bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mutations in the E1a gene of type 5 adenovirus result in oncogenic transformation of fischer rat embryo cells
Authors: Duigou Gregory J. , Babiss Lee E. , Liaw Wen‐Shing , Zimmer Stephen G. , Ginsberg Harold S. , Fisher Paul B. 
Published: December 2004

AbstractTransformation of a specific clone of Fischer rat embryo (CREF) cells with wild‐type 5 adenovirus (Ad5) or the Ela plus Elb transforming gene regions of Ad5 results in epithelioid transformants that grow efficiently in agar but that do not induce tumors when inoculated into nude mice or syngeneic Fischer rats. In contrast, CREF cells transformed by a host‐range Ad5 mutant. H5hrl, which contains a single base‐pair deletion of nucleotide 1055 in Ela resulting in a 28‐kd protein (calculated) in place of the wild‐type 51‐kd acidic protein, display a cold‐sensitive transformation phenotype and an incomplete fibroblastic morphology but surprisingly do induce tumors in nude mice and syngeneic rats. Tumors develop in both types of animals following injection of CREF cells transformed by other cold‐sensitive Ad5 Ela mutants (H5dl 101 and H5in l06), which contain alterations in their 13S mRNA and consequently truncated 289AA proteins. CREF cells transformed with only the Ela gene (0–4.5 m.u.) from H5hrl or H5dll01 also produce tumors in these animals. To directly determine the role of the 13S Ela encoded 289AA protein and the 12S Ela encoded 243AA protein in initiating an oncogenic phenotype in adenovirus‐transformed CREF cells, we generated transformed cell lines following infection with the Ad2 mutant pm975. which synthesizes the 289AA Ela protein but not the 243AA protein, and the Ad5 mutant H5dl 520 and the Ad2 mutant H2dl 1500, which do not produce the 289AA Ela protein but synthesize the normal 243AA Ela protein. All three types of mutant adenovirus‐transformed CREF cells induced tumors in nude mice and syngeneic rats. Tumor formation by these mutant adenovirus‐transformed CPEF cells was not associated with changes in the arrangement of integrated adenovirus DNA or in the expression of adenovirus early genes. These results indicate, therefore, that oncogenic transformation of CREF cells can occur in the presence of a wild‐type 13S Ela protein or a wild‐type 12S Ela protein when either protein is present alone, but does not occur when both wild‐type Ela proteins are present.
Structure and evolution of the human genes encoding protein C and coagulation factor IX
Authors: Long George L. 
Published: December 2004

AbstractHuman protein C is a vitamin K‐dependent plasma protein that serves as a feedback down‐regulator of the coagulation cascade by specifically degrading the protein cofactors VIIIa and Va. The protein C precursor consists of the following domains: leader peptide, “gla” region, two epidermal growth factor segments, and the activation peptide/serine protease. Comparison of amino acid sequences reveals that protein C and factor IX are homologous. A comparison of the genes for protein C and factor IX shows that all seven of the introns within the protein coding regions are in identical positions and correspond to protein structure‐function domain boundries. However, the base compositions of the two genes (coding and noncoding regions) are remarkably different: ∼60% guanine + cytosine (G + C) for protein C versus ∼40% G + C for factor IX. One possible explanation for this phenomenon is that the factor IX gene (located on the X chromosome) has undergone extensive deoxycytosine methylation and subsequent spontaneous deamination mutagenesis, resulting in a net C to thymine (and G to adenine) transition. This would suggest that the protein C gene may represent a more primitive form of the gene duplication precursor.
Tyrosyl and phosphatidylinositol kinases of human erythrocyte membranes
Authors: Vossler Mark R. , Coco Anna , Strausser Bimmie T. , Zaricznyj Christine , Macara Ian G. 
Published: December 2004

