The receptor for erythropoietin (Epo) belongs to the cytokine receptor family and lacks a tyrosine kinase domain. However, it has been hypothesized that a tyrosine kinase, Jak2, associates with the membrane proximal cytoplasmic region of Epo receptor (EpoR) and mediates the growth signaling from the receptor through tyrosine phosphorylation of cellular substrates. To explore the growth signaling pathways from the EpoR, we analyzed substrates of tyrosine phosphorylation induced by Epo stimulation in cells expressing various mutant EpoRs. The vav proto- oncogene product was found to be tyrosine phosphorylated after Epo stimulation in cells expressing the wild-type EpoR or a truncated receptor, H mutant, that retains the growth signaling function. In these cells, Epo also induced the expression of a serine/threonine kinase, Pim-1. However, Epo stimulation did not have any effect on Vav or Pim-1 in cells expressing a mutant EpoR, PM4 mutant, inactivated by a point mutation, Trp282 to Arg, in the membrane proximal region, which abrogates the interaction with Jak2. On the other hand, both tyrosine phosphorylation of Vav and expression of Pim-1 were observed constitutively in cells expressing a mutant EpoR that is constitutively activated by a point mutation, Arg 129 to Cys, in the extracellular domain. Jak2 was also constitutively tyrosine phosphorylated and activated in cells expressing this mutant, which confirms the crucial role of Jak2 in growth signaling from the EpoR. Taken together, these observations suggest that the tyrosine phosphorylation of Vav and the expression of Pim-1 may play important roles in growth signaling from the EpoR.