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Biomedical subjects

T L Simpson

Publications and source records attributed to T L Simpson.

27 records · Page 2Linked to original sources
PubMed1985

Posttranslational processing of the insulin proreceptor.

Following translation of the insulin proreceptor by 3T3-L1 adipocytes, about 1.5 hours is required for its conversion into active receptor; an additional 1.5 hours is needed for the active receptor to reach the plasma membrane. During this 3-hour period the proreceptor undergoes a complex series of processing events, glycosylation being an essential processing step. Thus, treatment of 3T3-L1 adipocytes with tunicamycin causes the depletion of cellular insulin binding activity and the accumulation of an inactive aglyco proreceptor. To investigate posttranslational processing of normal proreceptor and the role of glycosylation in active receptor formation, metabolic labeling experiments were conducted. The first 35S-labeled intermediate detected is a 190-kDa polypeptide (proreceptor) which is rapidly (t1/2 = 15 minutes) processed into a 210-kDa species. Both polypeptides contain N-linked core oligosaccharide chains, but in the latter case these chains appear to contain terminal N-acetylglucosamine. The 210-kDa precursor is converted slowly (t1/2 = 2 hours) by proteolytic processing into a 125-kDa (alpha') and 83-kDa (beta') species. Immediately prior to insertion into the plasma membrane, 3 hours after its synthesis, the alpha' and beta' precursors are converted to mature receptor composed of alpha (135 kDa) and beta (95 kDa) subunits. The 125-kDa alpha' and 83-kDa beta' precursors are endoglycosidase H-sensitive and their oligosaccharide chains do not contain terminal sialic acid. Just prior to insertion into the plasma membrane the alpha' and beta' precursors are sialylated, giving rise to the 135-kDa alpha and 95-kDa beta receptor subunits and becoming Endo H resistant and neuraminidase sensitive. In the presence of tunicamycin, a 180-kDa aglyco receptor polypeptide accumulates which is not further processed and does not reach the cell surface. It is concluded that N-linked oligosaccharide chains on the proreceptor are required either for its intracellular translocation to the proteolytic cleavage site or for its identification as a target of the cleavage enzyme. Thus, glycosylation of the insulin proreceptor is crucial for proper processing and formation of functional receptor.

Cell Membrane↗
PubMed1984

Role of glycosylation in the processing of newly translated insulin proreceptor in 3T3-L1 adipocytes.

A procedure was developed for the immunoprecipitation of glycosylated and nonglycosylated forms of the insulin receptor and its precursors without prior purification using lectins. 3T3-L1 adipocytes were labeled with [35S]methionine after which 35S-labeled receptor polypeptides were specifically immunoprecipitated and characterized by sodium dodecyl sulfatepolyacrylamide gel electrophoresis. The first 35S-polypeptide detected was a 190-kDa glycosylated proreceptor which was rapidly (t1/2 approximately equal to 15 min) processed to a 210-kDa intermediate. The latter precursor was more slowly (t1/2 approximately equal to 2 h) proteolytically processed to 125-kDa (alpha') and 83-kDa (beta') precursors of the mature alpha- and beta-receptor subunits. Immediately prior to insertion into the plasma membrane, i.e. about 3 h after translation, the alpha'- and beta'-precursor polypeptides were converted to the mature 135-kDa alpha- and 95-kDa beta-receptor subunits. The characteristics of the oligosaccharide moieties of the receptor precursors and products were investigated. The 210-kDa precursor and its two products, the 125-kDa alpha'- and 83-kDa beta'-species, and the mature alpha- and beta-receptor subunits bind tightly to wheat germ lectin, whereas the 190-kDa proreceptor species is not bound. Upon incubation with endoglycosidase H, both the 210- and 190-kDa species are converted to a 180-kDa species. The 125-kDa alpha'- and 83-kDa beta'-species are also cleaved by endoglycosidase H, being reduced in size to 97 and 79 kDa, respectively. Based on their sensitivity to endoglycosidase H and insensitivity to neuraminidase, the oligosaccharide chains of the receptor precursors (190, 210, 125, and 83 kDa) do not contain terminal sialic acid (or other capping sugars). However, near the time of insertion into the plasma membrane, capping of the alpha'- and beta'-species by sialic acid occurs, giving rise to the mature 135-kDa alpha- and 95-kDa beta-receptor subunits, which are partially endoglycosidase H-resistant and neuraminidase-sensitive. When 3T3-L1 adipocytes are treated with tunicamycin, a 180-kDa proreceptor aglycopolypeptide is synthesized which is incapable of undergoing further processing and proteolytic cleavage to the alpha- and beta (or alpha'- and beta'-)-subunits. The 180-kDa species, which appears to be the aglyco-form of hte 190-kDa proreceptor generated by endoglycosidase H, is resistant to trypsin in the intact cell and apparently has not reached the cell surface. Thus, the oligosaccharide moieties of the insulin receptor precursor are crucial for proper processing, intracellular translocation, and formation of functionally competent insulin re

