BHLHE40 modulates gene transcription in breast cancer cells exposed to hypoxia and low glucose

Knockdown of BHLHE40 expression significantly reduced primary tumor growth and spontaneous lung metastasis in an orthotopic xenograft model of human breast cancer. Gene expression analysis implicated a role of BHLHE40 in hypoxia-induced exosomic secretion of Heparin Binding Epidermal Growth Factor HBEGF, which promotes cell survival and invasion. BHLHE40 induces HBEGF transcription by blocking DNA binding of HDAC1 and HDAC2. A lung metastatic subline of MDA-MB-231, designated as LM, was stably transfected with MISSION shRNA (TRCN0000232187) to knockdown BHLHE40 or control vector. The cells were cultured in medium containing low glucose (1 mg/ml) under hypoxia (1% O2) for 6 or 48 hr. Total RNA was purified by using the RNeasy kits (Qiagen). A total of 12 samples were subjected to microarray analysis, with two biological repeats for each experiment condition.