4μ8C Inhibits Insulin Secretion Independent of IRE1α RNase Activity

IRE1α plays an important role in the unfolded protein response (UPR), which is activated by the accumulation of unfolded proteins in the endoplasmic reticulum. 4μ8C, a well-known inhibitor of IRE1α RNase activity, is commonly used to analyze IRE1α function during ER stress in cultured mammalian cells. However, the off-target effects of 4μ8C remain elusive. Pancreatic β-cells synthesize a large amount of insulin in response to high glucose stimulation, and IRE1α plays an important role in insulin secretion from pancreatic β-cells. Here, to analyze the role of IRE1α in pancreatic β-cells, we examined insulin secretion after 4μ8C treatment. Although 4μ8C inhibited insulin secretion within 2 hr, neither insulin synthesis nor maturation was inhibited by 4μ8C under the same conditions. This result prompted us to examine the precise effects of 4μ8C on insulin secretion in pancreatic β-cells. Unexpectedly, with just 5 min of treatment, 4μ8C blocked insulin secretion in cultured pancreatic β-cells as well as in pancreatic islets. Furthermore, insulin secretion was prevented by 4μ8C, even in pancreatic β-cells lacking the IRE1α RNase domain, suggesting that 4μ8C blocked the late stage of the insulin secretory process, independent of the IRE1α-XBP1 pathway. Our results indicate that 4μ8C has an off-target effect on insulin secretion in pancreatic β-cells. These findings inform the researchers in the field that the use of 4μ8C requires the special consideration for the future studies.Key words: 4μ8C, XBP1, insulin, IRE1α, pancreatic β-cells.