Significance Hyperactivity of the hormone glucagon plays an important role in the pathophysiology of type 2 diabetes, but the factors that affect glucagon levels and α-cell proliferation are not entirely understood. This is particularly important for the development diabetes drugs based on glucagon receptor inhibition, which increase glucagon levels in plasma and α-cell mass. Here we show that increased levels of Angiopoietin-like 4 (Angptl4) in adipose tissue and plasma are sufficient to induce α-cell proliferation. Angptl4 is a conserved, secreted lipoprotein lipase inhibitor expressed by many tissues that is regulated by exercise and feeding. Moreover, Angptl4 is required for the compensatory hyperglucagonemia and α-cell proliferation following treatment with glucagon receptor antagonists.