bluuebunny/crossref_metadata_2025 huggingface dataset:
Production of erythroid-potentiating activity by a human T-lymphoblast cell line.
Authors: Golde D W , Bersch N , Quan S G , Lusis A J
Published: May 2006

We derived a human T-lymphoblast cell line (Mo) that constitutively elaborates certain lymphokines. The Mo cells produce a colony-stimulating factor necessary for the growth of human granulocyte-monocyte precursors in vitro as well as an erythroid-potentiating activity (EPA) that enhances the proliferation of human erythroid progenitors in vitro. In the presence of serum, the EPA in Mo-conditioned medium stimulated the growth of small and large erythroid colonies almost 2-fold. EPA was also produced in serum-free medium, and, when assayed in serum-free cultures of human erythroid progenitors, it stimulated colony growth about 3-fold. The EPA produced by the Mo cell line did not stimulate normal murine erythroid progenitors (CFU-E) or Friend erythroleukemia cell growth in vitro. EPA was inactivated by protease treatment but was remarkably heat stable, with most of the activity recovered after boiling for 15 min. Preliminary biochemical characterization suggests that EPA is an acidic glycoprotein with molecular weight approximately 45,000. EPA is clearly separable from colony-stimulating factor on the basis of heat stability and gel-filtration chromatography. The present observations provide strong support for the concept that activated T cells produce humoral factors important in the regulation of erythropoiesis. The availability of a cell line producing human EPA should facilitate the characterization of the protein and permit definitive studies of its biologic effects.
Generation of oncogenic mouse type C viruses: in vitro selection of carcinoma-inducing variants.
Authors: Rapp U R , Todaro G J
Published: May 2006

Type C retroviruses are endogenous in most vertebrate species. These viruses generally are of low pathogenicity in their natural hosts. Variants that contain cell-derived transforming genes have been isolated infrequently in the past upon continued passage in vivo. We report here a procedure that allows the isolation of new mouse leukemia-, sarcoma-, and carcinoma-inducing type C viruses entirely in cell culture. The viruses generated after passage in chemically transformed mouse cells and selectron in epithelial mink lung cells produced pulmonary adenomas and adenocarcinomas and also ovarian carcinomas. Viruses with cell-transforming capacity, as determined by their ability to induce normal murine or mink cells to form progressively growing colonies in soft agar, appeared only transiently (2-4 days) after acute infection of "spontaneously" or chemically transformed mouse cells by nontransforming helper viruses. These transiently appearing transforming viruses can be "captured" by selecting the cells that respond to their newly acquired growth-stimulating ability. This system may lend itself to the systematic isolation of tissue-specific transforming functions from any cell that can be efficiently infected by retroviruses.
Isolation and characterization of the pigment-protein complexes of Rhodopseudomonas sphaeroides by lithium dodecyl sulfate/polyacrylamide gel electrophoresis.
Authors: Broglie R M , Hunter C N , Delepelaire P , Niederman R A , Chua N H , Clayton R K
Published: May 2006

When purified photosynthetic membranes from Rhodopseudomonas sphaeroides were treated with lithium dodecyl sulfate and subjected to polyacrylamide gel electrophoresis at 4 degrees C, up to 11 pigment-protein complexes were resolved. Absorption spectra revealed that the smallest complex contained reaction center pigments and the others contained the antenna components B850 and B875 in various proportions. Of these antenna complexes, the largest was almost entirely B850 and the smallest contained only B875. After solubilization at 100 degrees C and electrophoresis on polyacrylamide gradient gels, the B850 complex gave rise to two polypeptide components migrating with apparent Mr of 10,000 and 8000, whereas with the B875 complex, two components were observed with apparent Mr of 12,000 and 8000. The reaction center complex gave rise to only the 24 and 21 kilodalton polypeptide subunits. Fluorescence emission spectra showed maxima at 872 and 902 nm for B850 and B875, respectively. Analyses of bacteriochlorophyll a and carotenoids indicated that, in the B875 complex, two molecules of each of these pigments are associated with the two polypeptides. The associations of B850 and B875 in large and small complexes obtained by lithium dodecyl sulfate treatment are consistent with models of their organization within the membrane.
Geminate recombination of O2 and hemoglobin.
Authors: Chernoff D A , Hochstrasser R M , Steele A W
Published: May 2006

The photolysis of HbO2 and HbCO has been studied by measuring transient absorption spectra in the Soret region after excitation with picosecond pulses at 530 nm. Dissociation occurred promptly in both cases, followed (for HbO2) by geminate recombination of ca. 40% of the photodissociated O2 with a lifetime of 200 +/- 70 psec (25 degrees C). No recombination of Hb + CO was observed up to 1200 psec after photolysis. The HbO2 and HbCO photoproduct spectra were broader, weaker, and red-shifted in comparison to the spectrum of stable Hb and Gibson's fast-reacting form, Hb. For HbO2 the spectrum was initially much broader to longer wavelengths but relaxed to a constant shape within 90 psec, whereas for HbCO there was no spectral evolution. The photophysics is analyzed by considering the effect of spin constraints as well as spin--orbit coupling and orbital correlation among the various electronic states of liganded and deoxy hemoglobins. The small quantum yield of HbO2 dissociation is not primarily due to rebinding but rather to electronic relaxation to nonreactive states.
Soliton dynamics in polyacetylene
Authors: Su W. P. , Schrieffer J. R.
Published: May 2006

