An electron microscopic study of the Golgi apparatus of amphibian oocytes

Conclusion An electron microscope study of the Golgi complex was carried out using amphibian oocytes both prior to and after vitellogenesis. In the former case it was noted that the Golgi complex consists of small vesicles and cisternae. After yolk formation, each Golgi mass was found to consist only of (a smaller number of) cisternae. The distribution, function, and multiplication of the Golgi material were also discussed.This work was supported by U.S.P.H.S. research grant RG 5803 (C 2) and was carried out during the course of a U.S.P.H.S. Special Research Fellowship (GF—5356—C 2).     

Evidence for nonrandom multiplicity of gene products in 22 plant genera

Correlations in the number (multiplicity) of molecular forms among 10 enzymes have been estimated from a sample of plants representing 22 genera. Significant correlations in multiplicity among some of the 10 enzyme systems suggest a nonrandom organization of variation in number of molecular forms. An analysis of the correlations among enzymes of a glycolytic and Krebs cycle subset supports the occurrence of patterns of influence on multiplicity which correspond to the known metabolic relatedness of the enzymes. We interpret these data as evidence for organization of enzyme multiplicity. This property may be of adaptive value to the species.This research was supported by U.S.A.E.C. Contract AT(11-1)-1552, U.S.P.H.S. Grant AM 09381 and Rackham Faculty Award NSF-GU-3470-Project 131, and U.S.P.H.S. Career Development Award 1-K3-AM7959 (GJB).     

A sex-limited serum protein variant in the mouse: Inheritance and association with the H-2 region

An alloantiserum produced in the mouse has been used to detect an antigen which is present only in male serum from certain inbred strains of mice, e.g., DBA/2J, A/J, and BALB/c. Genetic tests reveal that the presence of this antigen is controlled by a dominant autosomal gene which is expressed only in males of the proper genotype. Test crosses and analysis of congenic resistant strains indicate close linkage between the sex-limited protein (Slp) and the histocompatibility-2 (H-2) region of linkage group IX. Analysis of seven intra-H-2 recombinant strains is consistent with the placement of the genetic determinant for Slp within the H-2 region in the same position as the Ss (serum substance) determinant. Immunological evidence suggests that the Slp antigenic sites reflect structural variation in the Ss component of mouse serum.Supported by U.S.P.H.S. Research Grant GM-15419, U.S.P.H.S. Career Development Award K3-HE-24, 980 (D.C.S.), and U.S.P.H.S. Training Grant 2T01-GM-00071 (H.C.P.).     

A sex-limited serum protein variant in the mouse: Hormonal control of phenotypic expression

The sex-limited protein (Slp) antigen of the mouse is first detected in the serum of strain DBA/2J males at 5–6 weeks of age and reaches full adult levels by 10 weeks. This antigen is normally absent in females. Immature DBA/2J males castrated at 3 1/2 weeks of age failed to develop Slp antigen, while DBA/2J females treated with testosterone propionate starting at 3 1/2 weeks developed normal adult male levels of Slp antigen. Similar hormone-influenced effects were demonstrated in adult males and females of the same strain. Experiments indicated that testosterone does not act directly in the serum to expose Slp antigenic sites. Testosterone treatment of both males and females of strain C57BL/10JSf, which does not carry the gene for the presence of the Slp antigen, failed to stimulate the appearance of the antigen. Thus, the presence of Slp antigen in the serum is dependent on both the proper genotype and the presence of male hormone.Supported by U.S.P.H.S. Research Grant GM-15419, U.S.P.H.S. Training Grant 2T01-GM-00071 (H.C.P.), and U.S.P.H.S. Career Development Award K3-HE-24,980 (D.C.S.).     

A comparative scanning electron microscopic analysis of the human cerebral ventricular system

Summary A scanning electron microscopic analysis of the adult human third ventricular wall revealed ultra-architectural differences between dorsal and ventral portions. In the brains of thirteen and sixteen week old human fetuses regional differences in the surface organization of lining ependyma were more sharply defined than those of the adult. Alterations in the luminal surfaces of ependyma may reflect differences in the functional capacity of various ventricular areas. The potential role of certain ependyma (tanycytes) and their putative participation in neuroendocrine events is discussed.Supported by U.S.P.H.S. Grant NS 08171.U.S.P.H.S. Career Development Awardee K 04 GM 70001.     

