Electrophoretic map of boar sperm plasma membrane polypeptides and localization and fractionation of specific polypeptide subclasses

A high resolution, two-dimensional (2-D) electrophoretic map of the plasma membrane (PM) polypeptides from the ejaculated boar spermatozoon is described. 2-D silver-stained polyacrylamide gel electrophoresis (PAGE) gels revealed over 250 polypeptides; Coomassie blue staining revealed more than 100. Fifty Coomassie-staining polypeptides were catalogued and biochemically characterized, with twenty of these designated major sperm PM polypeptides. Cavitation pressures ranging from 50 PSI to 1000 PSI were used to enrich 2-D maps either in head PM (50 PSI) or in flagellar PM (1000 PSI) and provided tentative localization of certain PM polypeptides. Immunoabsorption chromatography showed that most major polypeptides seen in the 2-D map were antigenic. Major polypeptide bands from single dimensional (1-D) gels were excised, antibodies against them were produced in rabbits, and the polypeptides were localized via indirect fluorescein isothiocyanate (FITC) technique. Cross-reacting antigenic determinants in the PAGE PM polypeptides were determined by transblotting and staining the transblots by an indirect peroxidase technique. Cross-reactivity was extensive with several polypeptide groups, but specific enough with others to allow tentative localization. Lectin affinity chromatography using Con A, WGA, RCA-1, PNA, and DBA indicated the lectin specificity of PM polypeptides. These data together with periodic acid-Schiff (PAS) and carbohydrate-specific silver staining permitted identification of glycoproteins in the 2-D maps. FITC coupled to specific lectins showed the regional distribution of these lectins on the sperm surface. The 2-D polypeptide map and the catalogue of properties of major Coomassie-stained PM polypeptides provides a reference for future studies in the boar and other species.     

Cloned DNA fragment specifying major outer membrane protein a in Escherichia coli K-12.

Plasmid pMC44 is a recombinant plasmid that contains a 2-megadalton EcoRI fragment of Escherichia coli K-12 DNA joined to the cloning vehicle, pSC101. The polypeptides specified by plasmid pMC44 were identified and compared with those specified by pSC101 to determine those that are unique to pMC44. Three polypeptides specified by plasmid pMC44 were localized in the cell envelope fraction of minicells: a Sarkosyl-insoluble outer membrane polypeptide (designated M2), specified by the cloned 2-megadalton DNA fragment, and two Sarkosyl-soluble membrane polypeptides specified by the cloning plasmid pSC101. Bacteria containing plasmid pMC44 synthesized quantities of M2 approximately equal to the most abundant E. coli K-12 outer membrane protein. Evidence is presented that outer membrane polypeptide M2, specified by the recombinant plasmid pMC44, is the normal E. coli outer membrane protein designated protein a by Lugtenberg and 3b by Schnaitman.     

Polypeptides of the chloroplast envelope membranes as visualized by immunochemical techniques

The polypeptides of relative molecular masses (Mr) 22,000, 29,000, and 36,000 represent three major constituents of the chloroplast envelope of spinach (Spinacia oleracea L.) leaves. The Mr 22,000 polypeptide has been localized in the outer membrane, whereas the two other peptides have been attributed to the inner envelope membrane (Joyard et al., 1983). The Mr 29,000 polypeptide has been identified as the "phosphate translocator" (Flügge and Heldt, 1979). In this investigation, we studied the three envelope polypeptides by means of immunocytochemistry. Using indirect immunofluorescence, all three polypeptides were visualized in cryostat sections of formaldehyde-fixed leaf tissue. They were found in both palisade and spongy parenchyma cells and in guard cells, as indicated by a strong fluorescence in the chloroplast periphery. In contrast, fluorescein isothiocyanate or protein A-gold labeling of isolated fixed chloroplasts resulted only in visualization of the Mr 22,000 polypeptide, a constituent of the outer membrane. We further studied the morphological distribution and frequency of this peptide by electron microscopic evaluation of platinum-carbon replicas after freeze-etching or label-fracture and of ultra-thin sections. By use of these three methods, the polypeptide was found to be randomly distributed in the outer envelope membrane and easily accessible to the immunomarker. Average marker density, as obtained by freeze-etching and label-fracture, was approximately 130 gold particles per square micron.     

Identification and partial characterization of a novel bipartite protein antigen associated with the outer membrane of Escherichia coli.

