High molecular weight,cell surface-associated glycoprotein (fibronectin) lost in maglinant transformation

Fibronectin is a polymorphic glycoprotein found in blood and tissues of vertebrates and in cultures of adherent vertebrate cells. There are several forms of fibronectin is composed of two high molecular weight subunits held together by forms found in tissues and on and around the surfaces of cultured cells. Soluble fibronectin is composed of two high molecular weight subunits held together by disulfide bonds. Insoluble fibronectin may be covalently cross-linked in larger complexes. Fibronectin has affinities for collagen, fibrin, heparin, and cell surfaces. in culture, fibronectin in growth medium may mediate attachment of cells to substratum, and fibronectin synthesized by cells may mediate adhesion to substratum. The widespread occurrence of fibronectin in basal lamina indicates that many different cell types in vivo abut against a fibronectin-containing matrix. Cultured transformed cells usually lack cell-surface fibronectin, also called large, external transformation-sensitive (LETS) protein. The failure of transformed cells to synthesize or bind fibronectin is paralleled (at least in some systems) by failures to synthesize or bind collagen and proteoglycans. Abnormal synthesis of fibronectin and other matrix components and abnormal interactions with the tissue matrix may account for several phenotypic characteristics of transformed cultutred cells and for some of the malignant behavior of neoplastic cells in vivo.     

Friend小鼠白血病病毒（Fr.MuLV）对新生金地鼠的感染性研究

一般认为,Ｆｒｉｅｎｄ小鼠白血病病毒（Ｆｒｉｅｎｄｍｕｒｉｎｅｌｅｕｋｅｍｉａｖｉｒｕｓ：Ｆｒ．ＭｕＬＶ）属同种嗜性白血病病毒（ｅｃｏｔｒｏｐｉｃＭｕＬＶ）,只感染小鼠或大鼠,分别引起小鼠发生白血病和大鼠发生呆小病;但对小鼠或大鼠以外的动物（包括人等）无感染性。为进一步探讨该病毒的感染范围,本文进行了该病毒对金地鼠的感染性研究。结果表明,该病毒可在新生金地鼠体内增殖,引起新生金地鼠出现体重减轻、脾萎缩、并发早死等呆小病样症状,病理学检查发现接种该病毒后新生金地鼠的脾脏、胸腺呈明显的病理变化。提示Ｆｒ．ＭｕＬＶ对新生金地鼠具有感染作用,并产生病毒增殖和引起呆小病样症状,为阐明Ｆｒ．ＭｕＬＶ的感染范围和致病作用提供了实验依据。     

Preliminary evidence for novel fucose-containing lipids in normal and murine sarcoma virus-transformed rat cells.

Two fucose-containing lipids with long chain base as their apolar moiety are described. These compounds were shown to possess a free amino group and a net positive charge by derivatization with specific amino reagents and by cation exchange column chromatography. The presence of long chain base was demonstrated by periodate-NaB[³H]₄ treatment to yield long chain alcohol and by [³H]-dansylation followed by hydrolysis to yield dansyl-long chain base.     

Potential identification of chemical carcinogens in a viral transformation system

Chemical carcinogens from several diverse chemical classes i.e.; aromatic amines, polycyclic hydrocarbons, nitrosamines, hormonal derivatives, metals and direct alkylating agents cause a 6.2–60.5-fold increase in the frequency of murine sarcoma virus (MSV)-induced transformation in a normal rat kidney (NRK) cell system. Exogenous metabolic activation with a rat liver S-9 homogenate is required for expression of this activity by procarcinogens. Non-carcinogenic analogs of these compounds fail to cause significant increases in the transformation frequency either with or without prior metabolic activation. Iododeoxyuridine, a mutagen also does not cause enhancement of transformation. This system may serve as the basis for a rapid and quantifiable means of identifying chemical carcinogens while introducing a new model for the understanding of the interactions between oncorna-viruses and chemical carcinogens.     

Transfer ribonucleic acid nucleotidyltransferase activity in virions of type-C viruses.

A nucleotidyltransferase activity has been found associated with a number of mammalian and avian oncornaviruses. This activity catalyzes the incorporation of adenosine monophosphate and cytosine monophosphate into acid insoluble forms. The transferase activity from Rauscher murine leukemia virus has been characterized. The endogenous reaction is stimulated by various tRNAs particularly the 4S RNA isolated from Rauscher leukemia virus, whereas other RNAs have no effect. The product of the reaction is alkali and RNase sensitive, insensitive to DNase, and its size is similar to tRNA. Finally, the terminal nucleotide analysis of the product of the reaction indicates the presence of a CCA terminus. The properties of the activity found in the type-C viruses are in accord with those of known tRNA nucleotidyltransferases from other sources.     

Deoxyglucose uptake by mouse astrocytes: Effects of temperature and retrovirus infection

Deoxyglucose uptake by FVB/N mouse astrocytes was studied before and after infecton by tsl retrovirus which causes a neurodegenerative disease in mice similar to HIV-1 encephalopathy in man. The Michaelis-Menten kinetic parameters, Km and Vmax, of 2-deoxy-D-glucose uptake by brain and cerebellar astrocytes were measured following culture at 34°C where tsl retrovirus replicates optimally, and at 37°C. Compared to astrocytes cultured at 37°C, astrocytes cultured at 34°C had increased Km and decreased deoxyglucose uptake despite increased or unchanged Vmax. Following ts1 retrovirus infection, brain astrocyte deoxyglucose uptake doubled [132%] associated with decreased Km but unchanged Vmax, whereas cerebellar astrocyte deoxyglucose uptake doubled [102%] associated with increased Vmax but unchanged Km. These observations of altered deoxyglucose uptake kinetic parameters following retrovirus infection indicate different neurochemical mechanisms for the regional variation in deoxyglucose uptake observed following retrovirus infection of the CNS in vivo.     

