Journal
EUROPEAN JOURNAL OF PHARMACOLOGY
Volume 895, Issue -, Pages -Publisher
ELSEVIER
DOI: 10.1016/j.ejphar.2021.173881
Keywords
Volume-regulated chloride channel; DCPIB; TMEM16A; LRRC8A; Oral cancer; Cell proliferation
Categories
Funding
- KAKENHI from the Ministry of Education, Science, Sports, and Culture of Japan [16K11713]
- Grants-in-Aid for Scientific Research [16K11713] Funding Source: KAKEN
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The study found that VRAC is highly functional and associated with proliferation in HST-1 cells, but not in HaCaT cells. The pharmacological profile of VRAC in HST-1 was similar to previous reports, and inhibition of VRAC with DCPIB led to apoptosis in HST-1 cells. Knockdown of TMEM16A and LRRC8A attenuated VRAC activity in HST-1 cells.
Objectives: Volume-regulated anion channels (VRACs), expressed in various cells, play an important role in cell volume regulation. Despite being physiologically defined almost half a century ago, only the molecular candidates of VRAC, TMEM16A, LRRC8A, and bestrophin-1 (BEST1), are known. Here, we aimed to explore the functional significance of VRAC in, HST-1, an oral squamous cell carcinoma (OSCC) cell line. Methods: Cell proliferation assays, RT-PCR, Western blot, and flow cytometry were used to estimate changes in gene expression and cell proliferation. Ion channel activity was recorded using the patch-clamp technique. Specific genes were knocked-down by siRNA assays. Results: VRAC, identified as a hypotonicity-induced current, was highly functional and associated with the proliferation of HST-1 cells but not of HaCaT (a normal keratinocyte) cells. The pharmacological profile of VRAC in HST-1 was similar to that reported previously. DCPIB, a specific VRAC inhibitor, completely inhibited VRAC and proliferation of HST-1 cells, eventually leading to apoptosis. VRAC in HST-1 was attenuated by the knockdown of TMEM16A and LRRC8A, while knockdown of BEST1 affected cell proliferation. In situ proximity ligation assay showed that TMEM16A and LRRC8A co-localized under isotonic conditions (300 mOsM) but were separated under hypotonic conditions (250 mOsM) on the plasma membrane. Conclusions: We have found that VRAC acts to regulate the proliferation of human metastatic OSCC cells and the composition of VRAC may involve in the interactions between TMEM16A and LRRC8A in HST-1 cells.
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