A transcription-independent role for HIF-1α in modulating microprocessor assembly.

Microprocessor is an essential nuclear complex responsible for the initial RNase-mediated cleavage of primary miRNA, which is a tightly controlled maturation process that requires the proper assembly of Drosha and DGCR8. Unlike previously identified mechanisms directly targeting the enzymatic subunit Drosha, current knowledge about the biological ways of controlling miRNA nuclear maturation through DGCR8 is less addressed. In this study, we unveiled that the microprocessor assembly is governed by a master gene regulator HIF-1α irrespective of its canonical transcriptional activity. First, a widespread protein binding of HIF-1α with DGCR8 instead of Drosha was observed in response to biological stimulations. Similar protein interactions between their corresponding orthologues in model organisms were also observed. After dissecting the essential protein domains, we noticed that HIF-1α suppresses microprocessor assembly via binding to DGCR8. Furthermore, our results showed that HIF-1α hijacks monomeric DGCR8 thus reducing its dimer formation prior to microprocessor assembly, and consequently, the suppressed microprocessor formation and nuclear processing of primary miRNA were demonstrated. In conclusion, here we unveiled the mechanism of how microprocessor assembly is regulated by HIF-1α, which not only demonstrates a non-transcriptional function of nuclear HIF-1α but also provides new molecular insights into the regulation of microprocessor assembly through DGCR8.

TARBP2 Suppresses Ubiquitin-Proteasomal Degradation of HIF-1α in Breast Cancer.

TAR (HIV-1) RNA binding protein 2 (TARBP2) is an RNA-binding protein participating in cytoplasmic microRNA processing. Emerging evidence has shown the oncogenic role of TARBP2 in promoting cancer progression, making it an unfavorable prognosis marker for breast cancer. Hypoxia is a hallmark of the tumor microenvironment which induces hypoxia-inducible factor-1α (HIF-1α) for transcriptional regulation. HIF-1α is prone to be rapidly destabilized by the ubiquitination-proteasomal degradation system. In this study, we found that TARBP2 expression is significantly correlated with induced hypoxia signatures in human breast cancer tissues. At a cellular level, HIF-1α protein level was maintained by TARBP2 under either normoxia or hypoxia. Mechanistically, TARBP2 enhanced HIF-1α protein stability through preventing its proteasomal degradation. In addition, downregulation of multiple E3 ligases targeting HIF-1α (VHL, FBXW7, TRAF6) and reduced ubiquitination of HIF-1α were also induced by TARBP2. In support of our clinical findings that TARBP2 is correlated with tumor hypoxia, our IHC staining showed the positive correlation between HIF-1α and TARBP2 in human breast cancer tissues. Taken together, this study indicates the regulatory role of TARBP2 in the ubiquitination-proteasomal degradation of HIF-1α protein in breast cancer.

TARBP2-Enhanced Resistance during Tamoxifen Treatment in Breast Cancer.

Tamoxifen is the most widely used hormone therapy in estrogen receptor-positive (ER+) breast cancer, which accounts for approximately 70% of all breast cancers. Although patients who receive tamoxifen therapy benefit with respect to an improved overall prognosis, resistance and cancer recurrence still occur and remain important clinical challenges. A recent study identified TAR (HIV-1) RNA binding protein 2 (TARBP2) as an oncogene that promotes breast cancer metastasis. In this study, we showed that TARBP2 is overexpressed in hormone therapy-resistant cells and breast cancer tissues, where it enhances tamoxifen resistance. Tamoxifen-induced TARBP2 expression results in the desensitization of ER+ breast cancer cells. Mechanistically, tamoxifen post-transcriptionally stabilizes TARBP2 protein through the downregulation of Merlin, a TARBP2-interacting protein known to enhance its proteasomal degradation. Tamoxifen-induced TARBP2 further stabilizes SOX2 protein to enhance desensitization of breast cancer cells to tamoxifen, while similar to TARBP2, its induction in cancer cells was also observed in metastatic tumor cells. Our results indicate that the TARBP2-SOX2 pathway is upregulated by tamoxifen-mediated Merlin downregulation, which subsequently induces tamoxifen resistance in ER+ breast cancer.

HIF-1α promotes autophagic proteolysis of Dicer and enhances tumor metastasis.

HIF-1α, one of the most extensively studied oncogenes, is activated by a variety of microenvironmental factors. The resulting biological effects are thought to depend on its transcriptional activity. The RNAse enzyme Dicer is frequently downregulated in human cancers, which has been functionally linked to enhanced metastatic properties; however, current knowledge of the upstream mechanisms regulating Dicer is limited. In the present study, we identified Dicer as a HIF-1α-interacting protein in multiple types of cancer cell lines and different human tumors. HIF-1α downregulated Dicer expression by facilitating its ubiquitination by the E3 ligase Parkin, thereby enhancing autophagy-mediated degradation of Dicer, which further suppressed the maturation of known tumor suppressors, such as the microRNA let-7 and microRNA-200b. Consequently, expression of HIF-1α facilitated epithelial-mesenchymal transition (EMT) and metastasis in tumor-bearing mice. Thus, this study uncovered a connection between oncogenic HIF-1α and the tumor-suppressive Dicer. This function of HIF-1α is transcription independent and occurs through previously unrecognized protein interaction-mediated ubiquitination and autophagic proteolysis.

A novel application of E1A in combination therapy with EGFR-TKI treatment in breast cancer.

Epidermal growth factor receptor (EGFR) is commonly overexpressed in breast cancer and is associated with poor clinical outcomes; however, an increasing number of patients have shown a poor effective response to EGFR tyrosine kinase inhibitors (EGFR-TKI). Here, we found that AXL expression was positively correlated with poor progression in breast cancer patients. Suppression of AXL by an anti-tumor protein, E1A, enhanced EGFR-TKI (gefitinib, erlotinib and lapatinib) sensitization, resulting in significant inhibition of tumor growth in breast cancer cells. Additionally, AXL overexpression dramatically impaired E1A-mediated EGFR-TKI sensitization. These findings show that downregulation of AXL expression by E1A contributes to sensitization to EGFR-TKI in breast cancer, suggesting that combinatorial therapy of AXL inhibitors or E1A gene therapy with EGFR-TKI may be a potential therapeutic strategy for treatment of breast cancer patients.