Proteins of molecular mass 46,000 (p46) and 34,000-39,000 (p36) daltons are phosphorylated at tyrosine in Rous sarcoma virus-transformed chicken and mouse fibroblasts. p46 has recently been identified as an isozyme of enolase but the function of p36 is unknown. The expression of these proteins in various mouse and rat tissues has been examined. In most tissues, except muscle, p46 is found at relatively constant levels. In muscle, a more basic, related protein is present. In contrast, the abundance of p36 varies more widely from tissue to tissue, suggesting that it has a function in some but not all differentiated cells. By SDS gel electrophoresis and immunoblotting, high levels of p36 (60-120% of its relative abundance in fibroblasts) were found in small intestine, lung, and thymus, and intermediate levels (20-50%) were found in spleen, lymph nodes, and testes. No p36 was detectable in brain and muscle. Where studied, p36 mRNA expression paralleled protein levels. The cell types within each tissue expressing p36 were identified by immunofluorescence and immunoperoxidase staining. These cell types include all endothelial cells and fibroblastic cells examined, as well as various epithelial cells, cardiac muscle cells, macrophages, and testicular interstitial cells. We were unable to detect p36 in skeletal or smooth muscle cells, erythrocytes, nerve cells, or lymphocytes in any of the examined tissues. p36 appears to be concentrated in the terminal web region of intestinal columnar epithelial cells.