bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Possible Role for Ligatin and the Phosphoglycoproteins It Binds in Calcium‐Dependent Retinal Cell Adhesion
Authors: Marchase Richard B. , Koro Lillian A. , Kelly Charles M. , McClay David R. 
Published: December 2004

AbstractLigatin is a filamentous plasma membrane protein that serves as a baseplate for the attachment of peripheral glycoproteins to the external cell surface. Ligatin can be released from intact, embryonic chick neural retinal cells by treatment with 20 mM Ca++ without adversely affecting their viability. α‐Glucose‐1‐phos phate is also effective in removing ligatin‐associated glycoproteins from intact cells. After either of these treatments, the retinal cells seem not to exhibit Ca++ ‐dependent adhesion for one another. It is thus suggested that ligatin in neural retina may serve as a baseplate for the attachment to the cell surface of glycoproteins active in Ca++‐dependent adhesion. The finding that Ca++ serves to protect Ca++‐dependent adhesion molecules from digestion by trypsin is discussed in relation to steric constraints on trypsin's accessibility to these adhesion molecules because of their possible binding to arrayed ligatin filaments.
Spectrin Domains: Proteolytic Susceptibility as a Probe of Protein Structure
Authors: Speicher David W. , Marchesi Vincent T. 
Published: December 2004

AbstractMild treatment of human erythrocyte spectrin with trypsin produces discrete intermediate‐sized peptides. The effects of buffer composition, enzyme–substrate ratio, temperature, and other experimental parameters on the resulting peptide pattern have been examined. Spectrin is capable of regaining its proteolytic resistance after NaDodSO4‐induced denaturation, permitting the use of isolated subunits to study spectrin structure and function. Tryptic digestion of isolated subunits also has greatly facilitated the identification of the subunit origin of the intermediate‐sized peptides. Isolated subunits could also be recombined to form functional units similar but not identical to the native dimeric form of the molecule. Spectrin apparently is composed of numerous large protease‐resistant regions or domains connected by small protease sensitive segments. The structural integrity and accessibility of these sites is minimally affected by oligomeric state or proteolytic digestion conditions. The similarities of sizes, isoelectric points, and amino acid compositions of many intermediate‐size peptides from areas of both subunits suggest that at least part of spectrin's structure may have evolved via replication of a single gene. A possible structural repeat of approximately 50,000 daltons is hypothesized.
Inhibition of Neoplastic Cell Growth by Quiescent Cells Is Mediated by Serum Concentration and cAMP Phosphodiesterase Inhibitors
Authors: Bertram John S. , Bertram Barbara B. , Janik Przemyslaw 
Published: December 2004

AbstractWe have demonstrated that confluent monolayers of the mouse fibroblast cell line C3H/10T1/2 (10T1/2) have the ability to cause reversible growth inhibition of cocultured transformed cells. This was first demonstrated for de novo transformed cells and later extended to established cell lines of proven oncogenicity in vivo. This growth inhibition could be increased by growing the 10T1/2 cells to high density in increasing concentrations of serum or by elevating intracellular concentrations of cAMP using inhibitors of phosphodiesterase (PDE). These manipulations, which in cocultures of nontransformed and transformed cells caused complete inhibition of tumor cell growth, had no effect on growth rate or saturation density of either ceil type when cultured alone, demonstrating the cooperative nature of this phenomenon. This cooperation could not be produced by transfer of culture medium, demonstrating the requirement for intimate cell contact. Inhibition of the formation of transformed foci of cells in these mixed cultures was accompanied by a decrease in the incorporation of labeled thymidine into these cultures; the kinetics of this inhibition and recovery suggested a rapidly reversible effect on cell cycle transit times. The potent inhibitor of cAMP PDE, Ro 20‐1724 induced dose dependent increases in intracellular cAMP in both nontransformed and in transformed cells. However, at a concentration of 10−4 M Ro 20‐1724, which inhibited tumor cell growth in mixed cultures, cAMP was elevated 30‐fold in nontransformed versus only 3‐fold in transformed cells.The inhibitory effects of PDE inhibitors on tumor growth have been extended to an in vivo model system, utilizing Lewis lung carcinoma cells growing as metastases in the lungs of C57B1 mice. In these mice, inoculated intravenously with a single cell suspension of Lewis lung cells, the formation of lung metastases was dramatically decreased by the twice daily administration of either isobutylmethylxanthine or Ro 20‐1724; PDE inhibitors were shown to be active in vitro. The latter compound, which showed highest activity in vitro, was also substantially more potent in vivo as an inhibitor of lung tumor colony formation and doubled the life span of the tumor bearing animals. Cell cycle analysis of lung tumor colonies by the labeled mitosis method showed that both phosphodiesterase inhibitors caused a prolonged G1 phase in the cell cycle but failed to influence other phases. Although detailed analysis of host tissues is not complete, prolonged treatment with these drugs caused no statistically significant weight loss or changes in counts of red or white blood cells indicating a selective growth inhibition of transformed cells at these doses. Studies to determine the mechanism of the cellular communication and the nature of the signal are in progress.
A study of cAMP binding proteins on intact and disrupted sperm cells using 8‐azidoadenosine 3′,5′‐cyclic monophoshate
Authors: Schoff Patrick K. , Forrester Ian T. , Haley Boyd E. , Atherton Robert W. 
Published: December 2004

