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Relationships of the Structure and Function of the Interferon Receptor to Hormone Receptors and Establishment of the Antiviral State1

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Relationships of the Structure and Function of the Interferon Receptor to Hormone Receptors and Establishment of the Antiviral State1 [CANCER RESEARCH 38, 4172-4185, November 1978] Relationships of the Structure and Function of the Interferon Receptor to Hormone Receptors and Establishment of the Antiviral State1 Evelyn F. Grollman, George Lee, Sofia Ramos, Pedro S. Lazo, H. Ronald Kaback, Robert M. Friedman, and Leonard D. Kohn Sect/on on Biochemistry of Cell Regulation, Laboratory of Biochemical Pharmacology ¡E.F. G., G. L., P. S. L., L. D. K.] and Laboratory of Experimental Pathology [R. M. F.], National Institute of Arthritis, Metabolism, and Digestive Diseases, NIH, and Division of Bacterial Products, Bureau of Biologies, Food and Drug Administration, Bethesda, Maryland 20014 [S. R.], and Laboratory of Membrane Biochemistry, Roche Institute of Molecular Biology, Nutley, New Jersey 07110 [H. R. K.J Abstract between TSH and receptor involves electrostatic interac tions between positively charged residues on the TSH This report describes similarities between the structure molecule and spatially oriented negatively charged residues and function of the Interferon receptor and receptors for within the receptor binding site. It is believed that this initial glycoprotein hormones and several bacterial toxins. Spe electrostatic interaction determines the locus and nature of cifically, it describes several common molecular and supplementary short-range (hydrophobic) interactions mechanistic elements, including: (a) the presence of a which approximate the hormone more firmly within the glycoprotein as well as a ganglioside component in the receptor. These interactions exclude water and salts from receptor; (b) changes in membrane structure as a conse the 2 approximating surfaces and induce a conformational quence of Interferon action; (c) ¡nterferon-inducedintra- change in the hormone. The receptor-induced conforma cellular cyclic adenosine 3':5'-monophosphate changes; tional change, the exact nature of which is encoded by the and (d) alterations in the flux of certain ions across the oligosaccharide determinant of the receptor, allows the « membrane. Since Interferon has an antiviral effect, these subunit of the hormone (or a portion of the a subunit results define a relationship between hormonal perturba similar in sequence to the nonapeptide hormones oxytocin tion of cellular events and the ability of an agent to and vasopressin) to intercalate itself within the membrane prevent or suppress viral infections of cells. Further defi bilayer. Implicit in this series of events is the notion that a nition of these relationships should be important to our receptor with a structure similar to that of the TSH receptor, understanding of the oncogenic state, of hormonal effects but containing a different oligosaccharide determinant, on the oncogenic state, and of other human diseases in might accommodate TSH, but would not trigger the appro which hormonal perturbations of non-target tissues or priate conformational change necessary for message trans cross-reactivity of receptors could be pathogenic. mission. The consequence of these events is a change in the state Introduction of the membrane, which expresses itself (a) as a change in cell surface determinants, such as receptors for other Studies over the past few years indicate that the TSH2 effectors of cell function (23-25, 27), and (b) as a change in receptor is composed of glycoprotein and ganglioside com the electrical potential across the cell membrane (18). ponents and that a key determinant within the receptor is Finally, changes in cyclic AMP levels occur as a result of an oligosaccharide moiety (23-25, 27). Current results indi direct or indirect modulation of components in the ade- cate that the glycoprotein represents a high-affinity binding nylate cyclase system. As noted above, cyclic AMP is then a component which selects TSH from structurally unrelated cellular message which effects changes in the translational effectors such as insulin, glucagon, adrenocorticotropic and transcriptional machinery of the cell in the absence of hormone, prolactin, growth hormone, etc. (2, 25). The any input of additional genetic information in the form of ganglioside, in contrast, is believed to serve as a low-affinity DMA or RNA (as in the case for fertilization or for a virus discriminator which completes the selection process by infection). distinguishing between structurally related effectors, such Evidence has accumulated which indicates that the bind as the other glycoprotein hormones (luteinizing hormone, ing of interferon to specific cell surface receptors is neces follicle-stimulating hormone, and human chorionic gonad- sary for its antiviral