Hemoglobin is an abundant protein in the host vascular compartment and a source of iron, heme, and amino acids for many pathogens. The human fungal pathogen Candida albicans uses hemoglobin as an iron source as well as a signaling molecule to alter gene expression and induce adhesion to several extracellular matrix proteins. We now report that hemoglobin can promote true hyphal morphogenesis. Hemoglobin added to yeast cells at 37 degrees C rapidly induced expression of the hypha-specific genes HWP1 and ECE1 coincident with the pattern of hyphal development. A synthetic medium buffered with phosphate at pH 7.2 and containing physiological glucose (5 mM) and low ammonium ion (0.1 mM) was optimal for the response to hemoglobin. High glucose (110 mM), high ammonium ion (20 mM), and 0.1 mM glutamine were all inhibitory. Heme, free globin, or immobilized hemoglobin could not replicate the activity of hemoglobin to induce germ tubes or hypha-specific gene expression at 37 degrees C under optimized conditions. This implicates the previously described Hb-signaling receptor in hyphal formation. This response was also dependent upon the presence of the morphogenesis regulator Efg1p, but the MAP-kinase specific transcription factor Cph1p was not required. These data define a role for the host-factor hemoglobin in Efg1p-dependent hyphal development.