AbstractThe photoaffinity probe (32P)8‐N3 cAMP was used to label the cAMP binding proteins in washed ejaculated human sperm. Three saturable binding proteins were photolabeled in both intact and disrupted cells with apparent molecular weights of 55,000, 49,000 and 40,000 daltons corresponding to the regulatory subunits of type II and type I cAMP‐dependent protein kinase (cAMP‐PK) and to an endogenous proteolytic product of the regulatory subunits, respectively. Photoincorporation in the three proteins could be totally blocked by preincubating the cells with cAMP. Cell‐free seminal plasma was found to be free of detectable (12P) 8‐N3 cAMP‐binding proteins. The 8‐N3 cAMP was also effective in stimulating endogenous cAMP‐PK activity in intact and disrupted sperm. A substantial amount of (32P) 8‐N3 cAMP binding to types I and II regulatory subunits and cAMP‐PK activity was detected on washed intact cells, intact cells. Intact cell bound 1.80 pmol of (32P) 8‐N3 cAMP/mg protein and had cAMP‐PK activity of 824 units/108 cells. Disrupted cells bound 3.95 pmol (32P) 8‐N3 cAMP mg protein and had a cAMP‐PK activity of 2,206 units/108 cells. The data presented support the concept of two classes of cAMP receptors being differentially available to externally added (32P) 8‐N3 cAMP and proteases. Cellular membrane integrity and membrane sidedness are discussed as possible explanations for the observation reported.
Macromolecular synthesis and stability in growth‐arrested BALB/C‐3T3 cells
Authors: Wharton Walker , Pledger W. J. 
Published: December 2004

AbstractConcentrations of methylglyoxal bis‐(guanylhydrazone) (mGBG) that inhibited serum‐stimulated BALB/c‐3T3 cells in late G1 caused a marked inhibition of 3H‐leucine incorporation during a 20‐min incubation. No decrease was observed in the incorporation of 3H‐uridine during a 20‐min incubation; however, the amount of acid‐insoluble 3H‐uridine in mGBG‐treated cultures was decreased when the incubation period was longer than 20 min. The amount of the decrease in the accumulation of incorporated 3H‐uridine was directly proportional to the length of the incorporation time. Between 10 and 12 h after quiescent BALB/C‐3T3 cells were serum‐stimulated in mGBG no additional 3H‐uridine was accumulated. The stability of the incorporated 3H‐uridine, as determined by acid‐insoluble radioactivity remaining after the addition of actinomycin D, was less in cells cultured in mGBG. Exogenous spermine or spermidine reversed the inhibition of 3H‐uridine accumulation in acid‐insoluble material produced by mGBG as well as the decrease in stability of the incorporated 3H‐uridine in acid‐insoluble material. The effects of mGBG on both the incorporation of 3H‐uridine and the stability of the incorporated 3H‐uridine can apparently be accounted for by an effect on ribosomal RNA.
Heterogeneity of 14S and 30S dynein ATPase activities with respect to activation by calmodulin
Authors: Blum Jacob J. , Hayes A. , Vanaman T. , Schachat F. H. 
Published: December 2004

