A cytochemical ultrastructural study of the lysosomal system of different species of malaria parasites

We have used ultrastructural techniques in different malarial species to demonstrate a lysosomal system. First, we have tried to localize acid phosphatase, a typical lysosomal label. Its activity was localized in the endoplasmic reticulum and in endocytic vesicles, and in dense-cored vesicles near the digestive vacuoles, especially in Plasmodium falciparum (FCR3 strain). Then, we have studied the different cellular compartments of the malarial parasite by the zinc iodide-osmium tetroxide technique that heavily contrasted the cellular compartments of the parasite. This experiment led to the observation of a profound rearrangement of the endoplasmic reticulum, especially in P. berghei. A very atypical but functional Golgi apparatus was demonstrated in all the growing stages of the parasite and lysosome-like vesicles were observed, showing a structure very similar to those of the coated vesicles of a true Golgi complex. The presence of these organelles are in favor of the existence of a lysosomal system and of the endogenicity of some enzymes involved hemoglobin degradation.     

The Peripheral Vesicles of Trophozoites of the Primitive ProtozoanGiardia lambliaMay Correspond to Early and Late Endosomes and to Lysosomes

Giardia lamblia,a primitive eukaryotic cell, lacks organelles such as mitochondria, peroxisomes, and a typical Golgi complex and presents a system of vesicles located below the plasma membrane. We used fluorescence and electron microscopy to better characterize the peripheral vesicles. Incubation of living cells with acridine orange showed that the peripheral vesicles correspond to an acidic compartment. Incubation with lucifer yellow, and with horseradish peroxidase, showed labeling of the peripheral vesicles even after several hours. Acid phosphatase was localized in the endoplasmic reticulum and in most of the peripheral vesicles. On the other hand, glucose 6-phosphatase, an endoplasmic reticulum marker, was observed in the endoplasmic reticulum cisternae and in some peripheral vesicles. A similar labeling pattern was observed using the zinc iodide technique, which reveals SH-containing proteins. Three-dimensional reconstruction and electron microscopy tomography of cells stained for acid phosphatase and glucose-6-phosphatase revealed the connection between some vesicles and profiles of the endoplasmic reticulum. Taken together, our observations suggest that trophozoites ofG. lambliapresent an endosomal–lysosomal system concentrated in a single system, the peripheral vesicles, which may represent an ancient organellar system that later on subdivided into compartments such as early and late endosomes and lysosomes.     

Cytochemical characterization of the endomembranous system during the oocyte primary growth in Serrasalmus spilopleura (Teleostei, Characiformes, Characidae)

The morphophysiological changes that occur during oocyte primary growth in Serrasalmus spilopleura were studied using ultrastructural cytochemical techniques. In the previtellogenic oocytes endoplasmic reticulum components, Golgi complex cisternae and vesicles, lysosomes, multivesicular bodies and some electron-dense vesicles react to acid phosphatase (AcPase) detection. The endoplasmic reticulum components, Golgi complex cisternae and vesicles also react to osmium tetroxide and potassium iodide impregnation (KI). These structures, except for the Golgi complex cisternae, are strongly contrasted by osmium tetroxide and zinc iodide impregnation (ZIO). Some electron-dense vesicles are ZIO-stained, while microvesicles in the multivesicular bodies and other large isolated cytoplasmic vesicles are contrasted by KI. At primary oocyte growth, the activity of the endomembranous system and the proliferation of membranous organelles are intense. The biosynthetic pathway of the lysosomal proteins such as acid phosphatase, involves the endoplasmic reticulum, Golgi complex, vesicles with inactive hydrolytic enzymes and, finally, the lysosomes. The oocyte endomembranous system have reduction capacity and are involved in the metabolism of rich in SH groups.     

