Krev-1/rap1A is an evolutionarily conserved Ras-family GTPase whose cellular function remains unclear, but which has been proposed to function as a tumor suppressor gene, and may act as a Ras antagonist. To elucidate Krev-1 activity, we have used LexA-Krev-1 in a two-hybrid screen of a HeLa cell cDNA library. Of the two cDNA classes isolated, one contained a single isolate encoding the known Krev-1 interactor Raf, while the second contained multiple isolates coding for a previously undescribed protein which we have designated Kritl (for Krev Interaction Trapped 1). The full length Krit1 cDNA encodes a protein of 529 amino acids, with an amino-terminal ankyrin repeat domain and a novel carboxy-terminal domain required for association with Krit1. Krit1 interacted strongly with Krev-1 but only weakly with Ras, suggesting it might specifically regulate Krev-1 activities. Krit1 mRNA and protein are expressed endogenously at low levels, with tissue specific variation. Intriguingly, the Krit cDNA has been mapped by FISH to chromosome 7q21-22, a region known to be frequently deleted or amplified in multiple forms of cancer.