SULFATE METABOLISM IN PANCREATIC ACINAR CELLS

The metabolism of inorganic sulfate in pancreatic acinar cells was studied by electron microscope radioautography in mice injected with sulfate-³⁵S. Labeled sulfate was concentrated in the Golgi complex at 10 min. Within 30 min, much of the radioactive material had been transferred to condensing vacuoles. These were subsequently transformed into zymogen granules. By 4 hr after injection, some of the zymogen granules with radioactive contents were undergoing secretion, and labeled material was present in the pancreatic duct system. The Golgi complex in pancreatic acinar cells is known to be responsible for concentrating and packaging digestive enzymes delivered to it from the endoplasmic reticulum. Our work demonstrates that the Golgi complex in these cells is also engaged in the manufacture of sulfated materials, probably sulfated mucopolysaccharides, which are packaged along with the enzymes in zymogen granules and released with them into the pancreatic secretion.     

Sulfation of thyroglobulin: A ubiquitous modification in vertebrates

Summary Mammalian thyroglobulin is released by thyroid follicle cells as a sulfated glycoprotein; the sulfate residues are mostly linked to tyrosine, but they are also attached to the high-mannose carbohydrate side-chains. To decide whether sulfation of thyroglobulin is confined to mammals, representatives of other vertebrate classes were analyzed for the presence of sulfated thyroglobulin: fish (trout), amphibians (clawed toad) and birds (chicken). Mini-organs were prepared from thyroid tissue and suspended in a ³⁵SO ₄ ^-- -containing culture medium. Light- and electron-microscope autoradiographs prepared from the mini-organs showed that thyroid follicle cells from all species examined incorporate ³⁵SO ₄ ^-- and synthesize a sulfated secretory product which accumulates in the follicle lumen. The Golgi complex was detected as the primary intracellular site of sulfate organification. The ³⁵SO ₄ ^-- -radiolabeled secretory product of all species was shown by polyacrylamide-gel-electrophoretic analyses to consist of thyroglobulin, identified by comparison with biosynthetically ¹²⁵I-labeled thyroglobulin. The results indicate that the sulfation of thyroglobulin is a ubiquitous post-translational modification observed already in the thyroglobulin of lower vertebrates. Our observations suggest that sulfation of thyroglobulin was acquired in the early stages of thyroid evolution.     

Sulfated proteoglycans synthesized by Neuro 2a neuroblastoma cells: Comparison between cells with and without ganglioside-induced neurites

Mouse neuroblastoma Neuro 2a cells are known to extend neurite-like processes in response to gangliosides added to the culture medium. We compared the structural features of proteoglycans (PG) synthesized by conventional Neuro 2a cells with those of neurite-bearing cells. Two different proteoglycans labeled with [³⁵S]sulfate, namely, chondroitin sulfate proteoglycan (CS-PG) and heparan sulfate proteoglycan (HS-PG), were found both in the cell layer and in the culture medium of the conventional cells. CS-PG isolated from the cell layer had a K_av value of 0.38 on Sepharose CL-6B, and had CS side chains with Mr of 27,000. HS-PG in the cell layer was slightly larger (K_av of 0.33) in terms of hydrodynamic size than CS-PG, and the apparent Mr of the heparan sulfate side chains was 10,000. The structural parameters of CS-PG and HS-PG isolated from the medium were almost identical to those of the PGs in the cell layer. In addition to these PGs, single-chain HS, with an average Mr of 2,500, was observed only in the cell layer and this component was the major sulfated component in the cell layers of both control and ganglioside treated cells. The neurite-bearing cells also synthesized both CS-PG and HS-PG which were very similar in hydrodynamic size to those synthesized by the conventional cells, but the size of HS side chains was greater. Radioactivity, as³⁵S, of each sulfated component from the gangliosideteated culture seemed to be slightly less than that of the corresponding component from the control culture. These findings indicate that the marked morphological change in Neuro 2a cells, induced by gangliosides is not accompanied by major changes in the synthesis of PGs.     