Early stages of the secretory pathway, but not endosomes, are required for Cvt vesicle and autophagosome assembly in Saccharomyces cerevisiae

The Cvt pathway is a biosynthetic transport route for a distinct subset of resident yeast vacuolar hydrolases, whereas macroautophagy is a nonspecific degradative mechanism that allows cell survival during starvation. Yet, these two vacuolar trafficking pathways share a number of identical molecular components and are morphologically very similar. For example, one of the hallmarks of both pathways is the formation of double-membrane cytosolic vesicles that sequester cargo before vacuolar delivery. The origin of the vesicle membrane has been controversial and various lines of evidence have implicated essentially all compartments of the endomembrane system. Despite the analogies between the Cvt pathway and autophagy, earlier work has suggested that the origin of the engulfing vesicle membranes is different; the endoplasmic reticulum is proposed to be required only for autophagy. In contrast, in this study we demonstrate that the endoplasmic reticulum and/or Golgi complex, but not endosomal compartments, play an important role for both yeast transport routes. Along these lines, we demonstrate that Berkeley bodies, a structure generated from the Golgi complex in sec7 cells, are immunolabeled with Atg8, a structural component of autophagosomes. Finally, we also show that none of the yeast t-SNAREs are located at the preautophagosomal structure, the presumed site of double-membrane vesicle formation. Based on our results, we propose two models for Cvt vesicle biogenesis.     

Secretory pathway quality control operating in Golgi,plasmalemmal, and endosomal systems

Exportable proteins that have significant defects in nascent polypeptide folding or subunit assembly are frequently retained in the endoplasmic reticulum and subject to endoplasmic reticulum-associated degradation by the ubiquitin-proteasome system. In addition to this, however, there is growing evidence for post-endoplasmic reticulum quality control mechanisms in which mutant or non-native exportable proteins may undergo anterograde transport to the Golgi complex and post-Golgi compartments before intracellular disposal. In some instances, these proteins may undergo retrograde transport back to the endoplasmic reticulum with re-targeting to the endoplasmic reticulum-associated degradation pathway; in other typical cases, they are targeted into the endosomal system for degradation by vacuolar/lysosomal proteases. Such quality control targeting is likely to involve recognition of features more commonly expressed in mutant proteins, but may also be expressed by wild-type proteins, especially in cells with perturbation of local environments that are essential for normal protein trafficking and stability in the secretory pathway and at the cell surface .     

Cytochemical characterization of the endomembranous system during oocyte secondary growth in Serrasalmus spilopleura (Teleostei, Characiformes, Characidae)

The endomembranous system of Serrasalmus spilopleura oocyte secondary growth was analysed using structural and ultrastructural cytochemical techniques. In vitellogenic oocytes, the endoplasmic reticulum components, the nuclear envelope intermembranous space, some Golgi dictiossomes, lysosomes, yolk granules, regions of the egg envelope and sites of the follicle cells react to acid phosphatase detection (AcPase). The cortical alveoli, some heterogeneous cytoplasmic structures, regions of the egg envelope, and sites of the follicle cells are strongly contrasted by osmium tetroxide and zinc iodide impregnation (ZIO). The endoplasmic reticulum components, some vesicles, and sites of the follicle cells also react to osmium tetroxide and potassium iodide impregnation (KI). The biosynthetic pathway of lysosomal proteins, such as acid phosphatase, required for vitellogenesis, involves the endoplasmic reticulum, Golgi complex, vesicles with inactive hydrolytic enzymes, and, finally, lysosomes. In S. spilopleura oocytes at secondary growth, the endomembranous system takes part in the production of the enzymes needed for vitellogenesis, and in the metabolism of yolk exogenous components (AcPase detection). The endomembranous system compartments also show reduction capacity (KI reaction) and are involved in the metabolism of proteins rich in SH‐groups (ZIO reaction).     