AbstractThe tyrosyl kinase and phosphatidylinositol (PI) kinase activities of human red cells have been partially purified and characterized. Although the PI kinase required detergent for solubilization, the major tyrosyl kinase of the red cell could be extracted by high salt. A very small residual activity remained associated with the membranes, however, that was solubilized with the PI kinase and copurified through an ammonium sulfate precipitation and diethylaminoethyl (DEAE) ion‐exchange step gradient elution. However, the two activities were found to differ with respect to their apparent Kms for ATP and Mg2+; they showed different half‐lives for temperature inactivation, possessed different relative activities in the presence of Mn+ and Ca2+, and were separable by elution from a DEAE‐Trisacryl ion exchange column using a linear NaCl gradient. The kinetic parameters of the membrane‐associated tyrosyl kinase differed from those of the salt‐extracted enzyme. PI kinase was not activated by pretreatment with the tyrosyl kinase p68v‐ros or by addition of the phosphotyrosyl phosphatase inhibitor, vanadate, to intact membranes, and was not competitively inhibited by the tyrosyl kinase substrate poly(Glu4,Tyr). We conclude that the human red cell phosphatidylinositol and tyrosyl kinases are distinct and separable activities, and that at least two separable tyrosyl kinases are present in human erythrocytes.
Genes that cooperate with tumor promoters in transformation
Authors: Colburn Nancy H. , Smith Bonita M. 
Published: December 2004

AbstractTumor‐promoting phorbol esters, like growth factors, elicit pleiotropic responses involving biochemical pathways that lead to different biological responses. Genetic variant cell lines that are resistant to mitogenic, differentiation, or transformation responses to tumor promoters have been valuable tools for understanding the molecular bases of these responses. Studies using the mouse epidermal JB6 cell lines that are sensitive or resistant to tumor promoter‐induced transformation have yielded new understanding of genetic and signal transduction events involved in neoplastic transformation. The isolation and characterization of cloned mouse promotion sensitivity genes pro‐1 and pro‐2 is reviewed. A new activity of pro‐1 has been identified: when transfected into human cancer prone basal cell nevus syndrome fibroblasts but not normal fibroblasts mouse pro‐1 confers lifespan extension on these cells. Recently, we have found that a pro‐1 homolog from a library of nasopharyngeal carcinoma, but not the homolog from a normal human library, is activated for transferring promotion sensitivity. The many genetic variants for responses to tumor promoters have also proved valuable for signal transduction studies. JB6 P‐ cells fail to show the 12‐O‐tetradecanoyl‐phorbol‐13‐acctate (TPA)‐induced synthesis of two proteins of 15 and 16 kD seen in P+ cells. P−, P+, and TPA transformed cells show a progressive decrease in both basal and TPA‐inducible levels of a protein kinase C substrate of 80 kD. P− cells are relatively resistant both to anchorage‐independent transformation and to a protein band shift induced by The calcium analog lanthanum. It appears that one or more calcium‐binding proteins and one or more pro genes may be critical determinants of tumor promoter‐induced neoplastic transformation.
Mode of estrogen action on cell proliferation in CAMA‐1 cells: II. Sensitivity of G1 phase population
Authors: Leung Benjamin S. , Potter Anne H. 
Published: December 2004

AbstractThe mammary cancer cell line CAMA‐1 synchronized at the G1/S boundary by thymidine block or at the G1/M boundary by nocodazole was used to evaluate (1) the sensitivity of a specific cell cycle phase or phases to 17β‐estradiol (E2), (2) the effect of E2 on cell cycle kinetics, and (3) the resultant E2 effect on cell proliferation. In synchronized G1/S cells, E2‐induced 3H‐thymidine uptake, which indicated a newly formed S population, was observed only when E2 was added during, but not after, thymidine synchronization. Synchronized G2/M cells, enriched by Percoll gradient centrifugation to approximately 90% mitotic cells, responded to E2 added immediately following selection; the total E2‐treated population traversed the cycle faster and reached S phase approximately 4 hr earlier than cells not exposed to E2. When E2 was added during the last hour of synchronization (ie, at late G2 or G2/M), or for 1 hr during mitotic cell enrichment, a mixed response occurred: a small portion had an accelerated G1 exit, while the majority of cells behaved the same as controls not incubated with E2. When E2 addition was delayed until 2 hr, 7 hr, or 12 hr following cell selection, to allow many early G1 phase cells to miss E2 exposure, the response to E2 was again mixed. When E2 was added during the 16 hr of nocodazole synchronization, when cells were largely at S or possibly at early G2, it inhibited entry into S phase. The E2‐induced increase or decrease of S phase cells in the nocodazole experiments also showed corresponding changes in mitotic index and cell number. These results showed that (1) the early G1 phase and possibly the G2/M phase are sensitive to E2 stimulation, late G1, G1/S, or G2 are refractory; (2) the E2 stimulation of cell proliferation is due primarily to an increased proportion of G1 cells that traverse the cell cycle and a shortened G1 period, (3) E2 does not facilitate faster cell division; and (4) estrogen‐induced cell proliferation or G1/S transition occurs only when very early G1 phase cells are exposed to estrogen. These results are consistent with the constant transition probability hypothesis, that is, E2 alters the probability of cells entering into DNA synthesis without significantly affecting the duration of other cell cycle phases. Results from this study provide new information for further studies aimed at elucidating E2‐modulated Gl events related to tumor growth.
Expression of coronavirus E1 and rotavirus VP10 membrane proteins from synthetic RNA
Authors: Armstrong John , McCrae Malcolm , Colman Alan 
Published: December 2004

