In blood cells, changes in intracellular Ca²⁺ concentration ([Ca²⁺]_i) are associated with multiple cellular events, including activation of cellular kinases and phosphatases, degranulation, regulation of cytoskeleton binding proteins, transcriptional control, and modulation of surface receptors. Although there is no doubt as to the significance of Ca²⁺ signaling in blood cells, there is sparse knowledge about the molecular identities of the plasmalemmal Ca²⁺ permeable channels that control Ca²⁺ fluxes across the plasma membrane and mediate changes in [Ca²⁺]_i in blood cells. Using RNA expression analysis, we have shown that human leukemia K562 cells endogenously co-express transient receptor potential vanilloid channels type 5 (TRPV5) and type 6 (TRPV6) mRNAs. Moreover, we demonstrated that TRPV5 and TRPV6 channel proteins are present in both the total lysates and the crude membrane preparations from leukemia cells. Immunoprecipitation revealed that a physical interaction between TRPV5 and TRPV6 may take place. Single-channel patch-clamp experiments demonstrated the presence of inwardly rectifying monovalent currents that displayed kinetic characteristics of unitary TRPV5 and/or TRPV6 currents and were blocked by extracellular Ca²⁺ and ruthenium red. Taken together, our data strongly indicate that human myeloid leukemia cells co-express functional TRPV5 and TRPV6 calcium channels that may interact with each other and contribute into intracellular Ca²⁺ signaling.