Restoration of Transforming Growth Factor-β Signaling through Receptor RI Induction by Histone Deacetylase Activity Inhibition in Breast Cancer Cells

Abstract

The loss of transforming growth factor-␤ (TGF-␤) response due to the dysregulation of TGF-␤ receptors type I (RI) and type II (RII) is well known for its contribution to oncogenesis. Estrogen receptor-expressing breast cancer cells are refractory to TGF-␤-mediated growth control because of the reduced expression of TGF-␤ receptors. Although RII is required for the binding of TGF-␤ to RI, RI is responsible for directly transducing TGF-␤ signals through the Smad protein family. Treatment of estrogen receptor-expressing MCF-7L and ZR75 breast cancer cells with the histone deacetylase (HDAC) inhibitor suberoylanilide hydroxamic acid (SAHA) led to a dramatic induction of RI. Accumulation of acetylated histones H3 and H4 was observed in the SAHAtreated cells. Chromatin immunoprecipitation analysis followed by PCR with RI promoter-specific primers indicated an accumulation of acetylated histones in chromatin associated with the RI gene, suggesting that histone deacetylation was involved in the transcriptional inactivation of RI. SAHA treatment stimulated RI promoter activity through the inhibition of Sp1/Sp3-associated HDAC activity. Histone acetyltransferase p300 stimulated RI promoter activity, thus further confirming the involvement of HDAC activity in the transcriptional repression of RI. Significantly, SAHA-mediated RI regeneration restored the TGF-␤ response in breast cancer cells.Transforming growth factor-␤ (TGF-␤), 1 a 25-kDa homodimeric polypeptide, plays an important role in the growth inhibition of most normal epithelial and some cancer cells (1). TGF-␤ functions through cell surface receptors referred to as type I (RI) and type II (RII). RI requires RII for the binding of TGF-␤. However, RI is a direct player in the TGF-␤ signaling pathway as it conveys signals from TGF-␤ through the activation of Smad protein family members (2). The loss of TGF-␤ response is well known for its contribution to oncogenesis and tumor progression. Direct involvement of RI in TGF-␤ signal transduction would suggest that loss or reduced expression of RI could contribute to TGF-␤ resistance resulting in a growth advantage that contributes to tumor progression.TGF-␤ resistance due to methylation of the RI promoter or RI promoter repression by Sp1 deficiency was reported to be a cause of TGF-␤ resistance in gastric and colon carcinomas (3,4). The RI gene is frequently mutated in ovarian carcinomas (5). Decreased expression of RI is associated with poor prognosis in bladder transitional cell carcinomas (6). Reduced RI expression is also associated with unfavorable prognosis in esophageal squamous cell carcinoma (7). RI*6A, a polymorphic allele of RI, is emerging as a high frequency, low penetrance tumor susceptibility allele that predisposes to the development of breast, ovarian, and colorectal cancer, as well as hematologic malignancies (8). Polymorphisms in the microsatellite region of the RI gene were reported in head and neck cancers as well as non-small cell lung cancer (9, 10). Mutations in the kinase domain of the RI gene we...

Laser microdissection and microarray analysis of breast tumors reveal ER-α related genes and pathways

“…Transforming growth factor b (TGF-b) has been demonstrated to have both tumor suppression and stimulating effects during early and late stages of tumorigenesis (Akhurst and Derynck, 2001). The crosstalk between TGF-b signaling and estrogen signaling at DNA-dependent or -independent manners has been documented (Matsuda et al, 2001;Wu L et al, 2003;Ammanamanchi and Brattain, 2004). A few genes that have implied action on TGF-b signaling were identified in the common and LCM unique gene lists.…”

Regulation of RON Tyrosine Kinase-mediated Invasion of Breast Cancer Cells

“…Western Blot Analysis-Equal amounts of cell lysates from MCF-7, MDA MB 231, and MDA MB 468 breast cancer cells were resolved by 7.5% SDS-PAGE and Western analysis was performed as described previously (20). Rabbit anti-human RON, Sp1, and actin polyclonal antibodies were purchased from Santa Cruz Biotechnology.…”

“…Chromatin Immunoprecipitation Assay-The chromatin immunoprecipitation assay was performed as described previously (20). RON promoter primers were used to carry out PCR on DNA isolated from chromatin immunoprecipitation using control IgG or Sp1 antibodies.…”

Histone deacetylases and cancer

“…Likewise, overexpression of the HAT p300 increases TGFbRI promoter activity. HDAC inhibitor treatment restores TGF-b sensitivity by impairing HDAC1 activity associated with Sp1/Sp3 at the TGF-bRI promoter (Ammanamanchi and Brattain, 2004). By repressing the expression of a receptor for a growth-restraining signaling molecule, HDACs render a cancer cell unresponsive to this signal and allow for unfettered cell growth.…”