AbstractSome viruses acquire their envelopes by budding through internal membranes of their host cell. We have expressed the cloned cDNA for glycoproteins from two such viruses, the E1 protein of coronavirus, which buds in the Golgi region, and VP10 protein of rotavirus, which assembles in the endoplasmic reticulum. Messenger RNA was prepared from both cDNAs by using SP6 polymerase and either translated in vitro or injected into cultured CV1 cells or Xenopus oocytes. In CV1 cells, the El protein was localised to the Golgi region and VP10 protein to the endoplasmic reticulum. In Xenopus oocytes, the E1 protein acquired post‐translational modifications indistinguishable from the sialylated, O‐linked sugars found on viral protein, while the VP10 protein acquired endoglycosidase‐H‐sensitive N‐linked sugars, consistent with their localisation to the Golgi complex and endoplasmic reticulum, respectively. Thus the two proteins provide models with which to study targeting to each of these intracellular compartments. When the RNAs were expressed in matured, meiotic oocytes, the VP10 protein was modified as before, but the E1 protein was processed to a much lesser extent than in interphase oocytes, consistent with a cessation of vesicular transport during cell division.
Regulation of sperm flagellar motility by calcium and cAMP‐dependent phosphorylation
Authors: Brokaw Charles J. 
Published: December 2004

AbstractThere is substantial evidence that cAMP‐dependent phosphorylation is involved in the activation of motility of spermatozoa as they are released from storage in the male reproductive tract. This evidence includes observations that in vivo activation of motility can be inhibited by protein kinase inhibitors, can be reversed by protein phosphatase treatment of demembranated spermatozoa, and is associated with phosphorylation of sperm proteins, and observations that spermatozoa that have not been activated in vivo can be activated in vitro by cAMP‐dependent phosphorylation. Activation in vivo can often be triggered by conditions that increase intracellular pH, but the relevance of this to in vivo activation under natural conditions and the steps between pH increase and cAMP increase have not been fully established. The relationships between changes in the protein substrates for cAMP‐dependent phosphorylation and changes in axonemal function are still unknown.Sperm chemotaxis to egg secretions is widespread; in the sea urchin Arbacia, the egg jelly peptide resact has been identified as a chemoattractant. Response to chemoatlractants involves changes in assymmetry of flagellar bending waves, and similar changes in asymmetry can be produced in vitro by increases in [Ca++]. Temporal changes in resact receptor occupancy might lead to transient changes in intracellular [Ca++] and the asymmetry of flagellar bending, but many links in this hypothetical sequence remain to be established.Both of these signalling systems offer immediate opportunities for investigations of biochemical pathways leading to easily assayable biological responses. However, complications resulting from interactions between these two systems need to be considered.
Membrane association of collagenase stimulatory factor(s) from B‐16 melanoma cells
Authors: Biswas Chitra , Nugent Matthew A. 
Published: December 2004

AbstractPast studies have shown that contact between tumor cells and fibroblasts results in stimulation of collagenase production by the fibroblasts. Membrane fractions prepared by differential centrifugation of sonicated B‐16 melanoma cells were shown here to contain a collagenase stimulatory factor(s) (CSF). Trypsin treatment of intact B‐16 cells prior to membrane fractionation led to loss of 90% of the total activity, indicating that CSF is localized on the outer surface of the cells. Stimulation of fibroblast collegenase production was also observed with dialyzed octylglucoside extracts of the B‐16 membranes. Additional of exogeneous lipid, ie, a mixture of phosphatidylcholine and phosphatidylserine, to the detergent extract of the membranes followed by dialysis and centrifugation at 100,000g resulted in 80% recovery of the factor activity in the pellet containing reconstituted lipid vesicles. Fractionation of tritium‐labeled, reconstituted lipid vesicles on a Sephacryl S‐300 column revealed that the collagenase stimulatory factor coeluted with the radioactive lipid vesicles. The fractionated lipid vesicles lost stimulatory activity completely after trypsin treatment or heating at 65°C, indicating that the factor is a protein.
Identification of androgen response elements in mouse mammary tumour virus and the rat prostate C3 gene
Authors: Parker Malcolm G. , Webb Paul , Needham Maurice , White Roger , Ham Jonathan 
Published: December 2004

