Oncogene-induced senescence acts as a barrier against tumour formation and has been implicated as the mechanism preventing the transformation of benign melanocytic lesions that frequently harbour oncogenic B-RAF or N-RAS mutations. In the present study we systematically assessed the relative importance of the tumour suppressor proteins p53, p21Waf1, pRb and p16INK4a in mediating oncogene-induced senescence in human melanocytes. We now show that oncogenic N-RAS induced senescence in melanocytes is associated with DNA damage, a potent DNA damage response and the activation of both the p16INK4a/pRb and p53/p21Waf1 tumour suppressor pathways. Surprisingly neither the pharmacological inhibition of the DNA damage response pathway nor silencing of p53 expression had any detectable impact on oncogene-induced senescence in human melanocytes. Our data indicate that the pRb pathway is the dominant effector of senescence in these cells, as its specific inactivation delays the onset of senescence and weakens oncogene-induced proliferative arrest. Furthermore, we show that although both p16INK4a and p21Waf1 are upregulated in response to N-RASQ61K, the activities of these CDK inhibitors are clearly distinct and only the loss of p16INK4a weakens senescence. We propose that the ability of p16INK4a to inhibit the cyclin D-dependent kinases and DNA replication, functions not shared by p21Waf1, contribute to its role in senescence. Thus, in melanocytes with oncogenic signalling only p16INK4a can fully engage the pRb pathway to alter chromatin structure and silence the genes that are required for proliferation.