The development and progression of malignancies is a complex multistage process that involves the contribution of a number of genes giving growth advantage to cells when transformed. The role of transforming growth factor-B (TGF-B) in carcinogenesis is complex with tumor-suppressor or prooncogenic activities depending on the cell type and the stage of the disease. We have previously reported the identification of a novel WD-domain protein, STRAP, that associates with both TGF-B receptors and that synergizes with the inhibitory Smad, Smad7, in the negative regulation of TGF-B-induced transcription. Here, we show that STRAP is ubiquitously expressed and is localized in both cytoplasm and nucleus. STRAP is up-regulated in 60% colon and in 78% lung carcinomas. Stable expression of STRAP results in activation of mitogen-activated protein kinase/extracellular signal-regulated kinase pathway and in down-regulation of the cyclin-dependent kinase inhibitor p21Cip1 , which results in retinoblastoma protein hyperphosphorylation. In addition, we have observed that Smad2/3 phosphorylation, TGF-Bmediated transcription, and growth inhibition are induced in STRAP-knockout mouse embryonic fibroblasts compared with wild-type cells. Ectopic expression of STRAP in A549 lung adenocarcinoma cell line inhibits TGF-B-induced growth inhibition and enhances anchorage-independent growth of these cells. Moreover, overexpression of STRAP increases tumorigenicity in athymic nude mice. Knockdown of endogenous STRAP by small interfering RNA increases TGF-B signaling, reduces ERK activity, increases p21 Cip1 expression, and decreases tumorigenicity. Taken together, these results suggest that up-regulation of STRAP in human cancers may provide growth advantage to tumor cells via TGF-B-dependent and TGF-B-independent mechanisms, thus demonstrating the oncogenic function of STRAP. (Cancer Res 2006; 66(12): 6156-66)

Androgen receptor (AR) action throughout prostate development and in maintenance of the prostatic epithelium is partly controlled by interactions between AR and forkhead box (FOX) transcription factors, particularly FOXA1. We sought to identity additional FOXA1 binding partners that may mediate prostate-specific gene expression. Here we identify the nuclear factor I (NFI) family of transcription factors as novel FOXA1 binding proteins. All four family members (NFIA, NFIB, NFIC, and NFIX) can interact with FOXA1, and knockdown studies in androgen-dependent LNCaP cells determined that modulating expression of NFI family members results in changes in AR target gene expression. This effect is probably mediated by binding of NFI family members to AR target gene promoters, because chromatin immunoprecipitation (ChIP) studies found that NFIB bound to the prostate-specific antigen enhancer. Förster resonance energy transfer studies revealed that FOXA1 is capable of bringing AR and NFIX into proximity, indicating that FOXA1 facilitates the AR and NFI interaction by bridging the complex. To determine the extent to which NFI family members regulate AR/FOXA1 target genes, motif analysis of publicly available data for ChIP followed by sequencing was undertaken. This analysis revealed that 34.4% of peaks bound by AR and FOXA1 contain NFI binding sites. Validation of 8 of these peaks by ChIP revealed that NFI family members can bind 6 of these predicted genomic elements, and 4 of the 8 associated genes undergo gene expression changes as a result of individual NFI knockdown. These observations suggest that NFI regulation of FOXA1/AR action is a frequent event, with individual family members playing distinct roles in AR target gene expression.

Members of the transforming growth factor-b (TGF-b) family regulate a wide range of biological processes including cell proliferation, migration, differentiation, apoptosis, and extracellular matrix deposition. Resistance to TGF-b-mediated tumour suppressor function in human lung cancer may occur through the loss of type II receptor (TbRII) expression. In this study, we investigated the expression pattern of TbRII in human lung cancer tissues by RT -PCR and Western blot analyses. We observed downregulation of TbRII in 30 out of 46 NSCLC samples (65%) by semiquantitative RT -PCR. Western blot analyses with tumour lysates showed reduced expression of TbRII in 77% cases. We also determined the effect of TbRII expression in lung adenocarcinoma cell line (VMRC-LCD) that is not responsive to TGF-b due to lack of TbRII expression. Stable expression of TbRII in these cells restored TGF-b-mediated effects including Smad2/3 and Smad4 complex formation, TGF-b-responsive reporter gene activation, inhibition of cell proliferation and increased apoptosis. Clones expressing TbRII showed reduced colony formation in soft-agarose assay and significantly reduced tumorigenicity in athymic nude mice. Therefore, these results suggest that reestablishment of TGF-b signalling in TbRII null cells by stable expression of TbRII can reverse malignant behaviour of cells and loss of TbRII expression may be involved in lung tumour progression.

Mesenchymal stem cell recruitment to the bladder after bladder outlet obstruction appears to be associated with increased blood flow and decreased tissue hypoxia, which may contribute to improvement in histopathological and functional parameters. Mesenchymal stem cell recruitment may be related to CCL2 over expression. Additional studies in larger samples are needed but these initial results suggest a potential role for mesenchymal stem cell based therapy for bladder outlet obstruction related bladder injury.

Purpose We have previously reported that embryonic rat bladder mesenchyma has the appropriate inductive signals to direct pluripotent mouse embryonic stem cells toward endodermal derived urothelium and develop mature bladder tissue. We determined whether nonembryonic stem cells, specifically bone marrow derived mesenchymal stem cells, could serve as a source of pluripotent or multipotent progenitor cells. Materials and Methods Epithelium was separated from the mesenchymal shells of embryonic day 14 rat bladders. Mesenchymal stem cells were isolated from mouse femoral and tibial bone marrow. Heterospecific recombinant xenografts were created by combining the embryonic rat bladder mesenchyma shells with mesenchymal stem cells and grafting them into the renal subcapsular space of athymic nude mice. Grafts were harvested at time points of up to 42 days and stained for urothelial and stromal differentiation. Results Histological examination of xenografts comprising mouse mesenchymal stem cells and rat embryonic rat bladder mesenchyma yielded mature bladder structures showing normal microscopic architecture as well as proteins confirming functional characteristics. Specifically the induced urothelium expressed uroplakin, a highly selective marker of urothelial differentiation. These differentiated bladder structures demonstrated appropriate α-smooth muscle actin staining. Finally, Hoechst staining of the xenografts revealed nuclear architecture consistent with a mouse mesenchymal stem cell origin of the urothelium, supporting differentiated development of these cells. Conclusions In the appropriate signaling environment bone marrow derived mesenchymal stem cells can undergo directed differentiation toward endodermal derived urothelium and develop into mature bladder tissue in a tissue recombination model. This model serves as an important tool for the study of bladder development with long-term application toward cell replacement therapies in the future.