A developmentally regulated cell surface receptor for a density-sensing factor in Dictyostelium.
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abstract
Conditioned medium factor (CMF) is an 80-kDa glycoprotein which is the signal in a cell density-sensing system used by developing Dictyostelium cells. CMF is slowly secreted by cells when they starve, and the extracellular level of CMF then becomes an indicator of the density of starving cells. To examine how CMF is sensed, we have made bacterially synthesized recombinant CMF and found that it has as much activity as native CMF, indicating that glycosylation is not part of the active site of CMF. Expression of recombinant fragments of CMF indicates that the active site lies within an 88-amino acid region near the N terminus. To determine whether CMF is sensed by cell surface receptors, we examined binding of iodinated recombinant CMF to live cells. We found saturable binding to 6-h starved cells at 3.9 x 10(4) molecules/cell with a KD of 2.1 nM. The binding saturates in 30 min, and a Scatchard plot indicates that there is only one class of receptor. The binding is competed off by the addition of either the native or recombinant CMF, or the 88-amino acid active fragment region; no binding competition is seen from the nonactive regions or other proteins. Very little binding to vegetative cells is seen, with maximal binding seen in cells starved for 6-8 h. The amount of cell surface CMF binding then decreases during later development. Normal levels of CMF binding are seen to CMF- cells, indicating that CMF is not required for the accumulation of its own receptor.