The equations of motion of the coupled electron-phonon system are integrated in real time for the model of polyacetylene recently proposed. To illustrate the physical behavior of this nonlinear system we consider the time evolution starting from three physically relevant configurations: (
i
) end generated soliton, (
ii
) electron-hole pair generation of a charged soliton-antisoliton pair, and (
iii
) the dressing of an injected electron. The calculations show that the system relaxes within a time of order 10
-13
sec, converting excited electron-hole pairs into soliton-antisoliton pairs.
Sendai virus utilizes specific sialyloligosaccharides as host cell receptor determinants.
Authors: Markwell M A , Paulson J C
Published: May 2006

Purified sialyltransferases (CMP-N-acetyl-neuraminate:D-galactosyl-glycoprotein N-acetylneuraminyl-transferase, EC 2.4.99.1) in conjunction with neuraminidase (acylneuraminyl hydrolase, EC 3.2.1.18) were used to produce cell surface sialyloligosaccharides of defined sequence to investigate their role in paramyxovirus infection of host cells. Infection of Madin-Darby bovine kidney cells by Sendai virus was monitored by hemagglutination titer of the virus produced and by changes in morphological characteristics. By either criterion, treatment of the cells with Vibrio cholerae neuraminidase to remove cell surface sialic acids rendered them resistant to infection by Sendai virus. Endogenous replacement of receptors by the cell occurred slowly but supported maximal levels of infection within 6 hr. In contrast, sialylation during a 20-min incubation with CMP-sialic acid and beta-galactoside alpha 2,3-sialytransferase restored full susceptibility to infection. This enzyme elaborates the NeuAc alpha 2,3Gal beta 1,3GalNAc (NeuAc, N-acetylneuraminic acid) sequence on glycoproteins and glycolipids. No restoration of infectivity was observed when neuraminidase-treated cells were sialylated by using beta-galactoside alpha 2,6-sialytransferase, which elaborates the NeuAc-alpha 2,6Gal beta 1,4GlcNAc sequence. These results suggest that sialyloligosaccharide receptor determinants of defined sequence are required for Sendai virus infection of host cells.
Evidence for presence of an internal thiolester bond in third component of human complement.
Authors: Tack B F , Harrison R A , Janatova J , Thomas M L , Prahl J W
Published: May 2006

Treatment of the third component of human complement (C3) with methylamine results in a loss of hemolytic function and the appearance of a thiol group. Studies with [14C]methylamine have indicatd a stoichiometric and covalent reaction with the native protein. Hydrazine-inactivated C3 and C3b prepared with bovine trypsin were unreactive with [14C]-methylamine. Alkylation experiments with [1-14C]iodoacetamide have further established a 1:1 correspondence between methylamine incorporation and expression of the reactive thiol. Autoradiographic analyses of [14C]methylamine-treated C3 and methylamine-inactivated [1-14C]carboxyamidomethylated C3 after NaDodSO4/polyacrylamide gel electrophoresis have shown a specific incorporation of each radiolabel into the alpha polypeptide chain. [14C]Methylamine-treated C3 was immobilized on Sepharose 4B by reaction of the protein thiol with a mixed disulfide. Digestion with bovine trypsin in 4 M urea released 96% of the bound absorbance (at 280 nm) units; the radiolabel remained associated with the Sepharose beads. Peptide material labeled with 14C was eluted with dithiothreitol, carboxymethylated with [3H]iodoacetic acid, and chromatographed on Sephadex G-75. On Edman degradation S-[3H]carboxymethylcysteine was released at step 9 and gamma-glutamyl[14C]-methylamide was released at step 12. We interpret these data to indicate the presence of an internal thiolester bond in native C3. In addition, evidence is presented for an identical reactive site in alpha 2-macroglobulin.
Isolation and partial characterization of the Drosophila alcohol dehydrogenase gene.
Authors: Goldberg D A
Published: May 2006

The alcohol dehydrogenase (ADH; alcohol: NAD+ oxidoreductase, EC 1.1.1.1) gene (Adh) of Drosophila melanogaster was isolated by utilizing a mutant strain in which the Adh locus is deleted. Adult RNA from wild-type flies was enriched in ADH sequences by gel electrophoresis and then used to prepare labeled cDNA for screening a bacteriophage lambda library of genomic Drosophila DNA. Of the clones that hybridized in the initial screen, one clone was identified that hybridized with labeled cDNA prepared from a wild-type Drosophila strain but did not hybridize with cDNA prepared from an Adh deletion strain. This clone was shown to contain ADH structural gene sequences by three criteria: in situ hybridization, in vitro translation of mRNA selected by hybridization to the cloned DNA, and comparison of the ADH protein sequence with a nucleotide sequence derived from the cloned DNA. Comparison of the restriction site maps from clones of three different wild-type Drosophila strains revealed the presence of a 200-nucleotide sequence in one strain that was absent from the other two strains. The ADH mRNA sequences were located within the cloned DNA by hybridization mapping experiments. Two intervening sequences were identified within Adh by S1 nuclease mapping experiments.
Isopentenyladenine as a mediator of mevalonate-regulated DNA replication.
Authors: Huneeus V Q , Wiley M H , Siperstein M D
Published: May 2006

