The tumour suppressor alternative reading frame (ARF) is one of the most frequently mutated proteins in human cancer. It has been well established that ARF is able to stabilize and activate p53 by directly inhibiting Mdm2. ARF-mediated p53 activation in response to oncogenic stress is thought to be an important determinant of protection against cancer. However, little is known regarding the control of ARF in cells. Here, we show that Siva1 is a specific E3 ubiquitin ligase of ARF. Siva1 physically interacts with ARF both in vitro and in vivo. Through direct interaction, Siva1 promotes the ubiquitination and degradation of ARF, which in turn affects the stability of p53. Functionally, Siva1 regulates cell cycle progression and cell proliferation in an ARF/p53-dependent manner. Our results uncover a novel regulatory mechanism for the control of ARF stability, thereby revealing an important function of Siva1 in the regulation of the ARF-Mdm2-p53 pathway.