A novel role of gap junction connexin46 protein to protect breast tumors from hypoxia

Abstract

Connexin proteins are the principle structural components of the gap junctions. Colocalization and tissue-specific expression of diverse connexin molecules are reported to occur in a variety of organs. Impairment of gap junctional intercellular communication, caused by mutations, gain of function, or loss of function of connexins, is involved in a number of diseases including the development of cancer. Here we show that human breast cancer cells, MCF-7, and breast tumor tissues express a novel gap junction protein, connexin 46 (Cx46) and it plays a critical role in hypoxia. Previous studies have shown that connexin46 is predominantly expressed in lens and our studies find that Cx46 protects human lens epithelial cells (HLEC) from hypoxia induced death. Interestingly, we find that Cx46 is upregulated in MCF-7 breast cancer cells and human breast cancer tumors. Downregulation of Cx46 by siRNA promotes 40% MCF-7 cell death at 24 hour under hypoxic conditions. Furthermore, direct injection of anti-Cx46 siRNA into xenograft tumors prevents tumor growth in nude mice. This finding will provide an exciting new direction for drug development for breast cancer treatment and suggests that both normal hypoxic tissue (lens) and adaptive hypoxic tissue (breast tumor) utilize the same protein, Cx46, as a protective strategy from hypoxia.

Investigation of the Reciprocal Relationship between the Expression of Two Gap Junction Connexin Proteins, Connexin46 and Connexin43

“…Cx43 has anti-tumor properties, and expression of Cx43 can be down-regulated in tumors (22)(23)(24)(25). Moreover, in previous studies, we have shown that Cx46 is up-regulated in breast tumor samples, MCF-7 xenografts, and Y79 retinoblastoma xenografts and acts as an oncogene to favor early tumor growth (24,25).…”

“…Immortalized HLECs and rabbit lens epithelial NN1003A cells (NN) were cultured as described previously (24). Briefly, cells were cultured in low glucose DMEM (Invitrogen) supplemented with 10% premium fetal bovine serum (Atlanta Biologicals, Atlanta, GA) to at least 70% confluence prior to use for experimentation.…”

“…Western Blot and Antisera-Western blot analyses were performed as previously described (24,25,28). Mouse anti-N-terminal-Cx43 (anti-Cx43 NT) was purchased from Fred Hutchinson Cancer Center (Seattle, WA); rabbit anti-phosphoCx43 (Ser-368) was from Cell Signaling Technologies (Danvers, MA); rabbit anti-C-terminal-Cx43 (anti-Cx43 CT, 1:5000) and mouse anti-␤-actin (1:10000) were from Sigma-Aldrich; rabbit anti-Cx46 (1:2000) was from U.S. Biologicals (Swampscott, MA); mouse anti-Cx50 (1:4000) was from Zymed Laboratories Inc.-Invitrogen (San Franscisco, CA); and mouse anti-EGFP antibody (1:5000) was purchased from Clontech.…”

Carbon Monoxide (CO) Is a Novel Inhibitor of Connexin Hemichannels

“…Cx46 has gained considerable attention lately as a potential target for cancer treatment (20,21) because it has been suggested that Cx46 helps cancer cells survive in hypoxic conditions (20). Because of its potential role of Cx46 in cancer biology and its sensitivity to gaseous transmitters such as NO (15), we decided to evaluate the possibility that Cx46 HCs are affected by HO-1 effectors (CO or guanylyl cyclase signaling pathways).…”

Cytoplasmic Connexin32 and Self-Renewal of Cancer Stem Cells: Implication in Metastasis

“…Today, connexin proteins are, however, beyond gap junction (Goodenough & Paul, 2003)! A great variety of structures, functions, and behaviours of connexin proteins are known, i.e., hemichannel (Jiang & Gu, 2005), mitochondrial connexin (Boengler et al, 2005), oncogenic connexin (Banerjee et al, 2010;Boengler et al, 2005;Ito et al, 2000), cytoplasmic connexin , fragmented connexin, and then, gap junction. Therefore, the functions of connexin proteins in cancers are not always suppressive to either cell proliferation or tumour progression (Naus & Laird, 2010).…”