AbstractThe effect of steroid hormones on mouse mammary tumour virus (MMTV) promoter activity has been analysed in the breast cancer cell lines ZR‐75‐1 and T‐47D 11. Both androgens and progestins were stimulatory in both cell lines, whereas glucocorticoids were stimulatory in only ZR‐75 cells. A glucocorticoid response was restored to T‐47D cells by the transient expression of a glucocorticoid receptor expression vector, suggesting that endogenous receptors were limiting. The effect of androgen was mediated by a region of MMTV previously shown to be essential for glucocorticoid and progestin responses between nucleotides −201 and −69 upstream of the cap site.Similarly, the effect of androgens on the rat prostate C3(l) gene promoter was analysed in the rat prostate cell line T5. Although these cells had functional androgen receptors which could stimulate MMTV‐promoter activity, they were incapable of interacting with androgen response elements to stimulate transcription from the C3(l) promoter
Pro‐opiomelanocortin gene: A model for negative regulation of transcription by glucocorticoids
Authors: Drouin Jacques , Charron Jean , Gagner Jean‐Pierre , Jeannotte Lucie , Nemer Mona , Plante Richard K. , Wrange Örjan 
Published: December 2004

AbstractThe gene encoding pro‐opiomelanocortin (POMC) offers an interesting model system to study negative control of transcription in eucaryotes. Indeed, glucocorticoid hormones specifically inhibit transcription of the POMC gene in the anterior pituitary. The POMC gene is predominantly expressed in the anterior and intermediate lobes of the pituitary. However, only anterior pituitary POMC transcription is inhibited by glucocorticoids and stimulated by corticotropino‐releasing hormone (CRH). Rat POMC promoter sequences required for anterior pituitary‐specific expression were localized between positions −480 and −34 base pairs (bp) by DNA‐mediated gene transfer into the POMC‐expressing tumor cells, AtT‐20. These POMC promoter sequences also confer glucocorticoid inhibition of transcription. While two of the six in vitro binding sites for purified glucocorticoid receptor identified in the rat POMC gene are within these sequences, only one is required for glucocorticoid inhibition; this binding site is located at position −63 bp in the promoter and overlaps a putative CCA AT box sequence. The DNA sequence of the POMC −63 bp receptor binding site is homologous to receptor binding sites identified in the glucocorticoid responsive element (GRE) of glucocorticoid‐inducible genes. However, DNA sequence divergencies between these sites, in particular within the conserved hexanucleotide sequence 5′‐TGTYCT‐3′, may be involved in their opposite transcriptional activity. Alternatively, binding of the receptor in the promoter proximal region of the POMC gene may inhibit transcription by a hormone‐dependent represser mechanism.
Characterization of a hepatic proliferation inhibitor (HPI): Effect of HPI on the growth of normal liver cells—comparison with transforming growth factor beta
Authors: Huggett Anthony C. , Krutzsch Henry C. , Thorgeirsson Snorri S. 
Published: December 2004

AbstractImprovements in the purification of a hepatic proliferation inhibitor (HPI) from adult rat liver have yielded a product that has an inhibitory activity 1,000‐fold greater than previously reported. The growth inhibitory activity, which could be eluted from SDS‐PAGE at 17–19 kilodaltons (kD), was compared to that of transforming growth factor beta (TGF‐β). The ID50 of the HPI preparation in Fischer rat liver epithelial cells was 50 pg/ml (2.5 pM) compared to a value of 260 pg/ml (10.4 pM) obtained for pure human TGF‐β. Both inhibitors also modulated the stimulation of DNA synthesis in primary hepatocytes by either epidermal growth factor or a growth stimulatory activity prepared from serum of hepatectomized rats. The ID50S of HPI and TGF‐β in these cells were 250 pg/ml and 40 pg/ml, respectively. In contrast to TGF‐β the growth inhibitory activity of HPI was unaltered in the presence of an antibody raised against TGF‐β. The possible mechanism of action of HPI is discussed.
Uncoupling of translocation across microsomal membranes from biosynthesis of influenza virus hemagglutinin
Authors: Chao Chuck C.‐K. , Bird Phil 
Published: December 2004