Reduction capacity of untreated and of repeatedly isoproterenol treated parotid gland of the mouse

Osmium impregnation was used to show possible differences of reduction capacity of perinuclear space, rough endoplasmic reticulum and the Golgi apparatus of unstimulated mouse parotid gland and in the gland after repeated pharmacological doses of isoproterenol. There were some significant differences between the staining of acinar and duct cells. In all intercalated and striated duct cells the staining is dense in the perinuclear space and in the rough endoplasmic reticulum. Osmiophility was not detected in the Golgi complex of intercalated duct cells. The staining was also lacking in the perinuclear space and endoplasmic reticulum of the acinar cells. The cis face of the Golgi complex and numerous transitional vesicles in the acinar cells showed variability of the reduction capacity of their membrane segments. In chronically treated acinar cells Os black was lacking in the Golgi cisternae, except that the numerous transitional vesicles were heavily stained. These results reveal characteristic differences of reduction capacity of endomembrane compartments in different parotid glandular cells, as well as between untreated and treated acinar cells.     

A proteomic analysis of lysosomal integral membrane proteins reveals the diverse composition of the organelle

Lysosomes are endocytic subcellular compartments that contribute to the degradation and recycling of cellular material. Using highly purified rat liver tritosomes (Triton WR1339-filled lysosomes) and an ion exchange chromatography/LC-tandem MS-based protein/peptide separation and identification procedure, we characterized the major integral membrane protein complement of this organelle. While many of the 215 proteins we identified have been previously associated with lysosomes and endosomes, others have been associated with the endoplasmic reticulum, Golgi, cytosol, plasma membrane, and lipid rafts. At least 20 proteins were identified as unknown cDNAs that have no orthologues of known function, and 35 proteins were identified that function in protein and vesicle trafficking. This latter group includes multiple Rab and SNARE proteins as well as ubiquitin. Defining the roles of these proteins in the lysosomal membrane will assist in elucidating novel lysosomal functions involved in cellular homeostasis and pathways that are affected in various disease processes.     

78 kDa receptor for Man6P-independent lysosomal enzyme targeting: biosynthetic transport from endoplasmic reticulum to "high-density vesicles"

Recent work has shown that the cation-independent mannose 6-phosphate and the 78 kDa receptors for lysosomal enzyme targeting are located in different cell compartments. While the mannose 6-phosphate receptor is enriched in the Percoll fractions that contain Golgi apparatus, most of the 78 kDa receptor is localized in a heavy fraction at the bottom of the Percoll gradient. This report presents the biosynthetic transport of the 78 kDa receptor. Newly synthesized 78 kDa receptor was transported to Golgi from endoplasmic reticulum with a half life of 5 min. From the Golgi apparatus, the receptor takes two routes; about 15-25% is transported to the plasma membrane, and the rest migrates to late endosomes, subsequently to prelysosomes and finally to the dense vesicles. The 78 kDa receptor starts appearing at the dense vesicles 120 min after biosynthesis and reaches a maximum of 40-50% of the total receptor. Treatment of cells with NH4Cl causes depletion of the receptor from the dense vesicles and prelysosomes and corresponding augmentation in endosomes and plasma membrane. These results suggest that the 78 kDa receptor cycles between compartments and that the dense vesicles seem to represent the most distal compartment in the biosynthetic pathway of this receptor.     

Immunoenzyme localization of cathepsins in the Golgi region of rat hepatocytes and renal tubule cells