The mechanism by which the cholesterol precursor, mevalonate, regulates S-phase DNA replication was examined in synchronized BHK cells. As previously demonstrated by this laboratory, blocking 3-hydroxy-3-methylglutaryl-CoA reductase [mevalonate:NADP+ oxidoreductase (CoA-acylating), EC 1.1.1.34] with the competitive inhibitor compactin suppresses DNA synthesis specifically during the S phase of the cell cycle. In the present study, known mevalonate derivatives were examined as possible mediators by which mevalonate controls DNA replication. Of the compounds studied, only isopentenyladenine and its 4'-hydroxylated analogue, zeatin, could substitute for mevalonate in restoring DNA replication in compactin-blocked cells. Moreover, these two derivatives proved to be at least 100 times more active than mevalonate, and both restored DNA relication to normal within 15 min of their being added to the medium. In addition, isopentenyladenine, like mevalonate, stimulated DNA synthesis specifically during the S phase of the cell cycle. Isopentenyladenine also reversed the inhibition of DNA synthesis caused by nalidixic acid, an antibiotic that does not inhibit cholesterol synthesis. These findings indicate that isopentenyladenine or a closely related derivative may mediate the regulatory role of mevalonate in DNA replication and suggest that such isoprenes may act upon DNA replication at a site common to that inhibited by nalidixic acid.
Progesterone inhibition of uterine nuclear estrogen receptor: dependence on RNA and protein synthesis.
Authors: Evans R W , Leavitt W W
Published: May 2006

Progesterone treatment was found previously to reduce nuclear estrogen receptor (Re) rapidly (2-4 hr) in the hamster uterus in vivo. The present study was done to determine whether this inhibitory effect of progesterone on uterine nuclear Re could be demonstrated in vitro ad whether progesterone action was dependent on RNA and protein synthesis. A uterine strip system was used in which estradiol pretretment caused cytosol Re translocation to the nucleus and increased synthesis of cytosol progesterone receptor (Rp) during a 16-hr incubation. When progesterone was added to the medium 4 hr before the end of the incubation, cytosol Rp was depleted and nuclear Re was greatly reduced. Further experiments done with actinomycin D, puromycin, and cycloheximide indicated that the progesterone-induced loss of uterine nuclear Re was dependent on RNA and protein synthesis. These results suggest that progesterone reduced nuclear Re through a mechanism involving Rp translocation and the induction of RNA and protein synthesis, a product of which is active in degrading or otherwise inactivating nuclear Re.
Magnetic field-induced increase in chlorophyll
a
delayed fluorescence of photosystem II: A 100- to 200-ns component between 4.2 and 300 K
Authors: Sonneveld Arie , Duysens Louis N. M. , Moerdijk Adri
Published: May 2006

At room temperature the delayed fluorescence (luminescence) of spinach chloroplasts, in which the acceptor Q is prereduced, consists of a component with a lifetime of 0.7 μs and a more rapid component, presumably with a lifetime of 100-200 ns and about the same integrated intensity as the 0.7- μs component. Between 4.2 and 200 K only a 100- to 200-ns luminescence component was found, with an integrated intensity appreciably larger than that at room temperature. At 77 K the 150-ns component approached 63% of saturation at roughly the same energy as the variable fluorescence of photosystem II at room temperature. At 77 K the emission spectra of prompt fluorescence but not that of the 150-ns luminescence had a preponderant additional band at about 735 nm. The 150-ns emission also occurred in the photosystem I-lacking mutant FL5 of
Chlamydomonas
. These experiments indicate that the 150-ns component originates from photosystem II. At room temperature a magnetic field of 0.22 T stimulated the 0.7-μs delayed fluorescence by about 10%. At 77 K the field-induced increase of the 150-ns component amounted to 40-50%, being responsible for the observed ≈2% increase of the total emission; the magnetic field increased the lifetime about 20%. In order to explain these phenomena a scheme for photosystem II is presented with an intermediary acceptor W between Q and the primary donor chlorophyll P-680; recombination of P-680
+
and W
-
causes the fast luminescence. The magnetic field effect on this emission is discussed in terms of the radical pair mechanism.
Metabolism of native and of lactosylated human low density lipoprotein: evidence for two pathways for catabolism of exogenous proteins in rat hepatocytes.
Authors: Attie A D , Pittman R C , Steinberg D
Published: May 2006

