Studies of the folate chemotactic receptor ofvegetative Dictyostelium discoideum cells have been hampered by the presence of the degradative enzyme folate deaminase. The diaminopterin compounds aminopterin and methotrexate (MTX) are chemoattractants but are not attacked by the deaminase. [3', 5', 7, 9-3H] Vegetative Dictyostelium discoideum cells are chemotactic to folic acid and pterins (1). Because bacteria liberate these compounds, this chemotactic system has been considered to be a food-seeking device (2); however, growing evidence suggests that folate may have a role in early development (3, 4). Whatever its role, it should be of interest to compare the folate chemotactic system with the cyclic AMP (cAMP) system of aggregating cells. In particular, it seems likely that receptors for both chemoattractants share a common transducer mechanism by which the extracellular signal is coupled to the response of directional motility. A reflection of this general mechanism may be the finding that stimulation of cells in the appropriate stage ofdevelopment with either folate or cAMP results in a transient increase in intracellular cyclic GMP (cGMP) levels (5, 6).Whereas both the cAMP receptor (7, 8) and degradative enzyme (9) 3':5'-cycic-nucleotide phosphodiesterase (cNPDEase, EC 3.1.4.17) of differentiating cells have been well characterized, less is known about the analogous components ofthe folate system. In this vegetative chemotactic system, the degradative enzyme is a pterin deaminase (EC 3.5.4.11) (10,11) with a cellular distribution similar to that of the cNPDEase (7, 9-11). To perform radioligand binding studies with the folate receptor, it is necessary to either inhibit the deaminase or to use a ligand that is not attacked by the enzyme. We and others (11,12) For both methods, cells were suspended at the indicated density (see in 17 mM sodium/potassium phosphate (pH 6.4) at 40C. [3H]MTX was added at 50-70 nM, and a 0.5-ml aliquot periodically was taken for assay. Filtration was on 10-place Hoeffer manifolds with 0.8-,m polycarbonate filters (BioRad), followed by a 1.0-ml wash with cold buffer. For the centrifugation assay, cells were incubated with [3H]MTX for 1.5 min and the 0.5-ml aliquots were layered above 1.0 ml of 12% (wt/vol) polyethylene glycol 6000 in a 1.5-ml polypropylene tube (Bio-Rad) and centrifuged for 30 s in a Beckman Microfuge B at 40C. The filters or the resuspended cell pellets were measured for radioactivity in 10 ml of 3a70 scintillation fluid (Research Products, Elk Grove, IL). Nonspecific binding was determined in the presence of 50 ,uM MTX. Chemotaxis assays were performed and photographed as described (14). The gradient of MIX formed in the 2% (wt/vol) agar well plates was determined by placing 1 AM MIX (containing 200,000 dpm of [3H]MIX) in the center well ofthe plates. After 3 hr ofdiffusion at room temperature, several 3-mm strips ofagar were cut from three plates, cut into 1-mm slices with a gel slicer, and measured for radioactivity. The logarithm ofMIX concentration...

For over 30 years, the cellular slime molds, and in particular the species Dictyostelium discoideum, have been viewed as a eukaryotic model system for the study of a variety of regulatory and developmental processes [l]. In particular, the transition from the free-living amoeboid stage to the multicellular slug stage has been the focus of a great deal of attention by those investigators studying the regulation of development [ 1-31. The first morphological evidence that differentiation of a starved population of cells is proceeding on schedule is the appearance of ripples or streams in a lawn of cells resting on a moist surface [l-31. This prelude to the readily identified tight aggregate stage indicates the functioning of the cells' endogenous chemosensory system. In 1969, Bonner and coworkers [4] identified the endogenously produced chemoattractant or acrasin of these cells as cyclic AMP (CAMP). While cAMP is widely encountered as an intracellular regulatory agent or second messenger, the Dictyostelia provide one of the few well documented examples of cAMP as an extracellular messenger molecule.The Polysphondylia such as P violaceum and P pallidum appear to employ a nonnucleotide molecule as their acrasin during starvation induced chemotaxis. This compound has been studied by Bonner and coworkers [S] and has been tentatively identified as a peptide with a blocked amino terminus, although further characterization is required for unambiguous identification. Bonner's group has also been responsible for the identification of folic acid and pterin compounds as chemoattractants of vegetative Dictyostelium amoebae [6-81. This identification of a nonacrasin chemoattractant raises the possibility of studying the transition from a vegetative to a developmental mode of chemotaxis. DEVELOPMENTAL CHEMOTAXIS IN D discoideumThe phenomenology of the CAMP chemotactic system in D discoideum has been well established [9,10]. It is convenient for descriptive purposes to imagine