The bHLH-LZ (basic region/helix-loop-helix/leucine zipper) oncoprotein Myc and the bHLH-LZ protein Max form a binary transcription factor complex controlling fundamental cellular processes. Deregulated Myc expression leads to neoplastic transformation and is a hallmark of most human cancers. The dynamics of Myc transcription factor activity are post-translationally coordinated by defined protein-protein interactions. Here, we present evidence for a second messenger controlled physical interaction between the Ca2+sensor calmodulin (CaM) and all Myc variants (v-Myc, c-Myc, N-Myc, and L-Myc). The predominantly cytoplasmic Myc:CaM interaction is Ca2+-dependent, and the binding site maps to the conserved bHLH domain of Myc. Ca2+-loaded CaM binds the monomeric and intrinsically disordered Myc protein with high affinity, whereas Myc:Max heterodimers show less, and Max homodimers no affinity for CaM. NMR spectroscopic analyses using alternating mixtures of 15N-labeled and unlabeled preparations of CaM and a monomeric Myc fragment containing the bHLH-LZ domain corroborate the biochemical results on the Myc:CaM interaction and confirm the interaction site mapping. In electrophoretic mobility shift assays, addition of CaM does not affect high-affinity DNA-binding of Myc:Max heterodimers. However, cell-based reporter analyses and cell transformation assays suggest that increasing CaM levels enhance Myc transcriptional and oncogenic activities. Our results point to a possible involvement of Ca2+ sensing CaM in the fine-tuning of Myc function.