A study by crossed immunoelectrophoresis performed in conjunction with precipitate excision and polypeptide analysis identified a new antigen complex in the envelope of Escherichia coli ML308-225. This antigen corresponds to antigen 43 in the crossed immunoelectrophoresis profile of membrane vesicles (P. Owen and H. R. Kaback, Proc. Natl. Acad. Sci. USA 75:3148-3152, 1978). Immunoprecipitation experiments conducted with specific antiserum revealed that the complex was expressed on the cell surface and that it contained, in equal stoichiometry, two chemically distinct polypeptides termed alpha and beta (Mrs of 60,000 and 53,000, respectively). The beta polypeptide was heat modifiable, displaying an apparent Mr of 37,000 when solubilized at temperatures below 70 degrees C. Analysis of fractions obtained following cell disruption, isopycnic centrifugation, and detergent extraction indicated that both alpha and beta polypeptides were components of the outer membrane. The two polypeptides were not linked by disulfide bonds, and neither was peptidoglycan associated. The complex contained no detectable lipopolysaccharide, enzyme activity, fatty acyl groups, or other cofactors. Neither correlated with E. coli proteins of similar molecular weight which had previously been shown to be associated with the outer membrane. Antibodies were raised to individual alpha and beta polypeptides. Each of these sera was shown to be subunit specific when tested against denatured membrane proteins. In contrast, each immunoglobulin preparation coprecipitated both alpha and beta polypeptides when tested against undenatured proteins derived from Triton X-100-treated membranes. The results reveal the presence of a novel bipartite protein antigen in the outer membrane of E. coli.     

Subcellular distribution of the 1,4-dihydropyridine receptor in rabbit skeletal muscle in situ: an immunofluorescence and immunocolloidal gold- labeling study

The subcellular distribution of the 1,4-dihydropyridine receptor was determined in rabbit skeletal muscle in situ by immunofluorescence and immunoelectron microscopy. Longitudinal and transverse cryosections (5-8 microns) of rabbit gracilis muscle were labeled with monoclonal antibodies specific against either the alpha 1-subunit (170,000-D polypeptide) or the beta-subunit (52,000-D polypeptide) of the 1,4-dihydropyridine receptor by immunofluorescence labeling. In longitudinal sections, specific labeling was present only near the interface between the A- and I-band regions of the sarcomeres. In transverse sections, specific labeling showed a hexagonal staining pattern within each myofiber however, the relative staining intensity of the type II (fast) fibers was judged to be three- to fourfold higher than that of the type I (slow) fibers. Specific immunofluorescence labeling of the sarcolemma was not observed in either longitudinal or transverse sections. These results are consistent with the idea that the alpha 1-subunit and the beta-subunit of the purified 1,4-dihydropyridine receptor are densely distributed in the transverse tubular membrane. Immunoelectron microscopical localization with a monoclonal antibody to the alpha 1-subunit of the 1,4-dihydropyridine receptor showed that the 1,4-dihydropyridine receptor is densely distributed in the transverse tubular membrane. Approximately half of these were distributed in close proximity to the junctional region between the transverse tubules and the terminal cisternae. Specific labeling was also present in discrete foci in the subsarcolemmal region of the myofibers. The size and the nonrandom distribution of these foci in the subsarcolemmal region support the possibility that they correspond to invaginations from the sarcolemma called caveolae. In conclusion, our results demonstrate that the 1,4-dihydropyridine receptor in skeletal muscle is localized to the transverse tubular membrane and discrete foci in the subsarcolemmal region, possibly caveolae but absent from the lateral portion of the sarcolemma.     

Secretory protein biosynthesis in snail hemocytes: in vitro modulation by larval schistosome excretory-secretory products

