Heterotrimeric G proteins (Galphabetagamma) play an essential role in coupling membrane receptors to effector proteins such as ion channels and enzymes. Among the five mammalian Gbeta-subunits cloned, the human G protein beta4 has not been described. The purpose of the present study was to functionally characterize the newly identified human Gbeta4 subunit. The Gbeta4 open reading frame (ORF) was amplified utilizing PCR from brain cDNA. Amplification primers were generated following 5' rapid amplification of cDNA ends (5'-RACE) from an expressed sequence tag (EST) containing the predicted 3' end of the protein. Multiple tissue cDNA panel analysis showed that Gbeta4 mRNA was strongly expressed in lung and placenta, whereas it is weakly expressed in brain and heart. Heterologous overexpression of Gbeta4gamma2 or Gbeta4gamma4 in rat sympathetic neurons resulted in tonic modulation of N-type voltage-gated Ca(2+) and G protein-gated inwardly rectifying K(+) currents. Furthermore, coexpression of Gbeta4gamma2 and Galpha(oA) resulted in heterotrimer formation. These results show that the newly cloned Gbeta subunit shares several properties with other human Gbeta family members.