Id3 induces an Elk‐1–caspase‐8‐dependent apoptotic pathway in squamous carcinoma cells

• Published in: Cancer medicine (Malden, MA) Vol. 4; no. 6; pp. 914 - 924
• Main Authors: Chen, You‐Shin, Aubee, Joseph, DiVito, Kyle A., Zhou, Hengbo, Zhang, Weiyi, Chou, Fen‐Pi, Simbulan‐Rosenthal, Cynthia M., Rosenthal, Dean S.
• Format: Journal Article
• Language: English
• Published: United States John Wiley & Sons, Inc 01.06.2015; BlackWell Publishing Ltd
• Subjects: Amino acids; Animal models; Animals; Antineoplastic Agents - pharmacology; Apoptosis; Apoptosis - physiology; Breast cancer; Cancer Biology; Carcinoma, Squamous Cell - drug therapy; Carcinoma, Squamous Cell - pathology; Carcinoma, Squamous Cell - physiopathology; Caspase; Caspase 8 - metabolism; Caspase Inhibitors - pharmacology; caspase‐8; Cell culture; Cell cycle; Cell Death - drug effects; Cell differentiation; Cell Line, Tumor; Cell number; Cisplatin; Cisplatin - pharmacology; Clonal deletion; Cloning; Deletion mutant; Deoxyribonucleic acid; DNA; DNA microarrays; Drug Synergism; Elk; Elk‐1; ETS protein; ets-Domain Protein Elk-1 - metabolism; Fluorouracil - pharmacology; Gene expression; Heterografts; HLH; Homeostasis; Humans; Id protein; Id1 protein; Id3; Inhibitor of Differentiation Proteins - pharmacology; Inhibitor of Differentiation Proteins - physiology; Keratinocytes; Lymphoma; Mice, Nude; Mutation; Neoplasm Proteins - pharmacology; Neoplasm Proteins - physiology; Neoplasm Transplantation; Penicillin; Phosphorylation; Proteins; SCC; Signal Transduction; siRNA; Skin cancer; Software; Squamous cell carcinoma; Transcription factors; Tumor Burden; Tumor suppressor genes; Ultraviolet radiation; Xenografts; Id3; SCC; caspase-8; HLH; Elk-1; Apoptosis
• ISSN: 2045-7634; 2045-7634
• DOI: 10.1002/cam4.427

Inhibitor of differentiation/DNA‐binding (Id) proteins are helix–loop–helix (HLH) transcription factors. The Id protein family (Id1–Id4) mediates tissue homeostasis by regulating cellular processes including differentiation, proliferation, and apoptosis. Ids typically function as dominant negative HLH proteins, which bind other HLH proteins and sequester them away from DNA promoter regions. Previously, we have found that Id3 induced apoptosis in immortalized human keratinocytes upon UVB exposure, consistent with its role as a tumor suppressor. To investigate the role of Id3 in malignant squamous cell carcinoma (SCC) cells (A431), a tetracycline‐regulated inducible system was used to induce Id3 in cell culture and mouse xenograft models. We found that upon Id3 induction, there was a decrease in cell number under low serum conditions, as well as in soft agar. Microarray, RT‐PCR, immunoblot, siRNA, and inhibitor studies revealed that Id3 induced expression of Elk‐1, an E‐twenty‐six (ETS)‐domain transcription factor, inducing procaspase‐8 expression and activation. Id3 deletion mutants revealed that 80 C‐terminal amino acids, including the HLH, are important for Id3‐induced apoptosis. In a mouse xenograft model, Id3 induction decreased tumor size by 30%. Using immunofluorescent analysis, we determined that the tumor size decrease was also mediated through apoptosis. Furthermore, we show that Id3 synergizes with 5‐FU and cisplatin therapies for nonmelanoma skin cancer cells. Our studies have shown a molecular mechanism by which Id3 induces apoptosis in SCC, and this information can potentially be used to develop new treatments for SCC patients. Id3 is shown to regress squamous cell carcinoma xenografts through induction of apoptosis. This molecular pathway is mediated through Elk‐1, caspase‐8, and caspase‐3. This pathway is dependent on the HLH and C‐terminal domains of Id3.