Identification and initial characterization of an autocrine pheromone receptor in the protozoan ciliate Euplotes raikovi.
The polypeptide pheromone Er-1, purified from the ciliate Euplotes raikovi of mating type I and genotype mat-1/mat-1, was iodinated with 125I-Bolton-Hunter reagent to a sp act of 0.45-0.73 mu Ci/microgram of protein. This preparation of 125I-Er-1 bound specifically to high affinity binding sites on the same cells of mating type I. Binding of 125I-Er-1 occurred with an apparent Kd of 4.63 +/- 0.12 X 10(-9) M in cells in early stationary phase. It was estimated that these cells carry a total number of approximately 5 X 10(7) sites/cell, with a site density that falls in the range of 1,600-1,700/microns 2 of cell surface. Unlabeled Er-1, other homologous pheromones such as Er-2 and Er-10, antibodies specific for Er-1, and human IL-2 were shown to act as effective inhibitors of specific binding of 125I-Er-1 to mating type I cells. The "autocrine" nature of the identified specific high affinity binding sites for Er-1 was further substantiated by cross-linking experiments. These experiments revealed that mating type-I cell membranes contain one protein entity of Mr = 28,000 that is capable of reacting specifically with the homodimeric native form of Er-1.
Ortenzi
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C
Department of Cell Biology, University of Camerino, Italy.
Miceli
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C
Bradshaw
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RA
Luporini
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P
D000818:Animals; D001667:Binding, Competitive; D016798:Ciliophora / Q000502:physiology*; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008565:Membrane Proteins; D008970:Molecular Weight; D010455:Peptides / Q000378:metabolism*; D010675:Pheromones / Q000378:metabolism; D015800:Protozoan Proteins; D011956:Receptors, Cell Surface / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D051964:Receptors, Mating Factor; D018000:Receptors, Peptide; D014157:Transcription Factors
D008565:Membrane Proteins; D010455:Peptides; D010675:Pheromones; D015800:Protozoan Proteins; D011956:Receptors, Cell Surface; D051964:Receptors, Mating Factor; D018000:Receptors, Peptide; D014157:Transcription Factors; C057783:mating pheromone Er-1, Euplotes raikovi