Thymic epithelial cells synthesize a heparan sulfate with a highly sulfated region

Epithelial cells are important components of the thymus microenvironment and are involved in thymocyte differentiation. The production and secretion of sulfated glycosaminoglycans by these cells grown in culture were investigated using labeling with radioactive ³⁵S-Na₂SO₄ and ³H-glucosamine. The major glycosaminoglycans synthesized by these cells are heparan sulfate and hyaluronic acid. The structure of the heparan sulfate was investigated by the pattern of degradation products formed by deaminative cleavage with nitrous acid. The ratio ³⁵S-sulfate/³H-glucosamine is high in the segments of the heparan sulfate released during the deaminative cleavage with nitrous acid but low in the resistant portion of the molecule. Thus, the heparan sulfate synthesized by the thymic epithelial cells contains a highly sulfated region. Digestion with heparitinase reveals that this highly sulfated region is a heparin-like segment of the molecule. The heparan sulfate is rapidly incorporated into the cell surface but its secretion to the extracellular medium requires a longer incubation period. Finally, heparin was used to mimic the possible effect of this heparan sulfate with a highly sulfated region, as ascertained by its ability to modulate thymocyte adhesion to thymic epithelial cells. Since heparin actually enhanced thymocyte adhesion, it is suggested that the heparan sulfate described herein, secreted by the thymic epithelium, may play a role upon intrathymic heterotypic cellular interactions. J Cell Physiol 178:51–62, 1999. © 1999 Wiley-Liss, Inc.     

In vivo sulfation of the contact site A glycoprotein of Dictyostelium discoideum

During the development of Dictyostelium discoideum from the growth phase to the aggregation stage, a glycoprotein with an apparent mol. wt. of 80 kd is known to be expressed on the cell surface. This glycoprotein, referred to as contact site A, has been implicated in the formation of species-specific, EDTA-stable contacts of aggregating cells. When developing cells were labeled in vivo with [³⁵S]sulfate, the 80-kd glycoprotein was found to be the most prominently sulfated protein. Another strongly sulfated protein had an apparent mol. wt. of 130 kd and was, like the 80-kd glycoprotein, developmentally regulated and associated with the particulate fraction of the cells. The [³⁵S]sulfate incorporated into the 80-kd and 130-kd proteins was not present as tyrosine-O-sulfate, a modified amino acid found in many proteins of mammalian cells. D. discoideum cells incubated with [³⁵S]sulfate in the presence of tunicamycin, an inhibitor of N-glycosylation, produced a 66-kd protein that reacted with monoclonal antibodies raised against the 80-kd glycoprotein, but no longer contained [³⁵S]sulfate. These results suggest that sulfation of the 80-kd glycoprotein occurred on carbohydrate residues. The possible importance of sulfation for a role of the 80-kd glycoprotein in cell adhesion is discussed.     

Expression of Sulfated Gangliosides in the Central Nervous System

Abstract: Several sulfated lipids were detected in the ganglioside fraction isolated from a cell line of oligodendrocyte progenitors that had been metabolically labeled with [³⁵S]sulfate. Separation of the ganglioside fraction by two-dimensional TLC showed that, except for galactosylceramide-sulfate, none of the sulfate-labeled lipids comigrated with those glycosphingolipids visualized by orcinol staining, indicating that these sulfolipids were quantitatively minor components. At least eight sulfate-labeled lipid bands were susceptible to desialylation by Arthrobacter ureafaciens neuraminidase, which resulted in the formation of three new bands that retained the labeled sulfate. Six of the sulfate-labeled lipid bands containing sialic acid were also susceptible to Vibrio cholerae neuraminidase, which generated two labeled bands that appeared identical to the two major products formed after treatment with A. ureafaciens neuraminidase. In vivo labeling of lipids from 14-day-old rat brain with [³⁵S]sulfate demonstrated that the synthesis of sulfated lipids containing sialic acid also occurred in intact brain tissue. These results show that sulfated gangliosides are synthesized in the CNS and that oligodendrocytes are one cell type that contributes to this synthesis.     

Incorporation of [S]Sulfate and [C]Bicarbonate into Karyotype-specific Polysaccharides of Chondrus crispus

Gametophytic and sporophytic tissues of Chondrus crispus (Stackhouse) cultured in vitro were labeled with ³⁵S and ¹⁴C. The major sulfated polysccharides isolated from the two karyotypes were characterized by KCI fractionation, immunoprecipitation, and infrared spectroscopy. Reproducibility of data has been demonstrated by an experiment using gametophytic T₄ strain with five replicates per time point. The rate of sulfate uptake was similar in haploid and diploid plants from a given area cultured for a similar time. Cultures from different sources cultured for different times showed different uptake and incorporation levels. Although sulfate uptake did not appear to be karyotype-related, the pattern of incorporation of ³⁵S and ¹⁴C into polysaccharides was ploidy-specific.     

