Localization of the mate-recognition pheromone in Brachionus plicatilis O.F. Müller (Rotifera) by fluorescent labeling with lectins

Abstract

Mate-recognition in the rotifer Brachionus plicatilis O.F. Müller is based upon male contact chcmoreception of a glycoprotein pheromone on the body surface of females. The highest densities of this glycoprotein were found on the corona of females as determined by fluorescence microscopy. A time series experiment demonstrated that binding sites on the corona fluoresced before any other structure and did so most intensely. The lorica margin of females fluoresced weakly after 20 min of exposure, suggesting that the mate-recognition pheromone (MRP) is present, but in low density. In males, the corona also was the most intensely labeled structure on the body surface. Pretreatment of rotifers with the lectins Concanavalin A, (ConA), Lens culinaris, Vicia faba and Pisum sativum blocked subsequent labeling with ConA-fluoroisothiocyanate (FITC). Exposure to other lectins did not block ConA-FITC-labeling. Quantitation of fluorescence intensity using image analysis demonstrated that the contrast between a glycoprotein signal and background ranged from 2 to 19.5. In situ degradation of the MRP by the glycohydrolasc N-glycanase reduced lectin-binding and the intensity of fluorescent-labeling. Since N-glycanase deglycosylation of glycoprotcin on the body surface of live females sharply reduced their ability to elicit male-mating reactions, it is clear that the same oligosaccharides critical for lectin-binding also are essential for male-recognition. These lectin probes have enabled us to visualize the distribution of the MRP on the body surface of females and provided insight into the copulatory behavior of males. © 1993.