Human low density lipoprotein (LDL) covalently conjugated with 200-250 residues of lactose per LDL particle (Lac-LDL) was bound and rapidly taken up by the galactose-specific receptor of rat hepatocytes. Uptake of Lac-LDL was associated with inhibition of 3-hydroxy-3-methylglutaryl-CoA reductase and stimulation of cholesterol esterification. Uptake of native human LDL had no significant effects on these enzyme activities even when the rates of LDL uptake equaled those of Lac-LDL. When injected into rats, Lac-LDL was selectively removed by the liver (98% of injected dose). The hepatic subcellular distribution of simultaneously injected native 125I-labeled LDL and 131I-labeled Lac-LDL differed significantly, Lac-LDL was associated with fractions enriched in lysosomal hydrolases whereas native LDL was found predominantly in the supernatant fraction enriched in lactate dehydrogenase. Chloroquine (0.1 mM) markedly suppressed uptake of Lac-LDL by cultured rat hepatocytes (> 80%) but had only a small effect on uptake of native LDL. Leupeptin (0.625 mM) inhibited degradation of Lac-LDL more than it did degradation of native LDL. Colchicine (0.25 microM) dramatically suppressed uptake of Lac-LDL (> 70%) but did not affect native LDL uptake even at concentrations as high as 10 microM. Uptake of human LDL by rat hepatocytes occurs largely by nonspecific mechanisms, including fluid endocytosis, whereas Lac-LDL, as shown here, is taken up by a specific receptor-mediated mechanism. The results show further that native human LDL, representing an example of a protein taken up nonspecifically, is processed intracellularly by a pathway qualitatively distinct from that for Lac-LDL, an example of a protein taken up by a specific mechanism. Lac-LDL may serve as a vehicle for specifically delivering drugs, hormones, or radioactive compounds to hepatocytes for therapeutic or diagnostic purposes.
Malignancies of metastatic murine lymphosarcoma cell lines and clones correlate with decreased cell surface display of RNA tumor virus envelope glycoprotein gp70.
Authors: Reading C L , Brunson K W , Torrianni M , Nicolson G L
Published: May 2006

Variant sublines of the murine lymphosarcoma RAW117 have been derived by sequential cycles of intravenous inoculation of cells and harvesting of solid liver tumors in syngeneic BALB/c mice [Brunson K. W. & Nicolson, G. L. (1978) J. Natl. Cancer Inst. 61, 1499-1503] and also by sequential removal of lectin-reactive cells via repeated adsorption on immobilized-lectins [Reading, C. L. Belloni, P. N. & Nicolson, G. L. (1980) J. Natl. Cancer Inst. 64, 1241-1249]. These cell sublines and their clones were analyzed for abilities to form gross liver tumor metastases after injection intravenously or subcutaneously into syngeneic mice, and this response was related to certain cell surface properties including quantities of viral antigens and lectin-binding sites, exposure of specific cell surface proteins, and quantities of cell surface glycoproteins visualized in gels with 125I-labeled lectins or antibodies. Consistent differences were obtained between RAW117 sublines of low and high malignancy with respect to the amounts or exposures of cell surface glycoprotein components of Mr approximately 70,000 or 69,000 and 71,000, depending on the gel system. Competition radioimmunoassays for RNA tumor virus antigens in the RAW117 lines and clones indicated the presence of Moloney murine leukemic virus antigens gp70, p30, and p12. Enhanced malignancy and metastasis to liver was accompanied by decreases in the cellular contents of viral antigens and loss of gp70 cell surface exposure. Analysis of several clones obtained from sublines selected in vivo and in vitro for high or low malignancy confirmed the inverse relationship between metastasis and expression of gp70 in this system.
Transcription units of chicken ovalbumin gene observed after injection of cloned complete genes into Xenopus oocyte nuclei.
Authors: Trendelenburg M F , Mathis D , Oudet P
Published: May 2006

The organization of transcription of a well-characterized protein encoding gene was studied by microinjection and electron microscopy. Circular recombinant DNA molecules containing the complete chicken ovalbumin sequences (7.7 kilobases, contained in 11.5 kilobases of chicken DNA) were microinjected into germinal vesicles of living oocytes of the clawed toad Xenopus laevis, and their transcription was studied in nuclear spread preparations. Evaluation of spread chromatin showed a limited number of observed molecules transcribed in "specific" patterns--i.e., circular chromatin molecules containing transcription units approximately 2.3 micrometer long, consisting of regular series of densely packed lateral ribonucleoprotein fibrils gradually increasing in length. The appearance of these fibril gradients was similar to that of actively transcribed endogenous protein encoding genes contained in lampbrush-chromosome loops of the same nuclei and to the putative Bombyx silk fibroin transcription units. In addition, less-regular arrays of transcript fibrils were seen in some circles, including fully fibril-covered molecules, indicative of the occurrence of irregular transcriptional events. The results of this heterologous transcription experiment indicate that the transcriptional machinery of the amphibian oocyte nucleus is capable of transcribing protein encoding genes from an avian species in typical regular arrays of transcription units.
Antigen-liposome modification of target cells as a method to alter their susceptibility to lysis by cytotoxic T lymphocytes.
Authors: Hale A H , Ruebush M J , Lyles D S , Harris D T
Published: May 2006

A method of liposome modification of cell surfaces to render unsuitable target cells susceptible to lysis by anti-viral cytotoxic T lymphocytes (CTLs) is described. Liposomes containing the hemagglutinin-neuraminidase (HN) and fusion (F) glycoproteins of Sendai virus as well as purified H-2Kk cells and rendering those cells susceptible to lysis by B10. A anti-Sendai virus or anti-H-2Kk CTLs. The absence from the modifying liposomes of the HN or F proteins or H-2Kk antigens eliminated the ability of the target cells to be recognized and lysed by either effector cell population. Vesicles containing HN, H-2Kk molecules, and inactive fusion protein (Fo) were not capable of increasing the susceptibility of h-2-negative target cells to lysis. Liposomes containing inactive fusion protein were similarly unable to render H-2-positive target cells susceptible to lysis by anti-Sendai virus CTLs, suggesting that fusion of the liposomes to the cell surface is a prerequisite to lysis. It did not appear that attachement of liposomes to the cell surface was sufficient for generation of susceptible targets, however, because attachment to the cell surface was observed, as long as the HN glycoprotein was present in the liposomes. These results indicate that purified H-2Kk glycoproteins are target antigens for anti-H-2k CTLs and that B10 . A anti-Sendai virus CTLs recognize in an H-2-restricted manner the HN, F, or both glycoproteins of Sendai virus in the context of the purified H-2Kk glycoproteins. This technique of liposome modification of cell surfaces has potential applications in the examination of CTL antigen recognition and immunotherapy of many viral and neoplastic diseases.
Autoradiographic method for measuring the ischemic myocardium at risk: effects of verapamil on infarct size aftr experimental coronary artery occlusion.
Authors: DeBoer L W , Strauss H W , Kloner R A , Rude R E , Davis R F , Maroko P R , Braunwald E
Published: May 2006