中国绒泡菌属的分类学研究II.产自新疆的新种和稀有种

关于新疆的粘菌,过去仅知８种,且无一为绒泡菌。本文第一作者于１９９４年在新疆进行了粘菌资源调查,在随后的研究中共明确了５０多个种,有１０种为绒泡菌,其中的几个种,如垂头绒泡菌Ｐｈｙｓａｒｕｍｎｕｔａｎｓ（Ｂｕｌｌ．）Ｐｅｒｓ．是常见和广布的,但也有几种是特殊的和稀有的。本文报告了５种绒泡菌：橙红绒泡菌Ｐ.ａｕｒａｎｔｉａｃｕｍ　Ｓ．Ｌ．Ｃｈｅｎ,Ｙ．ＬｉｅｔＨ．Ｚ．Ｌｉ和侧扁绒泡菌Ｐ．ｌｏｒａｔｕｍＳ．Ｌ．ＣｈｅｎＹ．Ｌｉ　ｅｔＨ．Ｚ．Ｌｉ是新种,黄白绒泡菌Ｐ．ａｌｂｅｓｃｅｎｓＥｌｌｉｓｅｘＴ．Ｍａｃｂｒ．和团聚绒泡菌Ｐ．ｃｏｎｇｌｏｍｅｒａｔｕｍ（Ｆｒ．）Ｒｏｓｔａｆ．是中国新记录种,已知种黄绿绒泡菌Ｐ．ｖｉｒｅｓｃｅｎｓＤｉｔｍａｒ显然少见,国内此前仅知分布于福建。橙红绒泡菌Ｐ．ａｕｒａｎｔｉａｃｕｍＳ．Ｌ．Ｃｈｅｎ,Ｙ．ＬｉｅｔH.Ｚ．Ｌｉ的孢丝近似钙丝菌状,由或大或小的石灰结将许多细短的孢丝线联成致密的白色网体,这使其被归入绒泡菌属,并与同属其它种相区分;侧扁绒泡菌Ｐ．ｌｏｒａｔｕｍＳ．Ｌ．Ｃｈｅｎ,Ｙ．ＬｉｅｔＨ．Ｚ．Ｌｉ的联囊体虽发达而侧扁,但顶部无预成开裂线,在相似种中也易识别。本文对这两个新种     

中国绒泡菌属的分类学研究 Ⅱ.产自新疆的新种和稀有种

关于新疆的粘菌,过去仅知８种,且无一为绒泡菌。本文第一作者于１９９４年在新疆进行了粘菌资源调查,在随后的研究中共明确了５０多个种,有１０种为绒泡菌,其中的几个种,如垂头绒泡菌Ｐｈｙｓａｒｕｍｎｕｔａｎｓ（Ｂｕｌｌ．）Ｐｅｒｓ．是常见和广布的,但也有几种是特殊的和稀有的。本文报告了５种绒泡菌：橙红绒泡菌Ｐ.ａｕｒａｎｔｉａｃｕｍ　Ｓ．Ｌ．Ｃｈｅｎ,Ｙ．ＬｉｅｔＨ．Ｚ．Ｌｉ和侧扁绒泡菌Ｐ．ｌｏｒａｔｕｍＳ．Ｌ．ＣｈｅｎＹ．Ｌｉ　ｅｔＨ．Ｚ．Ｌｉ是新种,黄白绒泡菌Ｐ．ａｌｂｅｓｃｅｎｓＥｌｌｉｓｅｘＴ．Ｍａｃｂｒ．和团聚绒泡菌Ｐ．ｃｏｎｇｌｏｍｅｒａｔｕｍ（Ｆｒ．）Ｒｏｓｔａｆ．是中国新记录种,已知种黄绿绒泡菌Ｐ．ｖｉｒｅｓｃｅｎｓＤｉｔｍａｒ显然少见,国内此前仅知分布于福建。橙红绒泡菌Ｐ．ａｕｒａｎｔｉａｃｕｍＳ．Ｌ．Ｃｈｅｎ,Ｙ．ＬｉｅｔH.Ｚ．Ｌｉ的孢丝近似钙丝菌状,由或大或小的石灰结将许多细短的孢丝线联成致密的白色网体,这使其被归入绒泡菌属,并与同属其它种相区分;侧扁绒泡菌Ｐ．ｌｏｒａｔｕｍＳ．Ｌ．Ｃｈｅｎ,Ｙ．ＬｉｅｔＨ．Ｚ．Ｌｉ的联囊体虽发达而侧扁,但顶部无预成开裂线,在相似种中也易识别。本文对这两个新种     

The Thai variant and the distribution of alleles of 6-phosphogluconate dehydrogenase and the distribution of glucose 6-phosphate dehydrogenase deficiency in Thailand