Secretion of 35SO4-labeled proteins from isolated rat hepatocytes

Sulfation is a Golgi-specific modification of secretory proteins. We have characterized the proteins that are labeled with 35SO4 in cultures of rat hepatocytes and studied their transport to the medium. Analysis by polyacrylamide gel electrophoresis showed that of the five most heavily labeled proteins, four had well-defined mobilities--apparent molecular masses of 188, 142, 125, and 82 kDa--whereas one was electrophoretically heterogeneous--apparent molecular mass of 35-45 kDa. Judging by their relatively high resistance to acid treatment, the sulfate residues in the 125- and 35-45-kDa proteins were linked to carbohydrate. Some of the secreted proteins were sialylated. In samples of pulse-labeled cells, there appeared to be no unsialylated forms, indicating that sulfation occurred after sialylation, presumably in the trans Golgi. Kinetic experiments showed that the cellular half-life was the same for all the sulfated proteins--about 8 min--consistent with the idea that transport from the Golgi complex to the cell surface occurs by liquid bulk flow.     

Sulfate restriction induces hyposecretion of the adhesion proteoglycan and cell hypomotility associated with increased 35SO42− uptake and expression of a band 3 like protein in the marine sponge,Microciona prolifera

Sulfate is an important component relating to normal proteoglycan secretion and normal motility in the marine sponge, Microciona prolifera. The following alterations were observed in sponge cells when sulfate free artificial sea water was used as the suspension medium: (1) impairment of aggregation, (2) loss of cell movements, (3) a marked reduction in the secretion of the adhesion proteoglycan (AP). Reversal of this effect occurred if sulfate depleted cells were again rotated in sulfate containing artificial sea water. Motility and reaggregation of sulfate deprived cells could be completely restored by purified AP, but only if cells were first pre-conditioned in normal sea water. Comparisons of ³⁵SO₄^2? uptake between normal and sulfate deprived cells which had been treated to reduce preformed secretions showed a marked increase in ³⁵SO₄^2? uptake and incorporation which could be greatly augmented in the presence of Ca²⁺/Mg²⁺. Excessive retention of AP in sulfate starved cells demonstrated by immunostaining suggested that AP secretion and cellular motility may be controlled by a sulfate dependent secretogogue or that undersulfated AP itself had developed a secretory defect. SDS-PAGE of Triton treated cellular extracts demonstrated a 116 kDa ³⁵SO₄^2? sulfated band which co-migrated with AP, but only in extracts derived from sulfate starved cells. Western blots prepared from such extracts incubated in the presence of a monoclonal anti-band 3 antibody demonstrated labelling of a single 97 kDa band only in material from sulfate deprived cells. The absence of this component in normal cell extracts indicated that this protein may be involved in facilitated sulfate transport. This study lends support to a heretofore unrecognized role for sulfate in cell motility and secretion.     

Intermediates in the constitutive and regulated secretory pathways released in vitro from semi-intact cells

Regulated secretory cells have two pathways that transport secreted proteins from the Golgi complex to the cell surface. To identify carrier vesicles involved in regulated and constitutive secretion, PC12 pheochromocytoma cells were labeled with [35S]sulfate to identify markers for the two secretory pathways, then mechanically permeabilized and incubated in vitro. Small constitutive secretory vesicles, containing mostly sulfated proteoglycans, accumulated during an in vitro incubation with ATP. In the presence of GTP gamma S, the constitutive vesicles became significantly more dense, suggesting that a coated intermediate was stabilized. Larger immature regulated secretory granules, enriched in sulfated secretogranin II, also escaped from the permeabilized cells in vitro. During granule maturation, their density increased and the amount of cofractionating proteoglycans diminished. The data suggest that sorting continues during secretory granule maturation.     

Characterization of the major sulfated protein of mouse pancreatic acinar cells: A high molecular weight peripheral membrane glycoprotein of zymogen granules

