Adenovirus type 12 infection of defined mouse-human hybrid cell clones

Summary Human adenovirus type 12 does not multiply in mouse cells; only viral T-antigen is detected. Mouse-human cell hybrid clones containing human chromosomes A3, B5, C7, C11, C12, D14, E17, F19 and F20, support synthesis of adenovirus DNA and capsid antigens.The hybrid clones used in these experiments were kindly supplied by Drs.Carlo Croce andHilary Koprowski of the Wistar Institute, Philadelphia, Pennsylvania. Supported by Grants from National Cancer Institute.     

Hemagglutination by simian adenovirus 7]

Simian adenovirus 7, either complete virus or its capsid subunits, agglutinates Rat (Sprague-Dowley) red blood cells in the presence of heterotypical antiserum. Haemagglutination takes place at 4 degrees C and room temperature. The antigen could not be eluted and its haemagglutinin properties are heat-stable. The reaction is specific. It is inhibited by homologous antiserum only. This property and its characteristics permit a camparison of this strongly oncogenic adenovirus to the human adenovirus of subgroup III of Rosen.     

Changes in immune reactivity during growth of an adenovirus 12-induced transplantable tumour in CBA mice

Summary Early suppression, followed by a period of enhancement and finally, suppression, was seen when the spleen cell response to T and B cell mitogens was monitored during growth of an adenovirus 12-induced tumour in CBA black mice. The macrophage content of the tumour changed with time and these fluctuations correlated with the ability of tumour tissue extracts to enhance the normal spleen cell response to mitogen.Acknowledgments. Thanks are dur to the Yorkshire Cancer Research Campaign, and the Cancer Research Committee of Sheffield University for financial support, also to Professor C.W. Potter for advice.     

Localization of integrated adenovirus DNA in the hamster genome

Adenovirus DNA integrated into the genomes of adenovirus-transformed hamster cells or of adenovirus type 12 (Ad12)-induced hamster tumor cells can be located at many different chromosomal sites. This raises the question as to whether distinct isochores of the hamster cell genome might be more accessible to recombination with adenovirus DNA. In Ad12- or Ad2-transformed hamster cell lines, and in Ad12 revertants, the investigated integrated viral DNA sequences were assigned to isochore families by analyzing DNA fractions from preparative CsCl density gradients for their buoyant densities (and, therefore, GC levels) and for the presence of viral DNA. Adenovirus DNA sequences were found in different isochores, which did not generally match the base composition of viral sequences. This is in apparent contrast to what was previously observed with retroviral integration. However, in cell lines carried in culture for many years, the viral DNA sequences might have been transposed to different isochore positions, since the host sequences flanking the viral DNA appear to have been conserved.Received 6 July 2004; received after revision 23 August 2004; accepted 6 October 2004     

Specific interactions of the adenovirus proteinase with the viral DNA,an 11-amino-acid viral peptide,and the cellular protein actin

The adenovirus proteinase (AVP) is synthesized in an inactive form that requires cofactors for activation. The interaction of AVP with two viral cofactors and with a cellular cofactor, actin, is characterized by quantitative analyses. The results are consistent with a specific model for the regulation of AVP. Late in adenovirus infection, inside nascent virions, AVP becomes partially activated by binding to the viral DNA, allowing it to cleave out an 11-amino-acid viral peptide, pVIc, that binds to AVP and fully activates it. Then, about 70 AVP-pVIc complexes move along the viral DNA, via one-dimensional diffusion, cleaving virion precursor proteins 3200 times to render a virus particle infectious. Late in adenovirus infection, in the cytoplasm, the cytoskeleton is destroyed. The amino acid sequence of the C terminus of actin is homologous to that of pVIc, and actin, like pVIc, can act as a cofactor for AVP in the cleavage of cytokeratin 18 and of actin itself. Thus, AVP may also play a role in cell lysis.Received 14 November 2002; received after revision 28 April 2003; accepted 30 April 2003     

Tyrosinase-cytalyzed conjugation of dopa with glutathione

Summary A convenient method is described for the preparation of 5-S- and 2-S-glutathionyldopa, based on tyrosinase oxidation of dopa in the presence of glutathione. The yields of 5-S, 2-S, and 6-S isomers produced were about 76, 12, and 5%, respectively.One of the authors (G.P.) thanks the C.N.R. (P.F. Chimica Fine e Secondaria) for partial financial support.     

Decrease in stimulation and allogeneic response following experimental infection of mice with human adenovirus 5]

15 days after experimental infection with human adenovirus 5, C57 Bl/6 murine spleen cells were found to have altered in vitro properties. Proliferative response to phytohemagglutinine was slightly increased while the allogeneic response was decreased and the capability to stimulate allogeneic cells was significantly diminished. The degree of expression of surface antigens responsible for allogeneic stimulation is influenced by many regulatory factors, still poorly defined; alteration of some factors of antigenicity may result, directly or indirectly, from certain viral infections.     