AbstractThis communication presents our recent studies on the biosynthesis of influenza virus hemagglutinin (HA) in a mammalian‐cell‐free system and its translocation across microsomal membranes. RNAs coding for wild‐type (full‐length) and mutant (truncated) forms of HA were generated by in vitro transcription by using bacteriophage T7 DNA‐dependent RNA polymerase. These RNAs were translated in a rabbit reticulocyte system that was supplemented with dog pancreas membranes, either before translation was initiated or after it had been artificially terminated with the antibiotic cycloheximide. All forms of HA could be cotranslationally translocated. However, only truncated molecules (83% of full length) could translocate after protein synthesis had been terminated. Posttranslational translocation was dependent on the presence of a functional N‐terminal signal sequence and occurred only in the presence of ribosomes. The molecular mechanism of protein targeting and translocation across the membrane of the endoplasmic reticulum is discussed based on the signal hypothesis.
Invasive activity and chemotactic response to growth factors by Kaposi's sarcoma cells
Authors: Albini Adrians , Mitchell Charles D. , Thompson Erik W. , Seeman Ruth , Martin George R. , Wittek Alec E. , Quinnan Gerald V. 
Published: December 2004

AbstractKaposi's sarcoma (KS) is a relatively low grade neoplasm, classically occurring in the skin of elderly men. A more virulent and invasive form of Kaposi's sarcoma has been described in patients with acquired immune deficiency syndrome (AIDS). The origin and identification of the tumor cells in these lesions is controversial. Here we have studied the behavior of cells derived from KS lesions in an in vitro assay which measures the ability of cells to invade through a reconstituted basement membrane. In agreement with previous work, KS cells obtained under selective culture conditions were invasive showing activity comparable to that of malignant tumor cells. Normal fibroblasts, smooth muscle cells, and endothelial cells did not demonstrate invasive behavior under the same experimental conditions. To characterize further the nature of the KS cells we tested the chemotactic response of cells from the most invasive line to a variety of growth factors and compared their response to those of fibroblasts, smooth muscle, and endothelial cells. These studies suggest that normal cells respond to a unique repertoire of chemotactic factors. The chemotactic response of the KS cells most closely resembled that of smooth muscle cells and was quite distinct from endothelial cells. These results indicate that the KS‐derived cultures contain invasive cells with a smooth muscle cell‐like phenotype.
Aspects of signal transduction in stimulus exocytosis‐coupling in Paramecium
Authors: Satir Birgit H. , Busch Gerald , Vuoso Alice , Murtaugh Timothy J. 
Published: December 2004

AbstractThis paper deals with the detailed mechanisms of signal transduction that lead to exocytosis during regulative secretion induced by specific secretagogues in a eukaryotic cell, Paramecium tetraurelia. There are at least three cellular compartments involved in the process: (I) the plasma membrane, which contains secretagogue receptors and other transmembrane proteins, (II) the cytoplasm, particularly in the region between the cell and secretory vesicle membranes, where molecules may influence interactions of the membranes, and (III) the secretory vesicle itself.The ciliated protozoan Paramecium tetraurelia is very well suited for the study of signal transduction events associated with exocystosis because this eukaryotic cell contains thousands of docked secretory vesicles (trychocysts) below the cell membrane which can be induced to release synchronously when trioggered with secretagogue. This ensures a high signal‐to‐noise ratio for events associated with this process. Upon release the trichocyst membrane fuses with the cell membrane fuses with the cell membrane and the trichocyst content undergoes a Ca2+‐dependent irreversible expansion. Secretory mutants are available which are blocked at different points in the signal transduction pathway.Aspects of the three components mentioned above that will be discussed here include (a) the properties of the vesicle content, its pH, and its membrane; (b) the role of phosphorylation/dephosphorylation of a cytosolic 63‐kilodalton (kDa)Mr protein in membrane fusion; and (c) how influx of extracellular Ca2+ required for exocytosis may take place via exocytic Ca2+ channels which may be associated with specific membrane microdomains (fusion rosettes).