Localization of carboxyl proteinase (cathepsin D) and cysteine proteinases (cathepsins B, H, and L) in Golgi region was studied using an immunoenzyme technique. Rat livers and kidneys were used. The results obtained from the livers were similar to those from the kidneys. All cathepsins were detected in lysosomal compartments such as secondary lysosomes, multivesicular bodies (endosomes), and autophagosomes. Rough endoplasmic reticulum (rER), including nuclear envelope was focally stained. Most of Golgi cisternae were negative, but sometimes only one cisterna or the terminal portion of the cisterna were stained focally. Rarely, the trans Golgi network (TGN) was positive for the proteinases. Among numerous Golgi vesicles, only a few of them were stained. The positive vesicles were divided into two groups, one had a bristle coat and heavily stained, and other were smaller than 40 nm in diameter and weakly stained. The small vesicles seemed to bud from the ER and to fuse with the Golgi cisternae, while the large clathrin-coated vesicles seem to bud from the TGN. The results suggests that cathepsins are transported by vesicular system from the rER to lysosomes via Golgi apparatus. In addition, it is suggested that the small vesicles transport the proteinases from the ER to the Golgi cisternae and the large clathrin-coated vesicles from the Golgi cisternae to the lysosomes.     

The amounts and rates of export of polysaccharides found within the membrane system of maize root cells

1. Maize seedling roots were incubated in vivo with d-[U-(14)C]glucose for 2, 5, 10, 15, 30 and 45min. The total incorporation of radioactivity into polysaccharide components in isolated fractions was investigated, and the pattern of incorporation into different polysaccharide components within the rough endoplasmic reticulum, Golgi apparatus and exported material was analysed. 2. The membrane compartments reached a saturation value of radioactivity in polysaccharide components by 30min incubation. Radioactivity in exported polysaccharide continued to increase after that time. The latter was formed and maintained by a steady-state turnover of polysaccharide synthesis and transport from the membrane system. 3. If the only access of the slime polysaccharide to the cell surface is via dictyosome-derived vesicles, the amount of slime components in the Golgi apparatus would have to be displaced every 0.3min in order to maintain the observed rates of increase in slime. This is in contrast with a displacement time of about 2.5min that is necessary for polysaccharide components in the Golgi apparatus to produce the observed increase in cell-wall material. The activity of the membrane system in the production of maize root slime is 8 times as great as that of the membrane system involved in cell-wall synthesis. 4. If the amount of polysaccharide material in the Golgi apparatus is maintained only by inflow of polymeric material from the rough endoplasmic reticulum the total amount of slime components in the rough endoplasmic reticulum would have to be displaced every 7min to maintain a constant amount in the Golgi apparatus. If the endoplasmic reticulum contributed directly to the cell surface in the synthesis of cell-wall material, displacement times necessary to maintain the observed rate of polymer production would be very slow.     

Immunoenzyme localization of cathepsins in the Golgi region of rat hepatocytes and renal tubule cells

Summary Localization of carboxyl proteinase (cathepsin D) and cysteine proteinases (cathepsins B, H, and L) in Golgi region was studied using an immunoenzyme technique. Rat livers and kidneys were used. The results obtained from the livers were similar to those from the kidneys. All cathepsins were detected in lysosomal compartments such as secondary lysosomes, multivesicular bodies (endosomes), and autophagosomes. Rough endoplasmic reticulum (rER), including nuclear envelope was focally stained. Most of Golgi cisternae were negative, but sometimes only one cisterna or the terminal portion of the cisterna were stained focally. Rarely, the trans Golgi network (TGN) was positive for the proteinases. Among numerous Golgi vesicles, only a few of them were stained. The positive vesicles were divided into two groups, one had a bristle coat and heavily stained, and other were smaller than 40 nm in diameter and weakly stained. The small vesicles seemed to bud from the ER and to fuse with the Golgi cisternae, while the large clathrin-coated vesicles seem to bud from the TGN. The results suggests that cathepsins are transported by vesicular system from the rER to lysosomes via Golgi apparatus. In addition, it is suggested that the small vesicles transport the proteinases from the ER to the Golgi cisternae and the large clathrin-coated vesicles from the Golgi cisternae to the lysosomes.     