The major sulfated protein of the mouse pancreatic acinar cell, gp300, hsa been identified and characterized with monoclonal and polyclonal antibidies. gp300 is a glycoprotein of M_r = 300,000 which contains ～40% of metabolically incroporated [³⁵S] sulfate in the acinar cell. Sulfate on gp300 is resistant to hot 1N HCl, but sensitive to alkaline hydrolysis. demonstrating that the sulfate is carbohydrate-linked rather than tyrosine-linked. gp300 metabolically labeled with [^3H]glucosamine and [³⁵H]sulfate was chemically and enzymaticlly treated followed by Bio-Gel P-10 gel filtration. Both labels were resistant to treatments which degrade glycosaminoglycan. Treatment of dual-labeled gp300 with PNGase F to cleave N-linked oligosaccharides released ～17% of [₃S]. Mild alkaline borohydride treatment after removal of N-linked sugar relased the remainder of both labels, indicating the presence of sulfated O-linked oligosaccharides. Biosynthetic studies and PNGase F digestion indicating the presence of sulfated O-linked oligosaccharides. Biosunynthetic studies and PNGase digestion F digestion indicate that the core protein is ～210 KDa, with apparent contrinution of ～35 KDa N-linked sugar, and ～55 KDa O-linked sugar. Lectin blotting and glycosidase digestion demonstrated the presece of Galβ(1–3)GalNAc and sialic acid α(2–3)Gal in O-linked oligosaccharide, and Galβ(1–4)GLcNAc in N-linked oligosaccharide. Immunolocalization and subcellular fractionation showed that gp300 is a peripheral memberane protein localized to the lumenal face of the zymogen granule membrane. gp300 was not secreted in reponse to hormone stimulation ofacini, so it is not a secertroy product. Immunoblot analysis showed that gp300 is present in other gastrointestinal tissues and parotid glands. Localization of this nonsecreted sulfated glycoprotein to exocrine secretory granule membranes suggests that gp300 may have a role in granule bigeneses.     

MUCOPOLYSACCHARIDES SYNTHESIZED BY CULTURED GLIAL CELLS DERIVED FROM A PATIENT WITH SANFILIPPO A SYNDROME

—Glial cells were cultured from brain tissue obtained at autopsy of a patient with Sanfilippo A syndrome. Mucopolysaccharides were labeled by culturing the cells in the presence of [³⁵S]sulfate. After proteolysis, intracellular and media-elaborated mucopolysaccharides were fractionated by Dowex 1 chromatography. One fraction, identified as heparan sulfate by chromatographic, electrophoretic, and enzyme susceptibility properties, accumulated in Sanfilippo glial cells in greater amounts than in controls. Heparan sulfate was also excreted into the culture media by both Sanfilippo and normal cultures, and it constituted a major fraction of the sulfated mucopolysaccharides synthesized by glial cells. Sanfilippo and normal fibroblasts were also included in these studies for comparative purposes. Sanfilippo fibroblasts accumulated significantly increased amounts of heparan sulfate as compared to normal fibroblasts. Heparan sulfate was excreted into the culture media by Sanfilippo and normal fibroblasts in equivalent amounts, but in contrast to glial cells, it was only a minor component of the sulfated mucopolysaccharides produced. Cultured glial cells should provide a useful system for investigating the role of heparan sulfate in glial cell function.     

Production of sulfated proteoglycans by human breast cancer cell lines: Binding to fibroblast growth factor-2

The cellular distribution and nature of proteoglycans synthesised by human breast cancer cells in culture were studied. Proteoglycans were labelled with [³⁵S] sulfate, purified, and characterised after ion-exchange chromatography followed by gel-filtration chromatography and treatment with glycosaminoglycan degrading enzymes. Proteoglycans were isolated from the culture medium and from cell layers of the hormono-dependent well-differentiated MCF-7 cell line, the hormono-independent poorly-differentiated MDA-MB-231 and the HBL-100 cell line which is derived from non malignant breast epithelium. HBL-100 and MDA-MB-231 cells produced larger amounts of proteoglycans which had a lower degree of sulfation than MCF-7 cells. Gel-filtration chromatography on Sepharose CL-6B indicated that HBL-100 and MDA-MB-231 cells accumulated cell surface heparan sulfate proteoglycans (HSPG), with a high apparent molecular weight (K_av 0.1). In contrast, the MCF-7 cell monolayers synthesised small sulfated macromolecules (K_av 0.4) which possessed mostly chondroitin sulfate chains. Moreover, considerable differences in the nature of the sulfated proteoglycans released into the culture medium of these breast epithelial cell lines were observed. MCF-7 cells released into the culture medium HSPG as the main proteoglycan component while MDA-MB-231 and HBL-100 cells released mainly chondroitin sulfate proteoglycans. In these three cell lines, medium-released sulfated macromolecules have a higher hydrodynamic size than cell-associated ones. Proteoglycans purified by ion-exchange chromatography were tested for their ability to bind ¹²⁵I FGF-2. We demonstrated that HBL-100 and MDA-MB-231 cells bind more FGF-2 to their heparan sulfate proteoglycans than MCF-7 cells. Taken together, these results suggest that differences in proteoglycan synthesis of human breast epithelial cells could be responsible for differences in their proliferative and/or invasive properties. J. Cell. Biochem. 64:605–617. © 1997 Wiley-Liss, Inc.     