The samples were taken from 3185 subjects from ten provinces throughout Thailand. In 1577 males the frequency of glucose 6-phosphate dehydrogenase deficiency was 11.98%. In the far south the gene frequency was 2.83%; in the remainder of the country the frequency did not vary significantly about a mean of 13.76%. The deficiency is of a severe type. The G6PD of all of the nondeficient individuals had the electrophoretic mobility of type B. The mean frequency of the A/B electrophoretic phenotype of 6-phosphogluconate dehydrogenase is 8.47%. The maximum frequency was in central and southern Thailand with a decline to the north and northeast. A variant form of 6-PGD, referred to as the Thai variant, has been found in which two additional electrophoretic components migrate anodally to the normal A band, confirming that the molecule is at least a dimer. The hypothesis is advanced that erythrocyte 6-PGD is determined by two genetic loci, only one of which is translated in leukocytes.Supported by U.S. Army Contract DA-49-193 MD 2879, U.S.P.H.S. GM 09252, and U.S.P.H.S. 5-K3-GM-15325.     

Localization of neurophysin in the rat supraoptic nucleus

Summary Localization of neurophysin in neurons of the supraoptic nucleus was accomplished using an unlabeled-antibody, post-embedding, immunoperoxidase technique. Neurophysin was exclusively associated with neurosecretory granules within cell bodies of supraoptic neurons and their processes.Supported by U.S.P.H.S. Grant HD-08867     

Structural characterization of a six-nucleotide RNA hairpin loop found in Escherichia coli, r(UUAAGU)

The binding region of the Escherichia coli S2 ribosomal protein contains a conserved UUAAGU hairpin loop. The structure of the hairpin formed by the oligomer r(GCGU4U5A6A7G8U9CGCA), which has an r(UUAAGU) hairpin loop, was determined by NMR and molecular modeling techniques as part of a study aimed at characterizing the structure and thermodynamics of RNA hairpin loops. Thermodynamic data obtained from melting curves for this RNA oligomer show that it forms a hairpin in solution with the following parameters: DeltaH degrees = -42.8 +/- 2.2 kcal/mol, DeltaS degrees = -127.6 +/- 6.5 eu, and DeltaG degrees (37) = -3.3 +/- 0.2 kcal/mol. Two-dimensional NOESY WATERGATE spectra show an NOE between U imino protons, which suggests that U4 and U9 form a hydrogen bonded U.U pair. The U5(H2') proton shows NOEs to both the A6(H8) proton and the A7(H8) proton, which is consistent with formation of a "U" turn between nucleotides U5 and A6. An NOE between the A7(H2) proton and the U9(H4') proton shows the proximity of the A7 base to the U9 sugar, which is consistent with the structure determined for the six-nucleotide loop. In addition to having a hydrogen-bonded U.U pair as the first mismatch and a U turn, the r(UUAAGU) loop has the G8 base protruding into the solvent. The solution structure of the r(UUAAGU) loop is essentially identical to the structure of an identical loop found in the crystal structure of the 30S ribosomal subunit where the guanine in the loop is involved in tertiary interactions with RNA bases from adjacent regions [Wimberly, B. T., Brodersen, D. E., Clemons, W. M., Morgan-Warren, R. J., Carter, A. P., Vonrhein, C., Hartsch, T., and Ramakrishnan, V. (2000) Nature 407, 327-339]. The similarity of the solution and solid-state structures of this hairpin loop suggests that formation of this hairpin may facilitate folding of 16S RNA.     

In vitro identification of a soluble protein:geranylgeranyl transferase from rat tissues

