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Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation.

ABSTRACT: Casein kinase 2 (CK2) is a highly ubiquitous and conserved serine/threonine kinase that forms a tetramer consisting of a catalytic subunit (CK2?) and a regulatory subunit (CK2?). Despite being ubiquitous, CK2 is commonly found at higher expression levels in cancer cells, where it inhibits apoptosis, and supports cell migration and proliferation. The Ca2+-activated chloride channel TMEM16A shows similar effects in cancer cells: TMEM16A increases cell proliferation and migration and is highly expressed in squamous cell carcinoma of the head and neck (HNSCC) as well as other malignant tumors. A microscopy-based high-throughput screening was performed to identify proteins that regulate TMEM16A. Within this screen, CK2 was found to be required for proper membrane expression of TMEM16A. small interfering (si) RNA-knockdown of CK2 reduced plasma membrane expression of TMEM16A and inhibited TMEM16A whole cell currents in (cystic fibrosis bronchial epithelial) CFBE airway epithelial cells and in the head and neck cancer cell lines Cal33 and BHY. Inhibitors of CK2, such as TBB and the preclinical compound CX4549 (silmitasertib), also blocked membrane expression of TMEM16A and Ca2+-activated whole cell currents. siRNA-knockout of CK2 and its pharmacological inhibition, as well as knockdown or inhibition of TMEM16A by either niclosamide or Ani9, attenuated cell proliferation. Simultaneous inhibition of CK2 and TMEM16A strongly potentiated inhibition of cell proliferation. Although membrane expression of TMEM16A is reduced by inhibition of CK2, our data suggest that the antiproliferative effects by inhibition of CK2 are mostly independent of TMEM16A. Simultaneous inhibition of TMEM16A by niclosamide and inhibition of CK2 by silmitasertib was additive with respect to blocking cell proliferation, while cytotoxicity was reduced when compared to solely blockade of CK2. Therefore, parallel blockade TMEM16A by niclosamide may assist with anticancer therapy by silmitasertib.

SUBMITTER: Pinto MC

PROVIDER: S-EPMC7291285 | biostudies-literature | 2020 May

REPOSITORIES: biostudies-literature

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Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation.

Pinto Madalena C MC Schreiber Rainer R Lerias Joana J Ousingsawat Jiraporn J Duarte Aires A Amaral Margarida M Kunzelmann Karl K

Cells 20200505 5

Casein kinase 2 (CK2) is a highly ubiquitous and conserved serine/threonine kinase that forms a tetramer consisting of a catalytic subunit (CK2α) and a regulatory subunit (CK2β). Despite being ubiquitous, CK2 is commonly found at higher expression levels in cancer cells, where it inhibits apoptosis, and supports cell migration and proliferation. The Ca2+-activated chloride channel TMEM16A shows similar effects in cancer cells: TMEM16A increases cell proliferation and migration and is highly expr ...[more]

PMID: 32380794

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Project description:In cultures of normal human epidermal keratinocytes (NHEKs), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces the expression of the epidermal growth factor receptor ligands transforming growth factor-? (TGF-?) and epiregulin (EREG). TCDD also down-regulates EGF receptors (EGFR), suggesting that decreases in signaling contribute to the effects of TCDD. In this study, we treated post-confluent NHEKs with 10 nM TCDD and assessed its effects on EGFR binding, EGFR ligand secretion, basal ERK activity, and proliferation. TCDD caused time-dependent deceases in [(125)I]-EGF binding to levels 78% of basal cell values at 72 h. Amphiregulin (AREG) levels increased with time in culture in basal and TCDD-treated cells, while TGF-? and epiregulin (EREG) secretion were stimulated by TCDD. Inhibiting EGFR ligand release with the metalloproteinase inhibitor batimastat prevented EGFR down-regulation and neutralizing antibodies for AREG and EREG relieved receptor down-regulation. In contrast, neutralizing TGF-? intensified EGFR down-regulation. Treating NHEKs with AREG or TGF-? caused rapid internalization of receptors with TGF-? promoting recycling within 90 min. EREG had limited effects on rapid internalization or recycling. TCDD treatment increased ERK activity, a response reduced by batimastat and the neutralization of all three ligands indicating that the EGFR and its ligands maintain ERK activity. All three EGFR ligands were required for the maintenance of total cell number in basal and TCDD-treated cultures. The EGFR inhibitor PD1530305 blocked basal and TCDD-induced increases in the number of cells labeled by 5-ethynyl-2'-deoxyuridine, identifying an EGFR-dependent pool of proliferating cells that is larger in TCDD-treated cultures. Overall, these data indicate that TCDD-induced EGFR down-regulation in NHEKs is caused by AREG, TGF-?, and EREG, while TGF-? enhances receptor recycling to maintain a pool of EGFR at the cell surface. These receptors are required for ERK activity, maintenance of total cell number, and stimulating the proliferation of a small subset cells.

Dataset Information

Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation.

Publications

Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation.

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