AbstractDemembranated cilia of Tetrahymena pyriformis were extracted with KCl or Tris‐EDTA and the crude dyneins from each resolved by sucrose density gradient sedimentation into 14S‐K, 30S‐K, 14S‐E and 30S‐E dyneins, respectively. The calmodulin activation ratio (ATPase activity in presence of added calmodulin/ATPase activity in absence of added calmodulin) did not vary across the 30S dynein fractions regardless of the method of extraction nor did it vary across the 14S‐E region. However, in going from the “heavy” fractions to the “light” fractions of the 14S‐K region, it increased markedly.The concentration of calmodulin required for half‐maximal activation did not differ appreciably in the “light” versus the “heavy” fractions of the 14S‐K region, suggesting that the affinity for calmodulin does not vary in these fractions. SDS‐polyacrylamide gel electrophoresis studies showed the presence of several polypeptides that varied in a systematic fashion across the 14S‐K region and hence may be involved in regulating the sensitivity of 14S‐K dynein ATPase activity to added calmodulin.
Protein‐dependent lipid lateral phase separation as a mechanism of human erythrocyte ghost resealing
Authors: Minetti Maurizio , Ceccarini Marina 
Published: December 2004

AbstractThe hypothesis of a correlation between a 10°–20°C lipid phase transition and the resealing process of human erythrocyte membrane has been investigated. The conditions required to reseal human erythrocyte ghosts have been studied by measuring the amount of fluorescein‐labeled dextran (FD) that is trapped into the membrane.Temperature per se was sufficient to induce membrane resealing: (1) at 5 mM sodium phosphate, pH 7.8 (5P8), resealing began at 12°C; (2) at salt concentrations above 8 mM sodium phosphate, it occurred at lower temperature; and (3) in isotonic saline was detected just above 5°C.The removal of peripheral membrane proteins from unsealed membranes by chymotrypsin at 0°C in 5P8 was followed by membrane resealing.This seems to imply that the presence of proteins is necessary to maintain the membrane unsealed. Protein‐induced lateral phase separation of lipids may be a reasonable mechanism for the observed phenomena. In fact, the permeability of phosphatidylserine‐phosphatidylcholine mixed liposomes to FD is modified by lipid lateral phase separation induced by pH or poly‐L‐lysine.Electron spin resonance studies of membrane fluidity by a spin labeled stearic acid showed a fluidity break around 11°C, which may be due to a gel–liquid phase transition. Fluidity changes are abolished by chymotrypsin treatment.It is suggested that a lateral phase separation is responsible for the permeability of open ghosts to FD. Accordingly, disruption of phase separation apparently produces membrane reconstitution. In this respect peripheral proteins and particularly the spectrin‐actin network, may play a major role in membrane resealing.
Detection of a complex of SV40 large tumor antigen and 53K cellular protein on the surface of SV40‐transformed mouse cells
Authors: Santos Myriam , Butel Janet S. 
Published: December 2004

AbstractThe possible interaction between simian virus 40 (SV40) large tumor antigen (T‐ag) and cellular proteins in the plasma membrane of SV40‐transformed mouse cells was investigated. The presence of SV40 T‐ag, 53,000 (53K) cellular protein, and histocompatibility (H‐2) antigens on the surface of SV40‐transformed cells was demonstrated by immunofluorescence. The use of lactoperoxidase‐catalyzed cell surface iodination and a differential immunoprecipitation technique established that large T‐ag is associated with the 53K host‐coded protein on the surface of the transformed cells. In contrast, no detergent‐stable complex between large T‐ag and H‐2 antigens was detected. Both labeled T‐ag and 53K protein were coprecipitated from surface‐iodinated SV40‐transformed cells by monoclonal antibodies directed against either the viral or the cellular protein. Based on the unique antigenic sites recognized by the anti‐T monoclonal antibodies, it appears that both the carboxy and amino termini of the T‐ag polypeptide are exposed on the surface of SV40‐transformed mouse cells. The nature of the association between surface T‐ag and 53K protein, as well as that between the molecular complex and the plasma membrane, remains to be determined. The possible effect of the surface‐associated T‐ag/53K complex on cellular proliferation is considered.
Parameters for crystal growth of ribosomal subunits
Authors: Yonath A. , Müssig J. , Wittmann H. G. 
Published: December 2004

AbstractA systematic analysis of the parameters that control the crystal growth of the large subunit of ribosomes from B stearothermophilus has been carried out. Several parameters have been identified and classified according to their significance. It was found that only biologically active particles can crystallize and that the critical period for the crystallization process is the first few days, during which changes in the volume and content of the crystallization drop are of importance for both nucleation and crystal growth. Consequently, an experimental procedure for fine control of these variables has been developed. As a result of these studies, the reproducibility of crystal formation was increased, and larger and more stable crystals have been obtained.
Biochemical and morphological properties of bovine erythrocyte membrane glycoproteins
Authors: Fletcher M. A. , Brunschwig J. P. , Lo H. , Caldwell K. E. , Lo T. M. 
Published: December 2004