Circulating hemocytes of the snail, Biomphalaria glabrata, synthesize and secrete a variety of polypeptides when maintained in vitro in serum-free medium containing [35S] methionine. SDS-PAGE/fluorographic analysis of supernatants from resistant snail (10-R2-OK strain) hemocyte cultures revealed the presence of numerous labeled polypeptides ranging in Mr from 220 to 14 kDa. Most of these same proteins were also produced by hemocytes of a susceptible B. glabrata strain (M-line), but the overall rate of secretory protein synthesis was reduced from that of resistant snail cells. In addition, excretory-secretory (ES) products contained in supernatants from Schistosoma mansoni miracidial transformation and 1-day primary sporocyst cultures stimulated increases in the synthesis of various polypeptides. Particularly striking was a 3-fold increase in the synthesis of a 66-kDa secretory polypeptide by hemocytes of both snail strains, and a concomitant increase in M-line hemocytes and decrease in 10-R2-OK cells of a 63-kDa polypeptide. Overall, however, the level of ES product-induced secretory protein synthesis was greater in 10-R2-OK snail hemocytes than in those of the M-line strain. Exposure of a nonhemocytic B. glabrata cell line to parasite culture supernatants had no stimulatory/inhibitory effect on labeled protein ouput, suggesting that the observed hemocyte response may be snail cell-type specific. Finally, the larval ES components responsible for modulating hemocyte protein metabolism are mainly concentrated in a heat-stable fraction composed of molecules of greater than 30 kDa. However, the loss of the ability of heated parasite products to stimulate synthesis of certain hemocyte proteins and the presence of minor stimulating activity in a low molecular weight fraction (less than 10 kDa) implies the possible existence of multiple larval components affecting formation of specific hemocyte secretory polypeptides. It is concluded that snail hemocytes are capable of in vitro synthesis and secretion of a variety of methionine-containing polypeptides, and that ES products of early larval schistosomes can modulate (i.e., stimulate or inhibit) this metabolic process. A differential response of susceptible vs. resistant hemocytes to larval products suggests that the degree to which these cells can be metabolically activated may determine their cytotoxic effectiveness.     

Molecular dissection of radixin: distinct and interdependent functions of the amino- and carboxy-terminal domains

The ERM proteins--ezrin, radixin, and moesin--occur in particular cortical cytoskeletal structures. Several lines of evidence suggest that they interact with both cytoskeletal elements and plasma membrane components. Here we described the properties of full-length and truncated radixin polypeptides expressed in transfected cells. In stable transfectants, exogenous full-length radixin behaves much like endogenous ERM proteins, localizing to the same cortical structures. However, the presence of full-length radixin or its carboxy-terminal domain in cortical structures correlates with greatly diminished staining of endogenous moesin in those structures, suggesting that radixin and moesin compete for a limiting factor required for normal associations in the cell. The results also reveal distinct roles for the amino- and carboxy-terminal domains. At low levels relative to endogenous radixin, the carboxy-terminal polypeptide is associated with most of the correct cortical targets except cleavage furrows. In contrast, the amino-terminal polypeptide is diffusely localized throughout the cell. Low level expression of full-length radixin or either of the truncated polypeptides has no detectable effect on cell physiology. However, high level expression of the carboxy-terminal domain dramatically disrupts normal cytoskeletal structures and functions. At these high levels, the amino-terminal polypeptide does localize to cortical structures, but does not affect the cells. We conclude that the behavior of radixin in cells depends upon activities contributed by separate domains of the protein, but also requires modulating interactions between those domains.     

Antigen activation of murine T cells induces tyrosine phosphorylation of a polypeptide associated with the T cell antigen receptor

The antigen receptor complex on murine MHC class II-restricted T cells consists of disulfide-linked alpha and beta chains noncovalently associated with four additional polypeptides, two that are endoglycosaminidase F-sensitive, gp26 and gp21, and two that are endoglycosaminidase F-resistant, p25 and p16. We demonstrate here that treatment of murine T cell hybridomas with phorbol 12-myristate 13-acetate results in phosphorylation of p25 and gp21 on serine residues. However, activation of cells by antigen results in the phosphorylation of the gp21 chain and a heretofore unidentified 21 kd protein. This newly defined polypeptide, p21, is specifically immunoprecipitated with the antigen receptor complex, is endoglycosaminidase F-resistant, and is itself part of a disulfide-linked molecule. Unlike antigen-induced phosphorylation of gp21, which occurs on serine residues, phosphorylation of p21 occurs uniquely on tyrosine residues.     