Sulfation of fucoidan in Fucus embryos. I. Possible role in localization

Zygotes of the brown alga Fucus distichus L. Powell divide into two cells which are structurally and biochemically different from each other. Cytochemical staining and autoradiography indicate that a sulfated polysaccharide is localized in only one of the two cells. Up to 10 hr after fertilization, no localization of sulfated polysaccharides is detectable in zygotes, and little ³⁵S (Na₂³⁵SO₄) is incorporated into an acid-soluble carbohydrate fraction. Between 10 and 16 hr, during rhizoid initiation and several hours before the first cell division, there is a large increase in the amount of ³⁵S incorporated into this fraction. The label is found associated with the sulfated fucose polymer fucoidan. Various extraction techniques and labeling experiments demonstrate that fucoidan is unsulfated at fertilization and undergoes little metabolic activity or turnover during the first 24 hr. Thus, the incorporation of sulfate into this carbohydrate fraction appears to involve a sulfation of a preexisting, unsulfated fucan polymer. The degree of sulfation achieved at this time in vivo is sufficient for migration of fucoidan through an electric field in agarose or acrylamide gels. The possible role of sulfation as a mechanism for the localization of fucoidan in the rhizoid cell by means of an intracellular electrical gradient is discussed.     

Axonal transport of proteoglycans to the goldfish optic tectum

The study addressed the question of whether³⁵SO₄ labeled molecules that the have been delivered to the goldfish optic nerve terminals by rapid axonal transport include soluble proteoglycans. For analysis, tectal homogenates were subfractionated into a souluble fraction (soluble after centrifugation at 105,000g), a lysis fraction (soluble after treatment with hypotonic buffer followed by centrifugation at 105,000g) and a final 105,000g pellet fraction. The soluble fraction contained 25.7% of incorporated radioactivity and upon DEAE chromatographys was resolved into a fraction of sulfated glycoproteins eluting at 0–0.32 M NaCl and containing 39.5% of total soluble label and a fraction eluting at 0.32–0.60 M NaCl containing 53.9% of soluble label. This latter fraction was included on columns of Sepharose CL-6B with or without 4 M guanidine and after pronase digestion was found to have 51% of its radioactivity contained in the glycosaminoglycans (GAGs) heparan sulfate and chondroitin (4 or 6) sulfate in the ratio of 70% to 30%. Mobility of both intact proteoglycans and constituent GAGs on Sepharose CL-6B indicated a size distribution that is smaller than has been observed for proteoglycans and GAGs from cultured neuronal cell lines. Similar analysis of lysis fraction, containing 11.5% of incorporated³⁵SO₄, showed a mixture of heparan sulfate and chondroitin sulfate containing proteoglycans, apparent free heparan sulfate and few, if any, sulfated glycoproteins. Overall, the result support the hypothesis that soluble proteoglycans are among the molecules axonally transported in the visual system.     

Sulfhydrolase Activity in Sediments of Wintergreen Lake, Kalamazoo County, Michigan

The hydrolysis of p-nitrophenyl sulfate, p-nitrocatechol sulfate, and [³⁵S]sodium dodecyl sulfate was examined in anoxic sediments of Wintergreen Lake, Michigan. Significant levels of sulfhydrolase activity were observed in littoral, transition, and profundal sediment samples. Rates of sulfate formation suggest that the sulfhydrolase system would represent a major source of sulfate within these sediments. Sulfate formed by ester sulfate hydrolysis can support dissimilatory sulfate reduction as shown by the incorporation of ³⁵S from labeled sodium dodecyl sulfate into H₂³⁵S. Sulfhydrolase activity varied with sediment depth, was greatest in the littoral zone, and was sensitive to the presence of oxygen. Estimations of ester sulfate concentrations in sediments revealed large quantities of ester sulfate (~30% of total sulfur). Both total sulfur and ester sulfate concentrations varied with the sediment type and were two to three orders of magnitude greater than the inorganic sulfur concentration.     