AbstractThe major and minor sialoglycoproteins of the bovine erythrocyte have been solubilized and extensively purified. A comparison of composition revealed that the major glycoprotein had 77% carbohydrate and 23% peptide, and the minor one had 27% carbohydrate and 73% peptide. Molar ratios of sugars were related, however, the major glycoprotein had twice as much galactose and sialic acid as did the minor glycoprotein. Molecular weights, estimated from retardation coefficients of mobility in sodium dodecyl sulfate gel electrophoresis, were 55,000 for the major glycoprotein and 34,000 for the minor glycoprotein. The glycoproteins were studied by electron microscopy before and after delipidation and after ultracentrifugation. The major glycoprotein, prior to delipidation, formed large micelles. After delipidation, the major glycoprotein could not be visualized suggesting that it did not form aggregates in aqueous solution. The minor glycoprotein was visualized as rather uniform spherical aggregates (62 Å average diameter) which tended to form short chains and small clumps. These characteristic aggregates were seen both before and after delipidation. After ultracentrifugation, fixation and sectioning both glycoproteins appeared to have formed microcrystalline arrays with average periodicity of 49 Å.
The antigenic structure of the acetylcholine receptor from torpedo californica
Authors: Gullick W. J. , Lindstrom J. M. 
Published: December 2004

AbstractThe immunological structure of the acetylcholine receptor (AChR) from the electric organ of Torpedo californica was studied using a large number of monoclonal antibodies which were initially selected for their abilities to bind to intact AChRs. The monoclonal antibodies were tested for their ability to bind to denatured AChR subunits labeled with 125I. Antibodies derived from rats immunized with individual denatured subunits or a mixture of subunits of Torpedo AChR reacted well in the assay. A much smaller proportion of antibodies derived from rats immunized with native Torpedo AChR or native AChR from Electrophorus electricus electric organ, bovine muscle, or human muscle reacted with denatured subunits of Torpedo AChR. Many monoclonal antibodies reacted with more than one subunit, but they always reacted best with the subunit used for immunization.Those monoclonal antibodies that bound to intact subunits were mapped more precisely by their ability to bind characteristic fragments of each subunit generated by proteolysis with Staphylococcal V8 protease. These fragments were analyzed by SDS polyacrylamide gel electrophoresis, and monoclonal antibodies that precipitated the same fragment pattern were placed in groups. By this method, we define a minimum of 28 determinants on Torpedo AChR.
Modulation of the neuronal binding of the β subunit of nerve growth factor (NGF) by the α‐NGF subunit
Authors: Woodruff Nathaniel R. , Neet Kenneth E. 
Published: December 2004

AbstractThe effect of the α subunit of the 7S‐NGF on the binding of β‐NGF to its two classes of sites on target cells has been studied. The presence of μM concentrations of α‐NGF causes the displacement of 125I‐β‐NGF from one class of sites on dissociated dorsal root ganglia neurons from stage E9 chicken embryos. At O.1 nM 125I‐β‐NGF, increasing α‐NGF concentrations produce a monotonic displacement curve with half‐maximal displacement occurring at 10 μM α‐NGF. The affinity and number of sites of the 125I‐β‐NGF displaced by α‐NGF are similar to those of β‐NGF that binds to the higher affinity (site I) receptors. The binding to the lower affinity class of sites (site II) is not affected‐by concentrations of α‐NGF up to 30 μM. This modulation of 125I‐β‐NGF binding does not occur with equivalent concentrations of serum albumin. No detectable neuronal binding of 125I‐β‐NGF was found, suggesting that the mechanism does not involve direct competition for receptor sites. The dissociation constant for the α‐β complex is in the μM range, and formation of this complex in solution can thus compete with the process of 125I‐β‐NGF binding to neurons. A model accounting for these observations includes binding of the α‐β complex to the lower affinity but not to the higher affinity sites. We conclude that there are differences in the specificity of the two classes of receptors.
Multiple specific binding sites for purified glucocorticoid receptors on mammary tumor virus DNA
Authors: Payvar Farhang , Firestone Gary L. , Ross Susan R. , Chandler Vicki L. , Wrange Örjan , Carlstedt‐Duke Jan , Gustafsson Jan‐åke , Yamamoto Keith R. 
Published: December 2004