Central nervous system antigen P84 can serve as a substrate for neurite outgrowth

Neurite outgrowth promoting properties of neural cell surface proteins can be assessed by immobilizing isolated membrane proteins on nitrocellulose-coated petri dishes. Using this method, we have identified a unique cell surface antigen, designated P84, as a new neural cell adhesion molecule. Immunoaffinity purified P84 contains three polypeptides with molecular weights of 167, 85, and 66 kDa. When spotted onto nitrocellulose-coated plates, P84 supports adhesion of mouse cerebellar neurons and neurite outgrowth. Glial cell attachment was also observed. Intact monoclonal antibodies directed against P84 inhibit adhesion and outgrowth on a P84 substrate. This antigen is found on the surfaces of neurons in cultures of cerebellar cells. It is also found on a subclass of unidentified flat cells. P84 is not found on oligodendrocytes or GFAP-positive astrocytes. As early as E9, P84 could be detected in the floor plate region of the spinal cord. This pattern persists throughout embryonic development. Postnatally, widespread expression of P84 is observed in a variety of CNS tissues.     

Comparison of two cell surface molecules involved in neural cell adhesion

Two cell surface molecules found in mouse brain, N-CAM and the L1 antigen, were compared in terms of their cell adhesion function, polypeptide structures, antigenic determinants and distribution in cerebellar tissue. Fab fragments of polyclonal antibodies to either N-CAM or L1 antigen only partially inhibited the rate of calcium-independent aggregation of neuroblastoma N2A cells, whereas complete and more efficient inhibition was obtained when they were used in combination. Despite the functional similarity, comparison of the electrophoretic behaviour of the purified molecules and of their proteolytic fragments shows that the L1 antigen polypeptide is distinct from that of N-CAM. In addition, no antigenic cross-reactivity was detected between the two molecules. In cryostat sections of cerebellum from young post-natal mice, N-CAM was found to be present in all cell and neurite layers, whereas L1 antigen was expressed only in regions containing post-mitotic cells. These results indicate that two chemically and histochemically distinct cell surface polypeptides can contribute to the calcium-independent adhesiveness of neural cells, and suggest that their differential expression might cause adhesive specificity among cells of developing neural tissues.     

A novel adenylylation process in liver plasma membrane-bound proteins

Rat liver plasma membrane contains five distinct polypeptides of apparent molecular mass of 130, 120, 110, 100, and 86 kDa which are labeled upon incubation with [alpha-32P]ATP as well as with [gamma-32P]ATP. Covalently bound adenosine 5'-monophosphate to some of the polypeptides was identified using nonhydrolyzable analogues of ATP. Chase experiments of alpha-32P-nucleotide-labeled polypeptides with different nonradiolabeled phosphocompounds and sensitivity to different inhibitors demonstrate that the 86-kDa polypeptide is a phosphoesterase, forming a catalytic intermediate. On the other hand, the comparative slow rate of turnover of the polypeptides of higher molecular mass (130, 120, 110, and 100 kDa) suggests that the bound AMP could play a regulatory rather than a catalytic role. Using the nonhydrolyzable ATP analogue [alpha, beta-methylene]ATP and dilution experiments with Triton X-100-solubilized membranes, it has been possible to identify the 130-kDa adenylylated polypeptide as a possible target of an adenylylating system. These polypeptides, except the 86-kDa phosphoesterase, are affected in their electrophoretic mobility in the absence of beta-mercaptoethanol. An intercatenary disulfide bond(s) appear(s) to link the polypeptide(s) of 120 kDa and/or 110 kDa in a dimeric structure of apparent molecular mass of 240 kDa. All five polypeptides labeled with [alpha-32P]ATP are glycoproteins bound to the cell plasma membrane.     

DNA-binding activity of hepatitis B e antigen polypeptide lacking the protaminelike sequence of nucleocapsid protein of human hepatitis B virus.

The characteristics of binding of hepatitis B core antigen (HBcAg) and hepatitis B e antigen (HBeAg) polypeptides to hepatitis B virus (HBV) DNA were analyzed. HBcAg polypeptide from recombinant HBV core particles and HBeAg polypeptide from partially purified serum HBeAg were prepared and verified to have molecular weights of 21,500 (P21.5) and of 17,000 (P17) and 18,000 (P18), respectively, by immunoblot analysis. By reaction of these proteins on a nitrocellulose membrane with cloned 32P-HBV DNA, it was revealed that the HBeAg polypeptide, which lacks the C-terminal 34 amino acids of P21.5, as well as the HBcAg polypeptide, bound to the DNA. The secondary structures of nucleocapsid proteins of HBV, woodchuck hepatitis virus, and ground squirrel hepatitis virus were predicted by the Garnier algorithm. Amino acid sequences which, in addition to those of the C-terminal regions, may contribute to binding were proposed to be the 21-amino-acid residues located at amino acids 100 to 120 of the nucleocapsid proteins of these hepadnaviruses.     

