The angiosperm family Proteaceae is a distinct component of the Cape Floristic Region biodiversity hotspot with 330 endemic species. Phylogenetic analyses of subfamily Proteoideae using sequence data from one nuclear and six plastid loci show that most of this diversity is contained in two distinct Cape floral clades. Molecular dating analyses, using Bayesian and penalized likelihood methods and four phylogenetically supported fossil age constraints, reveal contrasting histories for these two clades. The genus Protea belongs to a lineage that may have been in Africa since the Late Cretaceous but began to diversify in the Cape only 5-18 Myr ago. In contrast, the Leucadendrinae clade presumably arrived in the region no earlier than 46 Myr ago by long-distance dispersal from an Australian ancestor and the extant members of this clade began to diversify in the Cape 22-39 Myr ago. These results join a growing number of case studies that challenge the commonly accepted view that most of the Cape flora radiated synchronously in the Late Miocene and Early Pliocene when a Mediterranean climate settled in the region.