Ultrastructural localization of insulin and C-peptide antigenic sites in rat pancreatic B cell obtained by applying the quantitative high-resolution protein A-gold approach

Insulin and C-peptide antigenic sites have been revealed in rat pancreatic B cells by applying immunohistochemical and cytochemical techniques. Fluorescein and rhodamine stains at the light-microscope level have detected both antigens in the same B cells. With the protein A-gold technique, labeling for both antigens was found in the cisternae of the rough endoplasmic reticulum, in those of the transitional elements, in all the cisternae of the Golgi apparatus except in the trans-most one, in the smooth but not in the coated vesicles, in the immature and mature secretory granules, and in some lysosomal (multigranular) structures. The fixation procedure used yielded excellent ultrastructural preservation which allowed for high resolution. The various control experiments demonstrated the high specificity of the results. Quantitative evaluations confirmed the qualitative observations in that they documented the specificity of the label and revealed the presence of an increasing gradient for both antigenic sites along the endoplasmic reticulum-Golgi-granule secretory pathway. The quantification also demonstrated various sites in which an increased labeling occurs: the rough endoplasmic reticulum, the smooth vesicles, the trans-cisternae of the Golgi apparatus, and the immature and the mature secretory granules. The Golgi apparatus was composed of three different subcompartments distinguished by their concentration of label. These include the cisternae on the cis-side, those on the trans-side, and the trans-most rigid cisternae. Since insulin and C-peptide form the proinsulin chain, their antigenic sites were found in the same locations along the secretory pathway; differences in location appeared only in the secretory granules, where insulin was concentrated in the core, while C-peptide was found in both the core and the halo of the granules. Furthermore, in the mature secretory granules displaying a crystalline core, insulin was restricted to the core, while C-peptide was confined to the halo. These results are in accord with the biochemical data, which indicate that simultaneous localization of both antigenic sites in compartments upstream to the immature secretory granules reflects their presence in the form of proinsulin. However, upon dissociation of proinsulin into insulin and C-peptide, both antigenic sites are segregated in different locations. The peptides appear to share parallel pathways and a fate which includes secretion through exocytosis or degradation by the lysosomal system.     

Reduction capacity of untreated and of repeatedly isoproterenol treated parotid gland of the mouse

Summary Osmium impregnation was used to show possible differences of reduction capacity of perinuclear space, rough endoplasmic reticulum and the Golgi apparatus of unstimulated mouse parotid gland and in the gland after repeated pharmacological doses of isoproterenol. There were some significant differences between the staining of acinar and duct cells. In all intercalated and striated duct cells the staining is dense in the perinuclear space and in the rough endoplasmic reticulum. Osmiophility was not detected in the Golgi complex of intercalated duct cells. The staining was also lacking in the perinuclear space and endoplasmic reticulum of the acinar cells. The cis face of the Golgi complex and numerous transitional vesicles in the acinar cells showed variability of the reduction capacity of their membrane segments. In chronically treated acinar cells Os black was lacking in the Golgi cisternae, except that the numerous transitional vesicles were heavily stained.These results reveal characteristic differences of reduction capacity of endomembrane compartments in different parotid glandular cells, as well as between untreated und treated acinar cells.     

Protein Kinase A Activity Is Necessary for Fission and Fusion of Golgi to Endoplasmic Reticulum Retrograde Tubules

It is becoming increasingly accepted that together with vesicles, tubules play a major role in the transfer of cargo between different cellular compartments. In contrast to our understanding of the molecular mechanisms of vesicular transport, little is known about tubular transport. How signal transduction molecules regulate these two modes of membrane transport processes is also poorly understood. In this study we investigated whether protein kinase A (PKA) activity regulates the retrograde, tubular transport of Golgi matrix proteins from the Golgi to the endoplasmic reticulum (ER). We found that Golgi-to-ER retrograde transport of the Golgi matrix proteins giantin, GM130, GRASP55, GRASP65, and p115 was impaired in the presence of PKA inhibitors. In addition, we unexpectedly found accumulation of tubules containing both Golgi matrix proteins and resident Golgi transmembrane proteins. These tubules were still attached to the Golgi and were highly dynamic. Our data suggest that both fission and fusion of retrograde tubules are mechanisms regulated by PKA activity.     