Investigation of the efficacy of pharmacologic agents affecting myocardial infarct size after coronary artery occlusion is complicated by the variability of collateral flow among experimental animals which results in variability of infarct size. To overcome this difficulty, we developed an autoradiographic method to delineate the ischemic area at risk of necrosis after coronary artery occlusion and we invetigated the potential protective effect of a calcium antagonist verapamil. The left anterior descending coronary arteries of 25 barbiturate-anesthetized dogs were occluded. Thirty minutes later, highly radioactive human albumin microspheres labeled with 99mTc were injected into the left atrium. One hour after coronary artery occlusion, dogs were randomized to control or treated groups; the latter received a 0.2 mg/kg loading dose and 0.6 mg/kg per hr maintenance dose of verapamil intravenously. Eight hours after coronary artery occlusion, the dogs were killed, the hearts were excised, and the left ventricle was sectioned parallel to the atrioventricular groove; infarct size was determined planimetrically after incubation in triphenyl tetrazolium chloride. The slices were then exposed to high-speed x-ray film with image-enhancing screens. The percentage of left ventricle that was ischemic, as determined by planimetry of autoradiographs, was similar in treated and control animals (36.6 +/- 2.0% compared to 37.3 +/- 2.8%, respectively). Of the ischemic area, 92.0 +/- 4.3% was infarcted in control animals and 70.5 +/- 5.1% was infarcted in treated animals (P < 0.01). Thus, this autoradiographic method using 99mTc-labeled human albumin microspheres is useful in delineating the area of ischemia after coronary artery occlusion and in evaluating the efficacy of pharmacologic agents designed to protect ischemic myocardium. Verapamil, administered 1 hr after coronary artery occlusion, is effective in limiting infarct size.
Frequency and levels of antibodies to Epstein-Barr virus-specific DNase are elevated in patients with nasopharyngeal carcinoma.
Authors: Cheng Y C , Chen J Y , Glaser R , Henle W
Published: May 2006

Sera from healthy individuals and patients with infectious mononucleosis, Burkitt lymphoma, nasopharyngeal carcinoma, or other malignancies were examined for their capacity to neutralize Epstein-Barr virus (EBV)-induced DNase activity. Sera were found that neutralized the EBV DNase but not herpes simplex virus type 1 or type 2 DNases, and vice versa. Sera from 46 of the 49 patients with nasopharyngeal carcinoma examined (94%) neutralized > 6 units of EBV DNase per ml of serum. In contrast, only 19% of 47 patients with Burkitt lymphoma, 12% of 183 patient with other malignancies, 4% of 58 patients with infectious mononucleosis, and none of 101 healthy individuals had such levels of neutralizing activity. The neutralizing factor was found in the IgG fraction derived from nasopharyngeal carcinoma sera. There was no correlation between the concentration of these antibodie and the titers of IgG ad IgA antibodies to the EBV capsid antigen, the early antigen complex, or the EBV-associated nuclear antigen.
Complementation of arylsulfatase A in somatic hybrids of metachromatic leukodystrophy and multiple sulfatase deficiency disorder fibroblasts.
Authors: Chang P L , Davidson R G
Published: May 2006

Metachromatic leukodystrophy and multiple sulfatase deficiency disorder are severe neurodegenerative diseases inherited as separate autosomal recessive traits. Arylsulfatase A (aryl-sulfate sulfohydrolase, EC 3.1.6.1) activity is deficient in both diseases but in multiple sulfatase deficiency disorder, activities of arylsulfatases B and C and other sulfatases are also reported to be reduced. Somatic hybrid cell clones produced by fusing cultured fibroblasts from patients with these diseases were isolated by a nonselective technique based on unit-gravity sedimentation. Arylsulfatase A activity was restored in these hybrids. The complemented enzyme resembled the normal arylsulfatase A in heat stability, pH optimum, Km, electrophoretic mobility, and immunologic reactivity. Because a structurally normal enzyme can be restored in a hybrid only though intergenic complementation, these results indicate that the mutations responsible for the deficiency of arylsulfatase A activity in metachromatic leukodystrophy and multiple sulfatase deficiency disorder are nonallelic and that at least two genetic loci control the expression of arylsulfatase A activity in the human genome. Furthermore, arylsulfatase C activity was also restored to normal in the hybrids, indicating that a common sulfatase inhibitor is not the cause of the multiple sulfatse deficiency.
Cord-blood tyrosine levels in the full-term phenylketonuric fetus and the "justification hypothesis".
Authors: Scriver C R , Cole D E , Houghton S A , Levy H L , Grenier A , Laberge C
Published: May 2006