AbstractGlucocorticoid hormones selectively stimulate the rate of transcription of integrated mammary tumor virus (MTV) sequences in infected rat hepatoma cells. Using two independent assays, we find that purified rat liver glucocorticoid receptor protein binds specifically to at least four widely separated regions on pure MTV proviral DNA. One of these specific binding domains, which itself contains at least two distinct receptor binding sites, resides within a fragment of viral DNA that maps 110–449 bp upstream of the promoter for MTV RNA synthesis. Three other binding domains lie downstream of the promoter and within the MTV primary transcription unit. Restriction fragments bearing separate binding domains have been introduced into cultured cells; transformants have been recovered in which the introduced fragments arc expressed under glucocorticoid control. Thus, it appears that this assay will be useful for assessing the biological significance of the receptor binding sites detected in vitro.
Cells transformed by a wide variety of agents express higher abundance levels of some cellular RNA species
Authors: Schutzbank T. , Levine A. J. 
Published: December 2004

AbstractA cDNA‐cloned library was prepared from mRNA synthesized by SV40‐transformed mouse cells. Eleven cDNA clones were selected based on their ability to hybridize higher levels of mRNA in SV40‐transformed 3T3 cells than in 3T3 cells. These cDNA clones were employed to screen the steady‐state levels of cytoplasmic RNAs in a wide variety of viral (SV40, polyoma, adenovirus, and Rous sarcoma virus) and nonviral (methylcholanthrene, embryonal carcinoma) transformed cell lines. Two of the cDNA clones—A17 and 104—detected greater than 40–100‐fold higher levels of mRNA in all the transformed cell lines tested when compared to nontransformed cells (3T3, C3HEF). The levels of mRNA complementary to these two cDNAs were regulated in a temperature‐sensitive fashion (87–100‐fold) in both SV40tsA‐ and RSV ts‐src‐transformed murine cell lines. These two cDNA clones detected greater than 100‐fold, higher levels of complementary RNA derived from SV40 tumor tissue than in normal mouse liver. RNA species complementary to cDNA clones A17 or 104 were not detected in either actively growing nontransformed cells or in serum‐stimulated 3T3 cells. The abundance levels of mRNAs detected by these two cDNA clones appear to be regulated 100‐fold or greater by the transformed state, independent of the transforming agent. The higher levels of these RNA species detected in transformed mouse cells appear not to be solely regulated by the state of growth of nontransformed cells.
Sequence analysis and in vitro transcription of portions of the epstein‐barr virus genome
Authors: Deininger P. L. , Bankier A. , Farrell P. , Baer R. , Barrell B. 
Published: December 2004

AbstractThe 17,180 base‐pair Eco‐RI‐C fragment of Epstein‐Barr virus has been sequenced in its entirety. This same fragment has also been analyzed for RNA polymerase II promoters, which are active in a soluble in vitro assay. These data are compared to the availability of predicted open reading frames and other potential nucleotide signals associated with transcription. In addition, the DNA sequence of a number of previously undetected repeated DNA sequences from this and several nearby regions of the viral genome are reported.
Characterization of viral genomes in the liver and serum of chimpanzee long‐term hepatitis B virus carriers: A possible role for supercoiled HBV‐DNA in persistent HBV infection
Authors: Ruiz‐Opazo Nelson , Chakraborty Prasanta R. , Shafritz David A. 
Published: December 2004

AbstractIn chimpanzee hepatitis B virus (HBV) carriers, the molecular mechanism for viral persistence has been examined by analyzing the properties of viral DNA molecules in liver and serum. Two extrachromosomal HBV‐DNA molecules migrating on Southern blots at 4.0 kb and 2.3 kb were observed in chimpanzee liver DNA. There was no evidence for integration of HBV sequences into the host genome. The HBV‐DNA molecule which migrated at 4.0 kb position represents a full‐length “nicked,” relaxed circular form, and the DNA molecules migrating at 2.3 kb position represents a supercoiled form of the HBV genome. Evidence for supercoiled HBV‐DNA in serum was obtained by production of the relaxed circular intermediate upon digestion of Dane particle DNA with specific nucleases S1 and Bal 31. A possible role of these two extrachromosomal HBV‐DNA molecules in the biology of hepatitis B virus infection and the mechanism for viral persistence arc discussed.