Biosynthesis of chondroitin sulfate side chains and hyaluronate time-course studies with cartilage slices of calf ribs under different conditions

The time course of double labeling with ³⁵SO₄²⁻ and [³H]glucosamine was followed in a semi-in vitro system of cartilage slices from calf ribs whose chondroitin sulfate peptide pool consistsof (A) <1% of very short undersulfated side chains of <10 disaccharide units length, (B) 3–5% of short undersulfated longer side chains (16 to 25 disaccharide units), (C) 3–5% of short, slightly oversulfated side chains (16–23 dissacharide units, very probably containing some dermatan sulfate), (D) the bulk material (74–82% of total uronate) of longest, slightly undersulfated or equally sulfated side chains (22–42 disaccharide units).After 10 min incubation rapid chain elongation with [³H]glucosamine and prelabeling with ³⁵SO₄²⁻ of endogenous acceptors are apparent. Chains of type A exhibit highest specific radioactivities. During 30–60 min incubation it is mainly chains of type B that show highest specific radioactivities, after 90 min chains of type C. On the after hand, chains of type D always incorporated the highest total amount of both precursors. Preincubation of slices for 40 min at 37°C strongly enhances labeling rates of all types whilst preincubation for 40 min in an ice-bath enhances mainly ³⁵SO₄²⁻ labeling of types A and B.After 10 min preincubation followed by ³⁵SO₄²⁻ labeling for 60 min a decrease of radioactivity of type A and a distinct increase with type B are observed during the post incubation period. After pulse chase experiment type B exhibit highest specific radioactivities. The data make it evident that under-sulfated short chondroitin sulfate side chains from very rapidly in a well organised manner and grow, by elongation and proceeding sulfation processes, to longer higher sulfated chains.The labeling of the hyaluronate pool is about half of that of the chondroitin sulfate pool after a lag phase of 10 min. The latter increases linearly after 35–45 min incubation time. However, after preincubation and chase experiments the hyaluronate pool is more highly labeled. The data indicate different precursor pools of both biosynthesis mechanisms, probably located in different cell compartments and/or different cartilage cells.     

Distribution of inorganic sulfate between plasma, liver, and bile

The distribution of ³⁵SO₄⁼ between inorganic and other forms in plasma, liver and bile of rats was studied by use of paper electrophoresis. The overall liver sulfate space varied from 50–100% but when determined for inorganic ³⁵SO₄⁼ was constant at 34%. This is considerably higher than that expected for passive distribution and suggests SO₄⁼ is actively transported into liver cells. Overall bile/plasma ratios were always greater than 1 while the ratio for inorganic sulfate was 0.44, a value consistent with the known bile/plasma electrical potential.     

Effect of stress on sulfated glycol, metabolites of brain norepinephrine.

The present study examined the effect of footshock stress on the formation of the two major metabolites of rat brain norepinephrine (NE) - the sulfate conjugates of 3-methoxy-4-hydroxyphenylglycol (MOPEG-SO₄) and 3,4-dihydroxyphenylglycol (DOPEG-SO₄). Rats receiving intraventricular injections of either ³HNE or Na₂³⁵SO₄ prior to 0.5 hour of footshock showed significant and comparable increases in both sulfated glycols labeled with ³H or ³⁵SO₄. Elevations were greatest in the hypothalumus using Na₂³⁵SO₄. In pheniprazine pretreated rats footshock did not increase the production of MOPEG-³⁵SO₄ from intraventricular labeled sulfate given alone or in combination with various doses of exogenous MOPEG. The results indicate that neuronally released brain NE is metabolized to form both MOPEG-SO₄ and DOPEG-SO₄. The increase in these metabolites results from an increased glycol production and not from a stress-induced activation of brain sulfation mechanisms.     

ON THE SITE OF SULFATION IN COLONIC GOBLET CELLS

The location of bound S³⁵ in the goblet cell of the rat colon at time points from 2 to 60 minutes after administration of S³⁵ as sodium sulfate has been observed in vivo and in vitro by radioautographic techniques. Grains were first observed by electron microscopy over the stacked lamellae of the paranuclear part of the Golgi apparatus. The label was subsequently found associated with the supranuclear Golgi lamellae and was then seen associated with the smooth membranes limiting the mucin granules in the goblet. Finally, between ½ and 1 hour, the secreted mucus product in the crypts became radioactive. Neither mitochondria nor the endoplasmic reticulum was labeled. It is concluded that the Golgi apparatus is the organelle in which sulfation occurs.