The "justification hypothesis" attributes mental retardation in phenylketonuria (PKU) to an inability of the heterozygous mother to deliver an appropriate amount of tyrosine to the PKU fetus who, in turn, is unable to correct for this deficiency because of its genetic constitution. We tested this hypothesis by measuring concentrations of tyrosine and phenylalanine in cord blood obtained at delivery from nine infants with PKU and five infants with persistent (non-PKU) hyperphenylalaninemia (PHP). For each of these specimens there were four control cord-blood specimens from infants born on the same day and, generally, in the same hospital. PKU and PHP groups were similar with respect to cord-blood tyrosine and phenylalanine values. There was no biologically significant deficiency of tyrosine in cord blood of the pooled PKU and PHP deficiency of tyrosine in cord blood of the pooled PKU and PHP groups (54 +/- 10 microM, mean +/- SD) compared with controls (61 +/- 16 microM, P = 0.13). On the other hand, phenylalanine in cord blood of the pooled PKU and PHP groups was significantly increased (144 +/- 30 microM, mean +/- SD) compared with controls (128 +/- 24, P = 0.004). The mangitude of the differences in cord-blood tyrosine and phenylalanine between control and PKU subjects are so small that it is unlikely that they have any consequences for physical and mental development. The justification hypothesis, as it pertains to blood tyrosine at term, is not upheld.
Chemical potential, Teller's theorem, and the scaling of atomic and molecular energies
Authors: March Norman H. , Parr Robert G.
Published: May 2006

For atoms and homonuclear diatomic molecules, it is argued that the electronic energies have the forms [Formula: see text] and [Formula: see text] [Formula: see text], respectively,where
Z
is the atomic number,
N
is the number of electrons, and
R
is the internuclear distance. By using the Lieb-Simon theorem that the Thomas-Fermi theory is exact in the limit of large atomic number and the Teller theorem that molecules are not bound in the Thomas-Fermi theory, it is then shown, among other results, that the electron-electron repulsion energy for neutral systems has no term in
Z
2
and that the nucleus-nucleus repulsion energy for neutral molecules is probably [unk](
Z
5/3
). For neutral atoms, it is predicted and verified that the chemical potential (electronegativity) is [unk](
Z
-1/3
) for large
Z
. Tetrahedral and octahedral molecules are briefly discussed.
Multiple mRNA species for the precursor to an adenovirus-encoded glycoprotein: identification and structure of the signal sequence.
Authors: Persson H , Jörnvall H , Zabielski J
Published: May 2006

Early region 3 of the adenovirus type 2 genome encodes three proteins with molecular weights of 16,000, 14,500, and 14,000 (E2/16, E3/14.5, and E3/14). The E3/16 protein is the precursor to the E3/19 glycoprotein and is around 1500 daltons larger than the unglycosylated E3/19O protein. The E3/14.5 and E3/14 proteins are structurally related to each other but different from E3/16. Three mRNA species were identified for E3/16; all have common 5' ends with the same spliced region but with different 3' ends. E3/14 was translated from a 13S mRNA with the same 5' structure as the E3/16 mRNA but followed by a second spliced region with a different 3' end. A partial amino acid sequence was determined for E3/16 after radioactive labeling in vitro and this sequence can be aligned with a known DNA sequence. It contains a hydrophobic signal sequence, two presumptive glycosylation sites, and a hydrophobic region close to the COOH terminus.
Biosynthesis of A,B procollagen.
Authors: Kumamoto C A , Fessler J H
Published: May 2006

Interest in cell-associated collagens led others to isolate A and B collagen chains, also known as type V collagen, from many tissues, but only after pepsin cleavage. Soluble precursors of these chains are synthesized in vitro by crop, a chicken embryo muscle tissue, and are converted by at least two processing steps from procollagens via intermediates to final forms which are large than the pepsin-derived A and B chains. Heterotrimeric, disulfide-bridged procollagen molecules corresponding to B2A exist. Components were separated by ion exchange chromatography, velocity sedimentation, and electrophoresis, and the relationships between them were established by sequential radioactive labeling and comparison of peptides generated by protease cleavage.
High-performance liquid chromatography technique for resolving multiple forms of hepatic membrane-bound cytochrome P-450.
Authors: Kotake A N , Funae Y
Published: May 2006

Solubilized hepatic microsomal cytochrome P-450 from rats was resolved into 12-different heme-containing components by ion-exchange high-performance liquid chromatography. Separation was accomplished by the utilization of Anpac ion-exchange resin. Phenobarbital (PB) and 3-methylcholanthrene (3-MC) treatment of rats resulted in selective increases or decreases of specific components observed in the elution profile of solubilized microsomes prepared from untreated rats. Treatment with polychlorinated biphenyls increased peaks that were selectively increased by PB or 3-MC treatment. Proteins collected from the major peaks induced by PB or 3-MC treatment were found to retain spectral characteristics of cytochrome P-450. The recovered cytochrome b5 and NADPH-cytochrome c reductase retained spectral properties and enzymic activity, respectively. Purified P-450b obtained from PB-treated rats was found to cochromatograph with one of the peaks selectively induced by PB treatment. These results suggested that it is possible to separate membrane-bound proteins without dramatically altering their physical properties.
Mass spectra of partial protein hydrolysates as a multiple phase check for long polypeptides deduced from DNA sequences: NH2-terminal segment of alanine tRNA synthetase.
Authors: Herlihy W C , Royal N J , Biemann K , Putney S D , Schimmel P R
Published: May 2006

