The structure of the chorion and associated surface filaments in Oryzias--evidence for the presence of extracellular tubules.

The structure of the chorion with its associated surface filaments has been examined in Oryzias latipes using several techniques, including scanning and transmission electron microscopy, enzymatic digestion, and sodium dodecylsulfate-polyacrylamide gel electrophoresis. The chorion of the recently fertilized egg was found to be organized into three zones: an outer, fuzzy electron-lucent zone that was continuous over the surface of filaments, a middle, homogeneous electron-dense zone, and an inner zone of ten to 12 horizontal, fibrous lamellae. Two topographically distinct types of filaments were found on the chorionic surface: nonattaching and attaching. Nonattaching filaments showed a regular spatial distribution over the chorion with an interfilament distance of about 60-70 microns. Attaching filaments originated from a localized portion of the chorion and united with those of neighboring eggs to anchor the egg cluster to the gonoduct of the female. Both nonattaching and attaching filaments were morphologically regionalized into basal and distal segments. Internally, nonattaching and attaching filaments were constructed of unbranched, packed tubules with an average outside diameter of approximately 19.5 and 18.8 nm, respectively. Using the attaching filament for further study, it was determined by rotational analysis (Markham et al., '63) that the wall of each tubule was a cylinder composed of 14 globular subunits. Two structural types of attaching filaments were identified. The type I attaching filament was similar in internal organization to the nonattaching filament and consisted of only tubules. The type II attaching filament, however, showed a highly osmiophilic, electron-dense bar surrounded by packed tubules. Tubules of attaching filaments of the adult were resistant to the action of Triton X-100 and colchicine, but sensitive to a 0.1% protease solution. However, colchicine-treated ovary tissue showed an absence and pattern of disorganization of tubules at the periphery of developing filaments. Solubilized attaching filament samples electrophoresed on 7.5% polyacrylamide-SDS gels were resolved into a pair of Coomassie-blue-positive bands that comigrated with purified porcine brain tubulin. The apparent molecular weight of the attaching filament polypeptide was determined to be approximately 55,000 daltons. These data suggest that the extracellular, tubular components of attaching filaments (as well as nonattaching filaments) are proteinaceous and show properties similar to those of cytoplasmic microtubules. Tubular precursor material was electron-dense and appeared to originate in the cisternae of the rough endoplasmic reticulum of ovarian foll