The gamma subunit of mammalian trimeric G proteins has been shown previously to be modified in vivo on a cysteine residue situated at the carboxyl-terminal sequence-Cys-Ala-Ile-Leu-COOH by a 20-carbon prenyl moiety geranylgeranyl (Mumby, S. M., Casey, P. J., Gilman, A. G., Gutowski, S., and Sternweis, P. C. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 5873-5877; Yamane, H. K., Farnsworth, C. C., Xie, H., Howald, W., Fung, B. K-K., Clarke, S., Gelb, M. H., and Glomset, J. A. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 5866-5872). A biotinylated peptide acceptor comprising the eight carboxyl-terminal amino acids of the gamma subunit and tritiated geranylgeranyl diphosphate were utilized to monitor a protein:prenyl transferase activity in rat organs of varying age. The transferase activity was dependent upon the presence of divalent metal ions and maximal activity was achieved with either 1 mM ZnCl2 or 20 mM MgCl2. Activity was shown to be linear with respect to time, protein concentration, substrate concentration, and the pH optimum was 7.5. Protein:geranylgeranyl transferase activity was detected in all rat organs studied with the highest specific activity in brain S100. No activity was detected in the membrane fraction. The specific activity in brain, liver, kidney, and heart increased with age. Radioactivity incorporated into the peptide acceptor from both [1-3H]geranylgeranyl diphosphate and [5-3H]mevalonate by 21-day-old rat brain S100 was released by treatment with methyl iodide, and in both cases, analysis of the cleavage products by reversed phase high performance liquid chromatography showed a peak of radioactivity co-eluting with a geranylgeraniol standard which was well resolved from a farnesol standard. This indicated that the rat brain S100 contained not only the protein:geranylgeranyl transferase but also geranylgeranyl synthetase activity and that the peptide acceptor was specific for geranylgeranyl under the conditions tested.     

Salicylate 5-hydroxylase from Ralstonia sp. strain U2: a monooxygenase with close relationships to and shared electron transport proteins with naphthalene dioxygenase

The genes from the oxygenase cluster nagAaGHAbAcAd of naphthalene-degrading Ralstonia sp. strain U2 were cloned and overexpressed. Salicylate 5-hydroxylase (S5H) activity, converting salicylate to gentisate, was present in vitro only in the single extract of cells with overexpressed nagAaGHAb or in a mixture of three cell extracts containing, respectively, NagGH (the oxygenase components), NagAa (ferredoxin reductase), and NagAb (ferredoxin). Each of the three extracts required for S5H activity was rate limiting in the presence of excess of the others but, when in excess, did not affect the rate of catalysis. S5H catalyzed the 5-hydroxylation of the aromatic rings of 3- and 4-substituted salicylates. However, the methyl group of 5-methylsalicylate was hydroxylated to produce the 5-hydroxymethyl derivative and the 6-position on the ring of 5-chlorosalicylate was hydroxylated, producing 5-chloro-2,6-dihydroxybenzoate. In an assay for the nag naphthalene dioxygenase (NDO) based on the indole-linked oxidation of NADH, three extracts were essential for activity (NagAcAd, NagAa, and NagAb). NDO and S5H were assayed in the presence of all possible combinations of the nag proteins and the corresponding nah NDO proteins from the "classical" naphthalene degrader P. putida NCIMB9816. All three oxygenase components functioned with mixed combinations of the electron transport proteins from either strain. The S5H from strain U2 is a unique monooxygenase which shares sequence similarity with dioxygenases such as NDO but is also sufficiently similar in structure to interact with the same electron transport chain and probably does so in vivo during naphthalene catabolism in strain U2.     

Quantitative measurement of sphingosine 1-phosphate by radioreceptor-binding assay

In Chinese hamster ovary cells overexpressing Edg-1, one of the sphingosine 1-phosphate (S1P) receptor subtypes, [(3)H]S1P binding was displaced by unlabeled S1P with IC(50), a half-maximal concentration to inhibit the binding, of about 20 nM. This radioreceptor binding was used for quantitative measurement of S1P. Among the various lipids employed, only sphingosylphosphorylcholine (SPC), other than S1P, practically displaced the binding; however, the potency of SPC was about 100 to 1000 times less than that of S1P. Thus, SPC bound to the S1P receptors inefficiently. Furthermore, before the application of test samples to this assay, S1P was partially purified: the lipid was extracted first into the aqueous phase and separated from other lipids under alkaline conditions, and then reextracted into the chloroform phase under acidic conditions. With this assay, we could specifically and quantitatively measure S1P from 2 to 40 pmol per assay well in biological samples including serum samples and various tissues. This assay also allowed us to measure the change in cellular S1P content in U937 cells after treatment with exogenous sphingosine.     

Fibrillar structures resembling leiomyofibrils in endothelial cells of mammalian pulmonary bloodvessels

Summary Endothelial cells of mammalian pulmonary arteries, arterioles and precapillaries (metarterioles) posses bundled filamentous structures which resemble leiomyofibrils.Supported by U.S.P.H.S. Grants A 5514-03, GM-K3-14834, 18573-R2 and CA-11, 77-01A1.     

Generation of a new adenovirus type 12-inducible fragile site by insertion of an artificial U2 locus in the human genome.