action (13). Other studies suggest that otropin). The ganglioside thus contributes to the TSH re the interferon receptor has a ganglioside or ganglioside- ceptor a 3-dimensional structure with the stereospecific like structure as one of its component parts and that an binding properties of an enzyme (2, 25). oligosaccharide moiety on this ganglioside or ganglioside- Current experiments suggest that the initial association like component is a critical feature for receptor function (5- 7, 62). Since cholera toxin and TSH interact with cell 1 Presented at the John E. Fogarty International Center Conference on surface receptors believed to contain a ganglioside as a Hormones and Cancer, March 29 to 31, 1978, Bethesda. Md. functional component (23-25, 27), the following possibili 2 The abbreviations used are: TSH, thyrotropin; cyclic AMP, cyclic aden osine 3':5'-monophosphate; Gsn, galactosyl-/V-acetylgalactosaminyl(N-ace- ties were raised. First, the mechanism by which these 3 tylneuraminyl)galactosylglucosylceramide; GU2, W-acetylgalactosaminyl(N- effectors and interferon transmitted their messages to the acetylneuraminyl)galactosylglucosylceramide, GM3, N-acetylneuraminylga- cell might have elements in common. Secondly, cholera lactosylglucosylceramide; TPMP". triphenylmethylphosphonium ion; EGTA, ethyleneglycol bis(/3-aminoethyl ether)-W,N'-tetraacetic acid; PHA, phytohe- toxin and TSH might be able to block the ability of interferon magglutinin. to induce an antiviral state. 4172 CANCER RESEARCH VOL. 38 Downloaded from cancerres.aacrjournals.org on September 27, 2021. © 1978 American Association for Cancer Research. Interferon Receptor Recent experiments (14, 26) have shown: (a) that cholera Preparation of Membranes, Ganglioside, and Glycopro toxin and TSH do in fact inhibit the establishment of the tein Extractions. Plasma membranes from mouse Ly and antiviral state by interferoni (b) that this action reflects an human KB-3 cells were prepared by adapting methods used effect of these agents on plasma membranes, and (c) that previously to obtain bovine thyroid plasma membranes (3, these effectors do not appear to act as simple competitive 26, 56, 57). Plasma membrane gangliosides were extracted antagonists. according to a modification of the methods of Yu and This report will summarize experiments which further Ledeen (65) and Saito and Hakomori (54). The details of characterize: (a) the structure of the interferon receptor; these procedures are described elsewhere (17). Ganglioside and (b) the mechanism by which interferon-receptor inter standards for these analyses (GM1,GN12,GM3,N - acetylneu- actions might initiate or propagate the antiviral state. In raminylgalactosyl - N - acetylgalactosaminyl[N - acetylneu- sum, the data show that the interferon receptor, like the raminyl]galactosylglucosylceramide, galactosyl-A/-ace- TSH receptor, is a 2-component (glycoprotein and ganglio- tylgalactosaminyl[/V - acetylneuraminyl - N - acetylneurami- side) receptor, that both components are necessary for nyljgalactosylglucosylceramide, and W-acetylneuraminyl- message transmission, that membrane changes are a con galactosyl - N - acetylgalactosaminyl[A/ - acetylneuraminyl - sequence of the interferon-receptor interaction, and that N - acetylneuraminyljgalactosylglucosylceramide) were ob phosphorylated intermediates such as cyclic AMP may be tained as previously described in (40-43, 46, 47). Identifica involved in mediating the response to interferon. In addi tion of the gangliosides in the mouse Ly and human KB-3 tion, this report describes several interferon-induced effects cell membranes was based on content of sialic acid and on ion fluxes across the membrane. terminal galactose residues, sensitivity to neuraminidase, In the context of this conference, we believe these studies and Chromatographie properties. are important in 2 respects. First, they relate hormonal Surface-exposed terminal galactosyl residues on glyco- interactions with cells to the interaction of an agent known lipids and glycoprotein components of membranes were to produce an antiviral state. Secondly, we believe that they labeled with tritium according to modifications of described offer additional insight into the mechanism by which inter procedures (16, 44). Sialic acid residues on the ganglio feron can prevent oncogenic viruses from transforming sides and glycoproteins were tritiated after mild periodate cells. oxidation (44). Details of these procedures are described elsewhere (17). Materials and Methods The glycoprotein component of the interferon receptor was solubilized by extracting membrane preparations with Cells and Viruses. Mouse Ly cells are a strain of inter- 0.1 M lithium diiodosalicylate in analogy with procedures feron-sensitive mouse fibroblasts originally obtained from used to isolate the glycoprotein component of the TSH Dr. J. S. Younger, University of Pittsburgh School of Medi receptor
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