Traffic COPs of the early secretory pathway

Intracellular transport between the endoplasmic reticulum and Golgi compartments is mediated by coat protein complexes (COPI and COPII) that form transport vesicles and collect the desired set of cargo. Although the COPI and COPII coats are molecularly distinct, a number of mechanistic parallels appear to be emerging, most notably a general role for small guanine triphosphatases in co-ordinating coat assembly with cargo selection. A combination of morphological, biochemical, and genetic methods is revealing a very dynamic relationship between these compartments, and highlights a central role for COPs in directing traffic through the early secretory pathway. This review focuses on recent advances in molecular mechanisms underlying coated-vesicle assembly and connections with cellular structures.     

Re-assessing the locations of components of the classical vesicle-mediated trafficking machinery in transfected Plasmodium falciparum

The malaria parasite, Plasmodium falciparum, exports proteins beyond the confines of its own plasma membrane, however there is debate regarding the machinery used for these trafficking events. We have generated transgenic parasites expressing chimeric proteins and used immunofluorescence studies to determine the locations of plasmodial homologues of the COPII component, Sar1p, and the Golgi-docking protein, Bet3p. The P. falciparum Sar1p (PfSar1p) chimeras bind to the endoplasmic reticulum surface and define a network of membranes wrapped around parasite nuclei. As the parasite matures, the endomembrane systems of individual merozoites remain interconnected until very late in schizogony. Antibodies raised against plasmodial Bet3p recognise two foci of reactivity in early parasite stages that increase in number as the parasite matures. Some of the P. falciparum Bet3p (PfBet3p) compartments are juxtaposed to compartments defined by the cis Golgi marker, PfGRASP, while others are distributed through the cytoplasm. The compartments defined by the trans Golgi marker, PfRab6, are separate, suggesting that the Golgi is dispersed. Bet3p-green fluorescent protein (GFP) is partly associated with punctate structures but a substantial population diffuses freely in the parasite cytoplasm. By contrast, yeast Bet3p is very tightly associated with immobile structures. This study challenges the view that the COPII complex and the Golgi apparatus are exported into the infected erythrocyte cytoplasm.     

Protein transport in plant cells: in and out of the Golgi

In plant cells, the Golgi apparatus is the key organelle for polysaccharide and glycolipid synthesis, protein glycosylation and protein sorting towards various cellular compartments. Protein import from the endoplasmic reticulum (ER) is a highly dynamic process, and new data suggest that transport, at least of soluble proteins, occurs via bulk flow. In this Botanical Briefing, we review the latest data on ER/Golgi inter-relations and the models for transport between the two organelles. Whether vesicles are involved in this transport event or if direct ER-Golgi connections exist are questions that are open to discussion. Whereas the majority of proteins pass through the Golgi on their way to other cell destinations, either by vesicular shuttles or through maturation of cisternae from the cis- to the trans-face, a number of membrane proteins reside in the different Golgi cisternae. Experimental evidence suggests that the length of the transmembrane domain is of crucial importance for the retention of proteins within the Golgi. In non-dividing cells, protein transport out of the Golgi is either directed towards the plasma membrane/cell wall (secretion) or to the vacuolar system. The latter comprises the lytic vacuole and protein storage vacuoles. In general, transport to either of these from the Golgi depends on different sorting signals and receptors and is mediated by clathrin-coated and dense vesicles, respectively. Being at the heart of the secretory pathway, the Golgi (transiently) accommodates regulatory proteins of secretion (e.g. SNAREs and small GTPases), of which many have been cloned in plants over the last decade. In this context, we present a list of regulatory proteins, along with structural and processing proteins, that have been located to the Golgi and the 'trans-Golgi network' by microscopy.     