A strategy has been developed for rapid and accurate determination of the amino acid sequence of large proteins, such as many of the members of the class of proteins known as aminoacyl tRNA synthetases. This strategy involves combining DNA sequencing of the gene for the protein of interest with gas chromatographic mass spectrometric identification of tetra- and pentapeptides in partial hydrolysates of the entire protein or very large fragments thereof. These peptides are matched to blocks of codons at locations scattered throughout the entire structural gene. Tetra- and pentapeptide sequences are sufficiently long that they are unlikely to be repeated in the protein sequence or to occur in an incorrect reading frame; therefore, they can be placed at unique clusters of codons on the DNA. This procedure rigorously establishes the proper phasing of the DNA throughout the entire length of the structural gene, and the protein sequence is thereby accurately read from the DNA sequence. This approach is being used to determine the amino acid sequence of EScherichia coli alanine tRNA synthetase, a protein that has approximately 900 amino acids. This paper reports the sequence of the first 165 amino acids from the NH2 terminus.
Growth of normal human glial cells in a defined medium containing platelet-derived growth factor.
Authors: Heldin C H , Wasteson A , Westermark B
Published: May 2006

DNA synthesis and cell division were measured in serum-free cultures of human normal diploid glial cells maintained in MCDB 105 medium. In growth factor-free cultures the cells remained viable but the cell number was essentially constant. Supplementation with 10 ng of epidermal growth factor or platelet-derived growth factor per ml significantly stimulated DNA synthesis and cell multiplication. Growth occurred both when cells were allowed to settle in serum-containing medium and when cells were plated in serum-free medium. In the latter type of cultures, the cell yield was improved bu incubating the cells in collagen-coated dishes. The use of a miniclone technique allowed the analysis of cell multiplication induced by platelet-derived growth factor at the clonal level and demonstrated that the growth factor induced several cell cycle rounds in a large fraction of clones. The results show that normal cells grown in a recently developed synthetic medium (MCDB 105) supplemented with pure growth factors may multiply without the addition of plasma-derived factors ("progression factors"). It is suggested that the need fo progression factors may simply depend on the composition of the synthetic nutrient medium.
Somatic cell genetic analysis of regulation of expression of 3-hydroxy-3-methylglutaryl-coenzyme A reductase.
Authors: Sinensky M , Armagast S , Mueller G , Torget R
Published: May 2006

Three new types of regulatory somatic cell mutants defective in expression of the enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase are described. They include a recessive mutant with abnormally high enzyme activity, apparently defective in degradation of the enzyme, and one, phenotypically a mevalonate auxotroph, that maintains permanently repressed levels of enzyme activity and are recessive, constitutive mutants. These mutants offer means for analyses of the mechanism of regulation of expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase in cultured fibroblasts.
Production of auto-anti-idiotypic antibody during the normal immune response: changes in the auto-anti-idiotypic antibody response and the idiotype repertoire associated with aging.
Authors: Goidl E A , Thorbecke G J , Weksler M E , Siskind G W
Published: May 2006

Hapten-augmentable plaque-forming cells (PFC) are cells whose secretion of antibody is specifically inhibited by surface-bound auto-anti-iodotype antibody that can be displaced by hapten. This study showed that the percentage of hapten-augmentable PFC present in mice during the primary response to trinitrophenylated Ficoll (TNP-F) increases with age. The data suggest that there is a relative increase in the auto-anti-idiotypic antibody response with age and therefore a greater down-regulation of antibody production. The effect of age on idiotype expression was also studied. Hapten-reversible inhibition of plaque formation was used as an assay for anti-inhibition of plaque formation was used as an assay for anti-idiotype antibody and idiotype-bearing antibody-secreting cells. Sera from aged (21- to 22-month-old) C57BL/6 mice immunized with TNP-F significantly inhibited plaque formation, in a hapten-reversible manner, by spleen cells from 81% of TNP-F-immunized aged mice. However, these sera inhibited plaque formation by cells from only 50% of similarly immunized young adult (6- to 8-week-old) mice and 20% of immature (3- to 4-week-old) syngeneic mice. Similarly, sera from TNP-F-immunized young adult or immature mice inhibited formation of plaques by cells from immunized donors of the same age as the mice from whom the serum was obtained, but only rarely inhibited plaque formation by cells from mice of other age groups. The data thus suggest that the repertoire of TNP-specific idiotypes that are produced in response to TNP-F varies with age in syngeneic mice.
Role of adenylate cyclase in immunologic release of mediators from rat mast cells: agonist and antagonist effects of purine- and ribose-modified adenosine analogs.
Authors: Holgate S T , Lewis R A , Austen K F
Published: May 2006