Effects of aphidicolin on retrovirus DNA synthesis in vivo

Renaturation of $\text{Aequorea}$ green-fluorescent protein (A-GFP) was achieved for the first time following denaturation in guanidine-HCl or acid. Denaturation was accompanied by the concerted loss of visible fluorescence, alteration of absorption characteristics, and large negative deflection of CD signal in the far UV. Dialysis of a guanidine-denatured sample at pH 8 resulted in 64% renaturation (return to native absorption) and neutralization of an acid-denatured sample restored 90% of the native absorption. Renatured GFP is highly fluorescent and indistinguishable from native GFP with respect to the shape of excitation and emission spectra. Both native and denatured proteins exhibit resistance to trypsin hydrolysis and have identically broad pH and heat stability profiles, all of which suggest full renaturation.     

Murine leukemia virus clearance by flocculation and microfiltration

Clearance of murine leukemia virus from CHO cell suspensions by flocculation and microfiltration was investigated. Murine leukemia virus is a retrovirus that is recommended by the U.S. Food and Drug Administration for validating clearance of retrovirus-like particles. Due to biosafety considerations, an amphotropic murine leukemia virus vector (A-MLV) that is incapable of self-replication was used. Further, A-MLV is incapable of infecting CHO cells, thus ensuring that infection of the CHO cells in the feed did not result in a reduced virus titer in the permeate. The virus vector contains the gene for the enhanced green fluorescent protein (EGFP) to facilitate assaying for infectious virus particles. The virus particles are 80-130 nm in size. The feed streams were flocculated using a cationic polyelectrolyte. Microfiltration was conducted using 0.1 and 0.65 microm pore size hollow fiber membranes. The level of virus clearance in the permeate was determined. For the 0.1 microm pore size membranes a 1,000-fold reduction in the virus titer in the permeate was observed for feed streams consisting of A-MLV, A-MLV plus flocculant, A-MLV plus CHO cells, and A-MLV plus flocculant and CHO cells. While the flocculant had little effect on the level of virus clearance in the permeate for 0.1 microm pore size membranes, it did lead to higher permeate fluxes for the CHO cell feed streams. Virus clearance experiments conducted with 0.65 microm pore size membranes indicate little clearance of A-MLV from the permeate in the absence of flocculant. However, in the presence of flocculant the level of virus clearance in the permeate was similar to that observed for 0.1 microm pore size membranes. The results obtained here indicate that significant clearance of A-MLV is possible during tangential flow microfiltration. Addition of a flocculant is essential if the membrane pore size is greater than the diameter of the virus particles. Flocculation of the feed stream leads to an increase in the permeate flux.     

Delimitation of a DNA sequence which confers inducibility by glucocorticoid hormones

A chimeric long terminal repeat-thymidine kinase (LTR-tk) gene has been used to define the sequence requirements for glucocorticoid induction of gene expression. The original LTR-tk gene contains an entire mouse mammary tumor virus (MMTV) LTR preceding the tk gene. This gene can be expressed in a hormone-responsive fashion upon transfection into L tk--cells to produce a chimeric LTR-tk mRNA. Stepwise deletion of nucleotide sequences 5' of the viral RNA initiation site revealed that 202 nucleotides upstream of the viral cap site are sufficient for the hormonal regulation. Deletion of 5' sequences up to 59 nucleotides upstream of the viral cap site abolished RNA initiation in the LTR and hormonal induction.     

Characterization of a β-nerve growth factor expression vector for mammalian cells

A mammalian expression vector that directs expression of murine β-nerve growth factor (β-NGF) from a murine sarcoma virus long terminal repeat (LTR) promoter element was constructed and characterized. The vector, designated pLTRSNGF, was stably transfected into murine L-cells, and β-NGF mRNA and protein levels were quantified and compared to endogenous levels in control L-cells. Transfection of pLTRSNGF resulted in an approximate doubling of both β-NGF mRNA and mature β-NGF protein secreted into the media. Transfection of pLTRSNGF into rat PC 12 cells resulted in colonies of autocrine-differentiating cells that extended dense networks of neurites in the absence of added NGF, indicating that the β-NGF produced from the vector is biologically active.     

Lysine tRNA is the predominant tRNA in murine mammary tumor virus

The method of aminoacylation and subsequent identification of the esterified amino acids was used to characterize the transfer RNAs in murine mammary tumor virus. Lysine tRNA was the major tRNA in both “free” 4S RNA and “7OS-associated” 4S RNA in virus derived from either tissue culture or mouse milk.     

Isolation and characterization of phosphoprotein pp 105 from simian virus 40-transformed mouse fibroblasts

Investigation of the cellular distribution of a 105 kDa phosphoprotein (pp 105) in transformed mouse fibroblasts, showed that only a minor amount was located on the surface of logarithmically grown suspension cells. More than 90% of total pp 105 was contained in the cytosolic fracion representing about 0.2% of total cytosolic proteins. Surface and cytosolic pp 105 had identical phosphopeptide patterns. Cytosolic pp 105 was highly purified by ammonium sulfate precipitation followed by three chromatographic steps and gel electrophoresis. The purified pp 105 was capable of weak autophosphorylation. In the stationary growth phase of suspension cells, the amount of pp 105 detectable by endogenous phosphorylation was only 10–15% of that observed during logarithmic growth. pp 105 was also detected in normal mouse tissue and its distribution determined.