, 1997) and activates ERα in T47D cells (Gonzalez et al, 2009), PrlR mRNA is induced by P in T47D cells (Tseng and Zhu, 1998) and by E in MCF7 cells (Dong et al, 2006).…”

“…In each species, the major cell-associated receptors contain the same extracellular and transmembrane domains (Clevenger et al, 2003), with splice variants from the remaining exons resulting in different intracellular/signaling domains. The PrlR gene in both mice and humans has several (5 for mouse, 6 for human) exon 1s (Dong et al, 2006;Ormandy et al, 1998). Each exon 1 has its own promoter, several of which have been identified.…”

“…The promoter/exon 1 combination that is utilized may depend on which transcription factors are present in a particular tissue at a particular time (Ormandy et al, 1998). The most generic of these promoters and the one that is functional in both the mouse and human mammary gland is PrlR promoter region III (Dong et al, 2006;Ormandy et al, 1998). The human promoter in this region (−439/−179) is 81% similar in sequence to the mouse promoter, as is the rat PIII promoter (Hu et al, 1996).…”