The Caenorhabditis elegans gene unc-89, required fpr muscle M-line assembly, encodes a giant modular protein composed of Ig and signal transduction domains

Mutations in the Caenorhabditis elegans gene unc-89 result in nematodes having disorganized muscle structure in which thick filaments are not organized into A-bands, and there are no M-lines. Beginning with a partial cDNA from the C. elegans sequencing project, we have cloned and sequenced the unc-89 gene. An unc-89 allele, st515, was found to contain an 84-bp deletion and a 10-bp duplication, resulting in an in- frame stop codon within predicted unc-89 coding sequence. Analysis of the complete coding sequence for unc-89 predicts a novel 6,632 amino acid polypeptide consisting of sequence motifs which have been implicated in protein-protein interactions. UNC-89 begins with 67 residues of unique sequences, SH3, dbl/CDC24, and PH domains, 7 immunoglobulins (Ig) domains, a putative KSP-containing multiphosphorylation domain, and ends with 46 Ig domains. A polyclonal antiserum raised to a portion of unc-89 encoded sequence reacts to a twitchin-sized polypeptide from wild type, but truncated polypeptides from st515 and from the amber allele e2338. By immunofluorescent microscopy, this antiserum localizes to the middle of A-bands, consistent with UNC-89 being a structural component of the M-line. Previous studies indicate that myofilament lattice assembly begins with positional cues laid down in the basement membrane and muscle cell membrane. We propose that the intracellular protein UNC-89 responds to these signals, localizes, and then participates in assembling an M-line.     

Translocation of the Retinoblastoma Gene Product during Mitosis

This study reports the immunocytochemical localization of the retinoblastoma gene product within synchronized normal human keratinocytes. Data suggest that mitotic spindles function in the transport of the retinoblastoma tumor suppressor gene product during cell division. A diffuse anti-pRB reactivity was detected within the nuclei of G₁-phase keratinocytes, although staining was not evident within the nucleoli. During S-phase and G₂-phase the anti-pRB reactivity was localized to discrete regions within the nuclear compartment. The anti-pRB reactivity of M-phase cells was localized to the mitotic spindles and microtubule nucleation centers. Immunoprecipitation and Western blotting of the pRB antigen from synchronized keratinocytes showed that the apparent polypeptide molecular weight of pRB increased from 105 kDa during G₁-phase to 115 kDa during M-phase. Immunoprecipitation of the pRB antigen from mitotic spindles resulted in the coprecipitation of two polypeptides with apparent polypeptide molecular weights of 115 and 50 kDa. Western blotting of the immunoprecipitates from purified keratinocyte mitotic spindles showed that β-tubulin was the 50-kDa polypeptide associated with hyperphosphorylated pRB.     

Localization on the viral genome and nucleotide sequence of the gene coding for the two major polypeptides of the hepatitis B surface antigen (HBs Ag).

The structural gene coding for both polypeptides I and II which are the two major polypeptides of the Hepatitis B surface antigen, is found to be localized on the viral genome. This gene, referred to as gene S, is located in the partially single stranded region. It maps between positions 73.6 and 95.1% of the genome length. It is composed of 678 nucleotides, which correspond to a theoretical polypeptide of 25,422 molecular weight.     

Expression of hepatitis B virus large envelope polypeptide inhibits hepatitis B surface antigen secretion in transgenic mice.

The outer membrane of the hepatitis B virus consists of host lipid and the hepatitis B virus major (p25, gp28), middle (gp33, gp36), and large (p39, gp42) envelope polypeptides. These polypeptides are encoded by a large open reading frame that contains three in-phase translation start codons and a shared termination signal. The influence of the large envelope polypeptide on the secretion of hepatitis B surface antigen (HBsAg) subviral particles in transgenic mice was examined. The major polypeptide is the dominant structural component of the HBsAg particles, which are readily secreted into the blood. A relative increase in production of the large envelope polypeptide compared with that of the major envelope polypeptide led to profound reduction of the HBsAg concentration in serum as a result of accumulation of both envelope polypeptides in a relatively insoluble compartment within the cell. We conclude that inhibition of HBsAg secretion is related to a hitherto unknown property of the pre-S-containing domain of the large envelope polypeptide.     