Infection with adenovirus type 12 (Ad12) induces four fragile sites in the human genome (H.F. Stich, G.L. van Hoosier, and J.J. Trentin, Exp. Cell Res. 34:400-403, 1964; H. zur Hausen, J. Virol. 1:1174-1185, 1967). The major site, at 17q21-22, contains the U2 gene cluster, which is specifically disrupted by infection in at least a percentage of the cells (D.M. Durnam, J.C. Menninger, S.H. Chandler, P.P. Smith, and J.K. McDougall, Mol. Cell. Biol. 8:1863-1867, 1988). For direct assessment of whether the U2 locus is the target of the Ad12 effect, an artificial locus, constructed in vitro and consisting of tandem arrays of the U2 6-kbp monomer, was transfected into human cells. We report that integration of this artificial locus on the p arm of chromosome 13 creates a new Ad12-inducible fragile site.     

Fine structure of trypsin-dissociated rat adrenal cells

Summary Electron micrographs of trypsin-dissociated rat adrenal showed predominantly intact rounded cells without internal damage. The population contained cells from the glomerular, intermediary and fascicular zones with cells from the zona fasciculata predominant. The presence or absence of cells from the reticular zone could not be determined. Cells from the medullary zone were absent. The addition of adrenocorticotropic hormone (ACTH) to the cellular suspension for 2 hours produced corticosterone. However, these stimulated cells did not display any significant ultrastructural change.Supported by research grants from the U.S.P.H.S. (No. GM15872), and the Research Council of Rutgers University, and the National Science Foundation (No. GB7427 to Dr. George Sayers). We are indebted to Mrs. Jean A. Gibney, Miss Rose-Marie Ma, and Mrs. Mary Vegh for their excellent technical assistance.Postdoctoral Trainee, U.S.P.H.S. Training Grant No. 5T01-GM00899-11.     

Uptake of radioactive thymidine and cytidine by Ehrlich ascites tumor cells in different stages of growth

Summary In Strong A female mice, the Ehrlich ascites tumor inoculated into the peritoneal cavity grows exponentially for the first 7 days with a doubling time of about 36 hours. The tumor enters then into a late stage during which the number of tumor cells in the peritoneal cavity does not increase. The uptake of intraperitoneally injected thymidine decreases from the exponential to the late stage, mostly because of a decrease in the fraction of cells in DNA synthesis. During the exponential phase, the uptake of thymidine is a function of the amount of radioactive thymidine injected per tumor cell, the utilization decreasing with increasing cell dose. The uptake of intraperitoneally injected cytidine decreases slightly with time after inoculation although the fraction of tumor cells in RNA synthesis remains constant.Dedicated to Professor Friedrich Wassermann with admiration and affection on the occasion of his 80th birthday.This investigation was supported by U.S.P.H.S. Grant CA-05667. The author is a U.S.P.H.S. Research Career Development Awardee.     

Epigenetic Marks Associated to the Study of Nucleolar Dominance in Urochloa P. Beauv.

The genus Urochloa P. Beauv. presents a prominent role in the tropical agricultural scenario being composed of species with different ploidy levels. Studies on the genomic relationship within this genus as well as specific analysis involving epigenetic marks are limited. The aim of the present study was to identify the cytosine methylation (5-mCyt) and histone H3 lysine 9 dimethylation (H3k9me2) in the different modulations of 45S ribosomal DNA (rDNA) sites in interphase nuclei and to associate these results with gene expression analysis in Urochloa ruziziensis (2n = 4x = 36), Urochloa brizantha cv. Marandu (2n = 4x = 36), and their respective interspecific hybrid H1863 (2n = 4x = 36). Immunolocalization techniques were performed in combination with Fluorescence in situ hybridization (FISH) for the location of the 45S rDNA sites. Predominantly, we observed intra- and perinucleolar sites, mostly hypomethylated and/or hyper/hypomethylated, decondensed or partially condensed. The gene expression analysis was performed qualitatively through the conventional PCR using complementary DNA and confirmed by the RT-qPCR technique and primers designed for the ITS-1 region of U. brizantha and U. ruziziensis. The molecular analyses performed on leaves showed that there is dominance of U. brizantha 45S rDNA gene expression on U. ruziziensis in the H1863 hybrid. In roots, the analyses showed that the 45S rDNA genes of the two parents are expressed in the hybrid genome. Thus, it is plausible to infer a tissue-specific nuclear dominance model in which the pattern of hypermethylated cytosine sites with heterochromatic marks and, therefore, silenced were mostly inherited from U. ruziziensis, whereas the rDNA originated from U. brizantha was characterized by cytosine and H3k9 hypomethylation.