Aberrant Polycystin-1 Expression Results in Modification of Activator Protein-1 Activity, whereas Wnt Signaling Remains Unaffected

Abstract

Polycystin-1, the polycystic kidney disease 1 gene product, has been implicated in several signaling complexes that are known to regulate essential cellular functions. We investigated the role of polycystin-1 in Wnt signaling and activator protein-1 (AP-1) activation. To this aim, a membrane-targeted construct encoding the conserved C-terminal region of mouse polycystin-1 reported to mediate signal transduction activity was expressed in human embryonic and renal epithelial cells. To ensure specificity and minimal cotransfection effects, we focused our study on the endogenous proteins that actually transduce the signals, ␤-catenin and T-cell factor/lymphoid-enhancing factor for Wnt signaling and (phosphorylated) c-Jun, ATF2, and c-Fos for AP-1. Our data indicate that the C-terminal region of polycystin-1 activates AP-1 by inducing phosphorylation and expression of at least c-Jun and ATF2, whereas c-Fos was not affected. Under our experimental conditions, polycystin-1 did not modulate Wnt signaling. AP-1 activity was aberrant in human autosomal dominant polycystic kidney disease (ADPKD) renal cystic epithelial cells and in renal epithelial cells expressing transgenic full-length polycystin-1, resulting in decreased Jun-ATF and increased Jun-Fos activity, whereas Wnt signaling remained unaffected. Since our data indicate that aberrant polycystin-1 expression results in altered AP-1 activity, polycystin-1 may be required for adequate AP-1 activity.Progressive development of fluid-filled cysts in autosomal dominant polycystic kidney disease (ADPKD) 1 results in chronic renal failure. In the majority of patients, the disease can be accounted for by a mutation in the PKD1 gene (1, 2), whereas a minority suffers from a mutation in the PKD2 gene (3, 4). The precise function of polycystin-1 and polycystin-2, the proteins encoded by the PKD1 and PKD2 gene, respectively, remains to be elucidated. Polycystin-1 is predicted to be a transmembrane protein of ϳ460 kDa. The large extracellular N terminus contains multiple domains thought to be involved in cell-cell and cell-matrix interactions. The intracellular C terminus of polycystin-1 contains putative phosphorylation sites and a coiled-coil domain that can mediate protein-protein interactions.Several studies have implicated a role for polycystin-1 in signal transduction. Overexpression of the C-terminal region of polycystin-1 in human embryonic kidney 293T (HEK293T) cells has been shown to activate the Wnt signaling pathway (5) and the activator protein-1 (AP-1) transcription factor complex (6, 7). Furthermore, overexpression of a full-length polycystin-1 construct has been reported to activate the Janus kinase and signal transducer and activator of transcription (JAK-STAT) signaling pathway (8). These signaling pathways are all involved in key cellular processes such as proliferation and differentiation, cell cycle regulation, and cell survival. Since these cellular processes are essential for normal function, the signaling pathways governing them are tightly regulated. We ...

Increased Activity of Activator Protein-1 Transcription Factor Components ATF2, c-Jun, and c-Fos in Human and Mouse Autosomal Dominant Polycystic Kidney Disease

“…These data are in line with previous reports that increased polycystin-1 expression results in polycystic kidney disease (8,34). Renal epithelial cells expressing transgenic PKD1 show similar defects in AP-1 transcriptional activity as human ADPKD renal cystic epithelial cells (8), and transgenic Pkd1 mice develop polycystic kidney disease (34).…”

“…However, to further study this hypothesis, cyst formation should be studied at earlier stages under more controlled laboratory conditions, for instance using conditional knockout mouse models for Pkd1 and Pkd2. The immortalized renal cystic epithelial cells that were analyzed in our previous report may represent a subset of endstage cystic cells from large cysts displaying decreased level of Ser73-phosphorylated c-Jun and increased c-Fos in addition to an increase in Thr71-phosphorylated ATF2 (8). Furthermore, cultured renal cystic cells may differ from renal cystic epithelium because these cells are surrounded by an abundant expanded fibrotic interstitium in vivo.…”

“…Western blot analysis was performed as described (8). Rabbit anti-phospho-Thr71-ATF2, rabbit anti-phospho-Thr69/71-ATF2, and rabbit anti-phospho-Ser73-c-Jun were diluted 1:500, rabbit anti-c-Fos was diluted 1:1000 (Upstate, Charlottesville, VA), and mouse anti-␤-actin was diluted 1:40,000 (MP Biomedicals, Amsterdam, The Netherlands).…”

“…Several studies have implicated a role for the PKD1 gene product, polycystin-1, in signal transduction pathways governing activator protein-1 (AP-1) activity (6 -8). Moreover, we have reported that AP-1 activity is aberrant in an immortalized renal cystic epithelial cell line derived from a patient with ADPKD (8). AP-1 transcription factors are composed of multiple homoor heterodimers that regulate key cellular processes such as cell proliferation, differentiation, and survival (9 -11).…”

The isolated polycystin-1 COOH-terminal can activate or block polycystin-1 signaling

“…For example: full-length Pc-1 inhibits cell proliferation via activation of JAK-STAT [2], whereas the truncation has been found to enhance proliferation, probably through modulation of cell calcium [3][4][5]. The capacity of the pc-1 truncation to activate wnt signaling appears to be inconsistent [6,7] and altered wnt signaling has not been observed in human cystic disease [7]. Although the truncation activates AP-1 [7,8], full-length Pc-1 might have the opposite effect, since disruption of the PKD-1 gene in mouse and human ADPKD seems to cause increased AP-1 activity [9].…”