20-Hydroxyecdysone is required for, and negatively regulates, transcription of Drosophila pupal cuticle protein genes.

Transcripts of ecdysone-dependent genes (EDGs) accumulate in isolated imaginal discs with 8 hr after exposure to a pulse of the steroid hormone 20-hydroxyecdysone (20-HE; 1 microgram/ml for 6 hr) but not in discs cultured in the continuous presence or absence of the hormone. Sequence analyses show that two of the EDGs are members of gene families encoding insect cuticle proteins. We conclude that a third EDG encodes a cuticle protein because the conceptual glycine-rich protein contains sequence motifs similar to those found in insect egg shell proteins and vertebrate cytokeratins and because expression of this gene is limited to tissues that deposit the pupal cuticle. Nuclear run-on assays show that the hormone-dependent expression of each of these EDGs is due to transcriptional regulation. Readdition of hormone to imaginal discs actively synthesizing the EDG messages causes rapid repression of EDG transcription. Thus, 20-HE acts as both a positive and a negative regulator of EDG transcription. Sequences in the promoter regions of two of the EDGs are similar to an ecdysone response element and may play a role in negative regulation.

Morphometric and cytochemical analysis of lysosomes in rat Peyer's patch follicle epithelium: their reduction in volume fraction and acid phosphatase content in M cells compared to adjacent enterocytes.

M cells are specialized epithelial cells over lymphoid follicles in Peyer's patches which take up viruses, bacteria, and antigenic macromolecules from the intestinal lumen. Unlike ordinary enterocytes which sequester pinocytosed material in lysosomes, M cells transport such material across the epithelium to antigen-processing areas in lymphoid follicle domes, suggesting a difference in lysosomal activity or a different route for movement of endocytic vesicles. Ileal Peyer's patches in rats were examined by electron microscopy to identify lysosomes by acid phosphatase activity. Acid phosphatase was found in dense bodies in enterocytes but not in M cells. Stereological analysis showed the volume fraction occupied by dense bodies in M cells to be 16 times less than in enterocytes (P less than .0005), even though the volume fractions of cytoplasm occupied by mitochondria in M cells and enterocytes were not significantly different. The small volume fraction of dense bodies and the absence of acid phosphatase activity in M cells thus correlate with absence of lysosomal degradation of luminal microorganisms during transport into lymphoid follicles by M cells and may provide not only a complete array of microbial antigens for initiation of immune responses, but also a route through the mucosal barrier for microorganisms which can evade local containment mechanisms.

M cell transport of Vibrio cholerae from the intestinal lumen into Peyer's patches: a mechanism for antigen sampling and for microbial transepithelial migration.

Viable Vibrio cholerae O1 were inoculated into the intestinal lumen of nonimmune rabbits. The vibrios were phagocytosed by M cells over Peyer's patch lymphoid follicles, carried in vesicles through the epithelium, and discharged among underlying lymphocytes and macrophages. Autoradiography of V. cholerae labeled with [2-3H]adenine confirmed transport. Indigenous bacteria with and without capsules were also taken up from control loops and carried through M cells into Peyer's patches. V. cholerae killed by acidification, formalin, heat, or UV irradiation were not taken up, a result that may have relevance for development of oral vaccines. Ruthenium red stain revealed gaps in the layer of mucus over M cells, glycocalyx bridging the space between vibrios and M cell microvilli, and knobby projections over membranes of M cell microvilli; these projections were not found over absorptive cells. M cells thus convey viable enteric microbes, including V. cholerae that are not otherwise invasive, into intestinal lymphoid tissue, where mucosal immune responses are initiated. Uptake and transport by M cells may also assist certain pathogenic bacteria in traversing the mucosal barrier and initiating systemic infection.