Isolation of a vesicular intermediate in the cell-free transfer of membrane from transitional elements of the endoplasmic reticulum to Golgi apparatus cisternae of rat liver

Preparations enriched in part-smooth (lacking ribosomes), part-rough (with ribosomes) transitional elements of the endoplasmic reticulum when incubated with ATP plus a cytosol fraction responded by the formation of blebbing profiles and approximately 60-nm vesicles. The 60-nm vesicles formed resembled closely transition vesicles in situ considered to function in the transfer of membrane materials between the endoplasmic reticulum and the Golgi apparatus. The transition elements following incubation with ATP and cytosol were resolved by preparative free-flow electrophoresis into fractions of differing electronegativity. The main fraction contained the larger vesicles of the transitional membrane elements, while a less electronegative minor shoulder fraction was enriched in the 60-nm vesicles. If the vesicles concentrated by preparative free-flow electrophoresis were from material previously radiolabeled with [3H]leucine and then added to Golgi apparatus immobilized to nitrocellulose, radioactivity was transferred to the Golgi apparatus membranes. The transfer was rapid (T1/2 of about 5 min), efficient (10-30% of the total radioactivity of the transition vesicle preparations was transferred to Golgi apparatus), and independent of added ATP but facilitated by cytosol. Transfer was specific and apparently unidirectional in that Golgi apparatus membranes were ineffective as donor membranes and endoplasmic reticulum vesicles were ineffective as recipient membranes. Using a heterologous system with transition vesicles from rat liver and Golgi apparatus isolated from guinea pig liver, coalescence of the small endoplasmic reticulum-derived vesicles with Golgi apparatus membranes was demonstrated using immunocytochemistry. Employed were polyclonal antibodies directed against the isolated rat transition vesicle preparations. When localized by immunogold procedures at the electron microscope level, regions of rat-derived vesicles were found fused with cisternae of guinea pig Golgi apparatus immobilized to nitrocellulose strips. Membrane transfer was demonstrated from experiments where transition vesicle membrane proteins were radioiodinated by the Bolton-Hunter procedure. Additionally, radiolabeled peptide bands not present initially in endoplasmic reticulum appeared following coalescence of the derived vesicles with Golgi apparatus. These bands, indicative of processing, required that both Golgi apparatus and transition vesicles be present and did not occur in incubated endoplasmic reticulum preparations or on nitrocellulose strips to which no Golgi apparatus were added.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of castration and of oestradiol treatment on the subcellular structure of the mouse seminal vesicles, with particular reference to the function of the Golgi apparatus and the lysosomes

Synopsis A number of changes were observed in the ultrastructure of seminal vesicles from castrated mice fed continuously with oestradiol. Treatment for two weeks was accompanied in the epithelial cells by the disappearance of secretion droplets, swelling of the Golgi structures and the appearance of many dense bodies and vesicles of various sizes. Subsequently, there was a decrease in the amount of rough endoplasmic reticulum followed by the disappearance of the vesicles. These changes were paralleled by a decrease in size and, finally, disappearance of the dense bodies, and by the appearance of abnormal nuclei. Ultimately, the epithelial cells became packed with free ribosomes and keratinization of the epithelium ensued.These metaplastic phenomena in the epithelium were accompanied by thickening and infolding of the basement membrane and by the formation of several layers of smooth muscle cells. The latter cells were very abnormal in that they contained prominent Golgi apparatuses and a vesiculated cytoplasm.When vesiculation occurred, both in the epithelium and in the cells of the basal areas of the acini (myoepithelial, basal and muscle cells), the vesicles, the Golgi bodies and the dense bodies (lysosomes) contained acid naphthol-AS-phosphatase activity. This enzyme was different from the more commonly described lysosomal acid -glycerophosphatase in that it was not inhibited by either sodium fluoride or sodium molybdate; in certain instances its activity in the cisternae of the endoplasmic reticulum could be shown to be enhanced by these compounds. The significance of these findings is discussed.