Evidence for the presence of calsequestrin in two structurally different regions of myocardial sarcoplasmic reticulum

Localization of calsequestrin in chicken ventricular muscle cells was determined by indirect immunofluorescence and immuno-Protein A-colloidal gold labeling of cryostat and ultracryotomy sections, respectively. Calsequestrin was localized in the lumen of peripheral junctional sarcoplasmic reticulum, as well as in the lumen of membrane-bound structures present in the central region of the I-band, while being absent from the lumen of the sarcoplasmic reticulum in the A-band region of the cardiac muscle cells. Since chicken ventricular muscle cells lack transverse tubules, the presence of calsequestrin in membrane bound structures in the central region of the I-band suggests that these cells contain nonjunctional regions of sarcoplasmic reticulum that are involved in Ca2+ storage and possibly Ca2+ release. It is likely that the calsequestrin containing structures present throughout the I-band region of the muscle cells correspond to specialized regions of the free sarcoplasmic reticulum in the I-band called corbular sarcoplasmic reticulum. It will be of interest to determine whether Ca2+ storage and possibly Ca2+ release from junctional and nonjunctional regions of the sarcoplasmic reticulum in chicken ventricular muscle cells are regulated by the same or different physiological signals.     

Integral membrane proteins specific to the inner nuclear membrane and associated with the nuclear lamina

We obtained a monoclonal antibody (RL13) that identifies three integral membrane proteins specific to the nuclear envelope of rat liver, a major 75-kD polypeptide and two more minor components of 68 and 55 kD. Immunogold labeling of isolated nuclear envelopes demonstrates that these antigens are localized specifically to the inner nuclear membrane, and that the RL13 epitope occurs on the inner membrane's nucleoplasmic surface where the nuclear lamina is found. When nuclear envelopes are extracted with solutions containing nonionic detergent and high salt to solubilize nuclear membranes and pore complexes, most of these integral proteins remain associated with the insoluble lamina. Since the polypeptides recognized by RL13 are relatively abundant, they may function as lamina attachment sites in the inner nuclear membrane. Major cross-reacting antigens are found by immunoblotting and immunofluorescence microscopy in all rat cells examined. Therefore, these integral proteins are biochemical markers for the inner nuclear membrane and will be useful models for studying nuclear membrane biogenesis.     

Evidence for an Association of the 15-kDa Proteolipid of Mediatophore with a 14-kDa Polypeptide

The present report shows that mediatophore, a nerve terminal membrane protein that translocates acetylcholine on calcium action, forms a complex with a 14-kDa polypeptide. The complex was identified based on the following results. (a) A polyclonal antimediatophore antiserum that immunoprecipitates activity precipitates both the 15- and 14-kDa polypeptides. (b) After HPLC purification of mediatophore, both antigens were found in the same peak. (c) After 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate solubilization of presynaptic membranes or of the purified mediatophore, an immunoaffinity column made with the anti-14-kDa antigen monoclonal antibody retained both the 14-kDa and the 15-kDa polypeptide. Similarly, immunoprecipitation experiments using protein A-coated beads sedimented an immunocomplex in which both antigens were found. (d) The 14-kDa antigen could be localized in the synaptosomal membrane where mediatophore and its 15-kDa component are found.     

Immunoelectron microscopic evidence for organ differences in the composition of peroxisome-specific membrane polypeptides among three rat organs: liver, kidney, and small intestine

We examined the distribution of peroxisome-specific membrane polypeptides (PMPs) among peroxisomes of the liver, renal cortex, and jejunal mucosa, using antibodies for 70 KD, 26 KD and 22 KD PMPs. Immunoblot analysis showed signals for 70 KD polypeptide in all three kinds of tissue, but for the other two only in the liver and renal cortex, with neither being detected in jejunal mucosa. The total amounts of PMPs increased in all three organs with DEHP (di-(2-ethylhexyl)phthalate) administration. By immunoelectron microscopic analysis using protein A-gold, the three PMPs were localized along the peroxisomal membrane. Quantitation of the gold particles associated with the peroxisomal membrane showed an increase in the density of 70 KD and 26 KD PMPs but a decrease in 22 KD PMP with the administration of DEHP. The presence of tissue-specific localizations of PMPs suggest the 70 KD PMP is a common constituent of peroxisomes of these three tissues, whereas 26 KD and 22 KD PMPs are absent in microperoxisomes of jejunal mucosal epithelium.