Adipose Tissue↗
PubMed1983

Interaction of germanium (Ge) with biosilicification in the freshwater sponge Ephydatia mülleri: evidence of localized membrane domains in the silicalemma.

In the presence of germanium (Ge) the needle-shaped silica spicules of the freshwater sponge Ephydatia m ulleri are very short and thin and possess bulbs with large spines. SEM-coupled X-ray analyses confirm the incorporation of Ge into the silica. A small number of bulbs are susceptible to erosion by HNO3 and hypochlorite and although the chemical basis of such erosion is presently unknown it suggests the presence of an organic matrix within the bulbs and/or an incomplete polymerization of the silica. Addition of Ge to control media in which silicification is newly initiated increases the incidence of erosion and results in centrally located eroded areas of the silica and discontinuities in its deposition. Removal of Ge from such newly forming structures results in a partial recovery of normal morphology (spine development and thickening of the silica) but only in the central region surrounding the bulbs. Both results establish the presence of a central, active region for silicification and further support the view that there is a distal spreading, away from this center, of transported forms of silica. Secondary centers may also be present. The newly assembled organic core of control structures is associated with tubular elements possibly derived from the surrounding membrane. In such newly silicifying structures the spicule tips contain oriented material in the form of "rays." Both of these new observations increase the likelihood of the presence of an organic matrix within the silica.

Animals↗
PubMed1980

Fluorescent antibody localization of Microciona prolifera aggregation factor and its baseplate component.

Specific rabbit antisera were prepared against purified aggregation factor and its membrane-associated receptor, baseplate, derived from the marine sponge. Microciona prolifera. They were utilized in conjunction with fluorescent-labeled goat anti-rabbit IgG in an assay to demonstrate the surface localizations of both components. The specificity of antibody preparations for AF and BP was demonstrated through inhibition of the rotation-mediated assay by homotypic antibody. This study confirms the presence of aggregation factor on the surface of disaggregated sponge cells maintained in the presence of the divalent cations, Ca++ and Mg++, and its absence when cells are maintained in Ca++ and Mg++-free seawater. The location of BP could also be demonstrated on the cell surface. Aggregation factors and baseplate appear to be heavily distributed on archeocytes and choanocytes, but are localized less intensely on gray cells. Gray cells are typified by yellowish autofluorescence of their intracellular granules in stained and control preparations. The reaction of anti-Microciona aggregation factor with its homotypic factor appeared to be species specificity judged by immunofluorescence assays and by inhibition of rotation-mediated assay by anti-homotypic AF since antibodies prepared against heterotypic AF preparations were unreactive.

Animals↗
PubMed1976

Sex reversal in a freshwater sponge.

Spongilla lacustris exhibits a type of alternative hermaphroditism, new to the phylum, in which a sponge may be exclusively male or female during the period of sexual reproduction one year and the opposite sex the next year. This form of sexuality may facilitate larva production and thus dispersal following colonization of a new habitat. Gametogenesis occurs shortly after gemmule hatching in both males and females but slightly later in males.

Animals↗
PubMedDate not supplied

Screening for childhood physical and sexual abuse among outpatient substance abusers.

Research demonstrates that substance-abusing individuals report substantially higher rates of childhood sexual and physical abuse than the general population. This study sought to test a method of identifying substance-abusing clients with histories of childhood sexual and/or physical abuse and to explore the differences between those reporting childhood abuse and those not. Files of substance abusing clients from two distinct time periods were examined for reports of childhood abuse. At Time 1 (n = 399) clients were not systematically asked about experiences of childhood abuse, and at Time 2 (n = 305) clients were routinely asked about this issue. Results indicate that significantly more male and female clients disclosed childhood abuse at Time 2. Additionally, male clients reporting childhood abuse appeared more distressed than those not reporting abuse; female clients reporting childhood abuse did not appear more distressed than their counterparts.

Adolescent↗