Max M. Burger

Sialic acid contents and controls of normal and malignant cells

The sialic acid contents of normal and malignant cells have been determined by two procedures. Three lines of virus-transformed cells had consistently lower contents than normal cells. These same lines showed clear differences in contact inhibition of growth. The first enzyme of sialic acid biosynthesis, l-glutamine: d-fructose-6-phosphate aminotransferase (EC 2.6.1.16) was determined also. No significant differences between the kinetic parameters for activity or feedback inhibition by uridine diphospho-N-acetylglucosamine was found. It is concluded that there is no positive correlation between sialic acid content and loss of contact inhibition in the cell lines examined.

Cell Surfaces in Neoplastic Transformation

Publisher Summary This chapter describes the properties, and the chemical and physiological aspects of cell surfaces in neoplastic transformation. Immunological surface disparities between normal and tumor cells have been studied for decades, while attempts to bring them into the realm of biochemistry have been extremely rare. It can be assumed that firm biochemical knowledge regarding the structure and formation of the surface antigens of the tumor cell would contribute significantly to an understanding of neoplastic transformation. The chapter describes two types of chemical changes that occur in the transformed surface membrane: (1) changes in the overall composition and (2) changes in macromolecular structure. Several reports point to a decrease of total carbohydrates in transformed membranes, which may involve glycolipids and glycoproteins, which are primarily lacking their terminal carbohydrate portions. A rearrangement of the surface layers leading to a state in which cells become agglutinable or adsorb various types of antibodies occurs at the same time.

Isolation and characterization of agglutinin receptor sites: II. Isolation and partial purification of a surface membrane receptor for wheat germ agglutinin☆

Four different chemical extraction procedures for the isolation of wheat germ agglutinin receptor sites from L1210 cells are described. Fractionation of the biologically active material on Sephadex G-200 columns in pyridine results in two major peaks, the lower molecular weight fraction having a higher inhibitory activity. Electrophoresis in polyacrylamide sodium dodecyl sulfate gels yields four bands. The most active fraction from Sephadex G-200 has an approximate molecular weight between 40000–60000. A preliminary analysis of the active material indicates the presence of sialic acid, neutral sugars and amino sugars, including N-acetylglucosamine.

The chromaffin granule surface: the presence of actin and the nature of its interaction with the membrane

The involvement of the cytoskeleton in stimulussecretion coupling has been implied through several lines of evidence (reviewed in [ 1] ). Secretion can be inhibited by drugs known to disrupt cytoskeletal function [2], microfilamentous structures have been detected morphologically in secretory cells [3] and actin has been identified as a prominent component in such tissues [4,5]. Microfilaments and/or actin have been found in association with isolated secretory granules [6] and purified actin has been shown capable of ‘reassociating’ with membranes of set, .-tory granules [7,8]. There has been a good deal of controversy surrounding the question of whether or not secretory granules from adrenal medulla (chromaffin granules) possess endogenous actin. Datahas been presented [9] suggesting that actin is not present in isolated granules, while earlier reports have suggested that there is actin that associates with granule membranes [7]. While the reports in [7,9] relied solely on data obtained by gel electrophoresis (and fingerprint analysis [7] but not on granule membrane fractions), we used immunoprecipitation with monospecific antibodies to actin to test the presence of actin on granule membranes. Due to the lack of information on secretory granule-

Cell-surface changes after infection with oncogenic viruses: requirement for synthesis of host DNA.

The change in the surface structure of cultutred mammalian cells infected with oncogenic DNA viruses was similar to that described for the fully and permanently transformed cell surfaces. The course of appearance of this change was established. Synthesis of host DNA is required for expression of the surface change triggered by infection with the oncogenic virus.

The chromaffin granule surface localization of carbohydrate on the cytoplasmic surface of an intracellular organelle

Intact chromaffin granules from bovine adrenal medulla are shown to have complex carbohydrates on their external (cytoplasmic) surface. This is demonstrated by the facts (1) that granules can be agglutinated by wheat germ agglutinin, and (2) that significant amounts of sialic acid can be removed from the granule surface with neuraminidase. Glycoproteins located in the granule membrane, and not glycolipids, are the molecules that mediate wheat germ agglutinin agglutination. The possible involvement of granule surface carbohydrate in the process of exocytosis is discussed.

A working hypothesis explaining the maintenance of the transformed state by SV40 and polyoma

Abstract A model is presented that attempts to explain how SV40 or polyoma maintains the transformed state or phenotype. It is proposed that SV40 or polyoma codes for a protein involved in the initiation of cellular DNA synthesis (positive control element). This viral directed cellular DNA synthesis is different in some way from normal cellular DNA synthesis. This difference, in addition to other factors (possibly a second viral coded protein), results in a permanent cell surface alteration. This same cell surface alteration is observed transiently (only during mitosis) in the normal cell cycle. The permanent cell surface change permits the synthesis or activation of the viral positive control element for cellular DNA replication by-passing the normal cellular control for the initiation of DNA synthesis and allowing the cell to enter a new S period. This results in loss of density growth and expression of the transformed phenotype. The model is presented as a working hypothesis that makes specific predictions for new experiments.

Isolation and characterization of agglutinin receptor sites: III. Studies on the interaction with other lectins

Abstract 1. 1. A wheat germ agglutinin receptor fraction isolated from surface membranes of leukemia cells failed to interact with concanavalin A but did interact to a lesser degree with two other agglutinins. 2. 2. A preparation of concanavalin A that was not multivalent but presumably monovalent and which blocked the agglutination of susceptible cells by intact concavalin A had no effect on the agglutinability of the same cells by wheat germ agglutinin. 3. 3. Antiserum against the isolated wheat germ agglutinin receptor fraction from leukemia cells did not react with the normal lymphocytes nor did it react with red blood cells of syngeneic mice. 4. 4. The same antiserum against the wheat germ agglutinin receptor fraction inhibited the agglutination of L1210 (leukemia), Py 3T3 and Py BHK cells by wheat germ agglutinin but had no effect on the agglutination by concanavalin A of Py 3T3 or Py BHK cells. 5. 5. It was concluded that at least some lectin receptors are chemically and topographically on the membrane distinct from each other.

Isolation of in situ crosslinked ligand-receptor complexes using an anticrosslinker specific antibody

Abstract A new method designed for the specific isolation and characterization of ligand-receptor complexes using a heterobifunctional crosslinking agent and immunoprecipitation is described. The complexes are first covalently crosslinked by photoactivation of the crosslinking agent. After lysis of the cells, the crosslinked complexes are immunoprecipitated using an antiserum directed against the crosslinking agent. With this method, ligand-receptor complexes formed in only minute amounts become available for further investigation. By using this anticrosslinker antiserum, different receptor systems can be investigated without raising new receptor- or ligand-specific antibodies for each system. As a test system, a radioiodinated lectin was used as ligand molecule and erythrocyte membranes acted as receptor carriers.

Surface Changes Detected by Lectins and Implications for Growth Regulation in Normal and in Transformed Cells

If transformed cells differ in as many behavioral traits as have been discussed by Dr. Stoker in the previous chapter, one would expect to find quite a few changes in the cell surface of these cells since the surface is probably involved either directly or at least as a mediator of the stimuli which lead to alterations in social behavior of cells among themselves.