Host growth factors induce proliferation of Trypanosoma cruzi amastigotes by mechanisms that remain poorly defined. Here we examined human epidermal growth factor (EGF) for its ability to bind to the mammalian multiplicative forms of T. cruzi and to induce growth of the parasites. EGF stimulated incorporation of [³H] thymidine into DNA and growth of amastigotes both in a concentration-dependent manner. Radiolabeled EGF was found to bind to amastigotes in a concentration-dependent and saturable manner but it did not bind to trypomastigotes. Scatchard analysis showed a single class of receptors with a K_d of 0.8 nM and numbering 3.1 × 10³ per amastigote. Results from internalization experiments provided evidence of receptor-mediated endocytosis of EGF. Northern analysis showed a 3.0-kb transcript for the putative EGF receptor (EGFR) homologue in amastigotes, but not trypomastigotes. Binding of EGF to amastigotes induced signal transduction events. EGF induced “in vitro” kinase activity as determined by γ;en[³²P] ATP incorporation into amastigote proteins. EGF also increased protein kinase C activity in a concentration-dependent manner and Mitogen Activated Protein (MAP) kinase activity in a time- and concentration-dependent manner. A specific inhibitor (AG14782) of the EGFR and a MAP kinase inhibitor (PD98059) decreased EGF-dependent T. cruzi MAP kinase activity. These results describe a novel mechanism used by amastigotes to regulate their proliferation mediated by an EGF-dependent signal transduction pathway.