We have previously shown that the activity of the Ca²⁺ channel TRPV6 was increased in the presence of bis- (N, N-dimethyl-hydroxamido) hydrooxovanadate (DMHV) an inhibitor of the tyrosine phosphatase PTP1B (Sternfeld et al. Cell Signal 17, 2005). When HEK293 cells were co-expressed with TRPV6 and or its mutant TRPV6 (Y161,162F) with different tyrosine kinases or phosphatases we found that tyrosine kinase Src and the protein tyrosine phosphatase 1B (PTP1B) phosphorylates and dephosphorylates, respectively, the tyrosines in position 161/162. The DMHV-effect on Ca²⁺ influx was abolished in the TRPV6 (Y161,162F) mutant as compared to the wild type TRPV6. Using the "bimolecular fluorescence complementation method", the 2-hybrid system and co-immunoprecipitation, interaction of TRPV6 and PTP1B could be shown for which the N-terminal cytosolic part of TRPV6 was essential. We conclude that phosphorylation/dephosphorylation of tyrosines in position 161/162 is essential for regulation of Ca²⁺ influx through TRPV6-Ca²⁺ channels.