The initial monophasic rise in cyclic AMP beginning 5-15 sec after bridging of rat mast cell IgE-Fc receptors precedes the secretion of granule constituents, thereby implying a causal relationship. Direct evidence for a relationship between IgE-dependent transmembrane activation of adenylate cyclase and granule secretion was provided by the capacity of purine-modified (R site active) and ribose-modified (P site active) adenosine analogs, respectively, to augment and suppress mediator release while simultaneously increasing and decreasing the activity of adenylate cyclase. R site stimulation alone does not cause granule secretion but augments the rate and magnitude of IgE-Fc receptor-induced secretion, reflecting the coupled relationship of such receptors. Inhibition of adenylate cyclase at the P site attenuates the rise in cellular cyclic AMP and suppresses IgE-dependent mediator release in a parallel and superimposable dose-response fashion. Further, the relationship between the attenuation in the rise in cyclic AMP and the diminution in immunologic mediator release is linear with the regression line passing through the origin, indicating a direct relationship between the IgE-dependent activation of adenylate cyclase and preformed mediator release. Although not the only events in coupled mast cell activation--secretion, there is a sequential relationship among perturbation of IgE-Fc receptors, transmembrane activation of adenylate cyclase, elevation of cytoplasmic levels of cyclic AMP, activation of cyclic AMP-dependent protein kinase, and secretion of mast cell granules.
Synaptic endings and their postsynaptic targets in neostriatum: synaptic specializations revealed from analysis of serial sections.
Authors: Groves P M
Published: May 2006

Neostriatal tissue from rat brain was examined in ribbons of serial sections, using tissue from subjects in which the synaptic marker 5-hydroxydopamine had been injected into the lateral ventricle. A total of 440 synaptic terminals, identified by the presence of a population of vesicles, was studied, including those labeled by 5-hydroxydopamine and all those unlabeled in the surrounding neuropil. Three categories of synaptic profiles innervating neostriatal dendrites could be discerned. One category contained small rounded or slightly pleomorphic vesicles and consisted of a sample of 375 endings, 53 of which exhibited light to heavy label inside the synaptic vesicles. All of these, labeled and unlabeled, showed evidence of membrane specializations and other conventional criteria of synaptic contact, mostly in the form of asymmetric synaptic contacts with the heads of dendritic spines. A second category of 40 terminals, 3 of which were lightly labeled, contained large rounded or slightly pleomorphic vesicles and made predominantly symmetric contacts with the shafts of dendritic spines or spine-free portions of dendritic membrane. None of these synaptic endings appeared to lack synaptic contact with postsynaptic targets in the neostriatum. A third category of 25 terminals, 3 of which were labeled, contained small flattened vesicles, and these endings also invariably made synaptic contact, mostly onto spine-free dendritic membrane, and were characterized by symmetric membrane thickenings at the point of apposition. Our evidence supports the view that dopaminergic and other synaptic terminals in the rat caudate-putamen make synaptic contact with postsynaptic targets in the neostriatum and, at least in the adult, do not exist in appreciable numbers in the form of terminals that are not apposed to membranes of postsynaptic targets in the neostriatum.
Regulation by guanosine 3':5'-cyclic monophosphate of phospholipid methylation during chemotaxis in Dictyostelium discoideum.
Authors: Alemany S , García Gil M , Mato J M
Published: May 2006

In Dictyostelium discoideum, the chemoattractant cyclic AMP activates the enzyme guanylate cyclase, giving a brief up to 10-fold increase in the intracellular cyclic GMP content. The addition of physiological cyclic GMP concentrations to a homogenate of D. discoideum cells markedly increased the incorporation of the 3H-labeled methyl group from S-adenosyl-L-[methyl-3H]methionine into mono- and dimethylated phosphatidylethanolamine and phosphatidylcholine. Lipid methylation was inhibited by S-adenosyl-L-homocysteine, which inhibits transmethylation. When whole cells prelabeled with L-[methyl-3H]methionine were exposed to cyclic AMP, a rapid transient increase in the amount of [methyl-3H]phosphatidylcholine was observed. The time course of [methyl-3H]phosphatidylcholine formation agrees with its being mediated by the intracellular increase in cyclic GMP originating during chemotactic stimulation. Addition of the 8-Br derivative of cyclic GMP to whole cells also increased the levels of labeled phosphatidylcholine. It is therefore likely that cyclic GMP contributes to chemotaxis by regulating membrane function via phospholipid methylation.
DNA polymerase alpha mutants from a Drosophila melanogaster cell line.
Authors: Sugino A , Nakayama K
Published: May 2006

Aphidicolin, a tetracyclic diterpenoid antibiotic, is a specific inhibitor of DNA synthesis in vivo and DNA polymerase (deoxynucleosidetriphosphate:DNA deoxynucleotidyltransferase, EC 2.7.7.7) alpha of eukaryotic organisms. After ethyl methanesulfonate mutagenesis, we have recovered mutants of Drosophila melanogaster Schneider cell line no. 2 that grow at concentrations of aphidicolin that completely inhibit wild-type cells. The DNA polymerase alpha from one of these mutants, aph-10, is much more resistant to inhibition by the drug; the apparent Ki of the wild-type enzyme is 12 nM aphidicolin, whereas the apparent Ki of the aph-10 polymerase is more than 100 nM. (The apparent Km for dCTP is the same for both enzymes.) Another mutant, aph-13, overproduces DNA polymerase alpha at least 8-fold. The DNA polymerase of this mutant has the same apparent Km and Ki for dNTPs and aphidicolin as does wild-type polymerase.