The secretory contents of Paramecium tetraurelia trichocysts: ultrastructural--cytochemical characterization.
The secretory contents ("matrix") of Paramecium tetraurelia trichocysts expand by a factor of 4.5 when they undergo a Ca2+-mediated decondensation in the course of exocytosis. This is paralleled by a concomitant increase in the interval of the periodic banding of the matrix from 12 nm to 45-51 nm, which becomes visible with different electron stains for proteins and negatively charged groups. Recent reports of actin in secretory contents led us to investigate its redistribution and artifactual adsorption to the trichocyst contents upon their expansion. To visualize this effect we used peroxidase-labeled F(ab) fragments from an IgG directed against Paramecium actin, a DNAase I-gold complex, and the induction of F-actin polymerization. The trichocysts were analyzed in situ as well as after isolation by density-gradient centrifugation. Additionally, in response to current reports in the literature, we reanalyzed trichocyst contents for any possible presence of calmodulin. We applied three independent in situ methods for this: autofluorescence after trifluoperazine affinity labeling, calmodulin-fluorescence affinity labeling, and an electron microscopic immunocytochemical method. All three methods failed to reveal any significant labeling of structurally intact trichocysts in situ, although we also showed that discharged trichocysts avidly adsorb calmodulin from the culture medium. From the present data we conclude that the decondensation of trichocysts during exocytosis is mediated by a sudden conformational rearrangement of secretory proteins in the trichocyst contents, without the involvement of any other regulatory or contractile proteins, which occur only in the cytoplasm. Trichocyst contents are not significantly--if at all--glycosylated.
D000199:Actins / Q000032:analysis; D000345:Affinity Labels; D002147:Calmodulin / Q000032:analysis; D002499:Centrifugation, Density Gradient; D003470:Culture Media; D003593:Cytoplasm / Q000032:analysis*; D005089:Exocytosis; D006651:Histocytochemistry; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D010247:Paramecium / Q000032:analysis* / Q000378:metabolism / Q000648:ultrastructure; D014268:Trifluoperazine
D000199:Actins; D000345:Affinity Labels; D002147:Calmodulin; D003470:Culture Media; D014268:Trifluoperazine