Obesity is a disorder of energy balance 1. Hormone-sensitive lipase (HSL) mediates the hydrolysis of triacylglycerol 2, the major form of stored energy in the body. Perilipin (encoded by the gene Plin), an adipocyte protein, has been postulated to modulate HSL activity 3, 4, 5. We show here that targeted disruption of Plin results in healthy mice that have constitutively activated fat-cell HSL. Plin−/− mice consume more food than control mice, but have normal body weight. They are much leaner and more muscular than controls, have 62% smaller white adipocytes, show elevated basal lipolysis that is resistant to β-adrenergic agonist stimulation, and are cold-sensitive except when fed. They are also resistant to diet-induced obesity. Breeding the Plin−/− alleles into Lepr db/db mice reverses the obesity by increasing the metabolic rate of the mice. Our results demonstrate a role for perilipin in reining in basal HSL activity and regulating lipolysis and energy balance; thus, agents that inactivate perilipin may prove useful as anti-obesity medications.