Evidence for the presence of virus in clonal insulin-producing RINm5F cells

Summary The ultrastructural morphology of the clonal insulin-producing cell line, RINm5F, was investigated. Virus-like particles, probably C type viruses, were identified both intra- and extracellularly. Because these particles could not be found in the original transplantable tumor, it is probable that viruses were induced at some later stage in the development of the RINm5F cell line. All investigators using the RINm5F cells should be aware of the fact that these cells may contain one or several types of viruses, and of the possibility that these particles may interfere with a variety of cellular functions.This work was supported by grants from the Lake County Medical Center Developmental Agency (to VH and PWB) and the American Heart Association, Indiana Affiliate (to PWB,^*USA) and by the Swedish Medical Research Council (12X-562), the Swedish Diabetes Association, the Nordic Insulin Foundation and Åke Wibergs Foundation (^**Sweden). Per-Olof Berggren is a recipient of a postdoctoral fellowship from the Swedish Medical Research Council.     

Rauwolfia alkaloids,XVI. Deserpidine,a new alkaloid fromRauwolfia canescens

Zusammenfassung AusRauwolfia canescens wurde Deserpidin, C₃₂H₃₈O₈N₂, isoliert und eine Konstitutionsformel für dieses neue Alkaloid vorgeschlagen.

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Communication XV,C. F. Huebner et al., J.A.C.S. (in press).

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From lecture given at Columbia University, New York, January 12th, 1955.     

Studies on the Dd antigen-antibody system. I. The nature of antigen Dd and its antibodies

Summary The nature of antigen Dd, an antigen present in the extracts of human dandruff which precipitates human sera selectively, and antibodies reacting with it are reported.This work was supported through U.G.C. Grant No. F.23-230/75/SR II. H.K. participated in this study, first as J.R.F. of U.G.C. and then as S.R.F. of I.C.M.R. We are grateful to Dr Baruch S. Blumberg for his invaluable suggestions and to Professor H. Walter for the gift of IgG, IgM and IgA immune sera.     

Macrophage origin of platelet activating factor]

Platelet-activating factor (P.A.F.) is a mediator of anaphylaxis released from human and Rabbit basophils which causes aggregation of platelets and release of their vasoactive amines. We have induced the release of P.A.F. from Rat peritoneal cells (P.C.) with ionophore A 23187. After fractionation of P.C. on 5-15% Ficoll gradients, P.A.F. was obtained from macrophage-rich but not from mastocyte-rich fractions and from adherent cells but not from non adherent cells. These data suggest an important new function for the macrophage: aggregation of platelets and release of their vasoactive amines and others mediators of inflammation.     

Genetic analysis of interspecific crosses Mus musculus L. x Mus spretus Lataste: linkage of Adh-1 with Amy-1 on chromosome 3 and Es-14 with Mod-1 on chromosome 9]

192 offsprings from interspecific back-crosses (male M. spretus x female BALB/c) F1 x male BALB/c or (male M. spretus x female C57BL6) F1 x male C57BL6 were analysed at thirteen structural loci. Linkage of ES-14 with Mod-1 on chromosome 9 and that of Adh-1 with Amy-1 on chromosome 3 are shown. The following order centromere/Car-2/Amy-1 is tentatively proposed for these loci on chromosome 3.     

Hybridization between Robertsonian karyotypic races of the common shrewSorex araneus

Summary British common shrews of the Aberdeen, Oxford and Hermitage Robertsonian karyotypic races were hybridized successfully in captivity. Hybrids, both simple Robertsonian heterozygotes and double Robertsonian heterozygotes with monobrachial homology, have been identified in an area of contact between the Oxford and Hermitage races. The relative fertility of these two types of hybrid is considered.I thank Dr. C. E. Ford F. R. S., Prof. F. W. Robertson and Dr A. E. Douglas for reading earlier drafts of this paper, Prof. F. W. Robertson, Dr J. R. K. Savage and Dr J. R. Clarke for laboratory facilities, Mrs J. E. Evans for technical assistance and S.E.R.C. for financial support.     

Purification of RNA from animal cells using diethyl-pyrocarbonate

Summary Extraction of RNA from animal cells by a method using diethyl-pyrocarbonate yielded 50–60% of the total RNA. RNA purified by a hot phenol-SDS method from adenovirus 2 infected cells showed about 9% homology with adenovirus DNA, and RNA purified by diethyl-pyrocarbonate-SDS showed over 7% hybridization. Profiles of RNA prepared by both methods were identical when studied by polyacrylamide gel electrophoresis.Acknowledgment. This investigation was supported by U.S. Public Health Services research grant No. 5R01-CA10724-03 from the National Cancer Institute.     

Evidence for the presence of virus in clonal insulin-producing RINm5F cells

The ultrastructural morphology of the clonal insulin-producing cell line, RINm5F, was investigated. Virus-like particles, probably C type viruses, were identified both intra- and extracellularly. Because these particles could not be found in the original transplantable tumor, it is probable that viruses were induced at some later stage in the development of the RINm5F cell line. All investigators using the RINm5F cells should be aware of the fact that these cells may contain one or several types of viruses, and of the possibility that these particles may interfere with a variety of cellular functions.     

Rate of tryptophan hydroxylation in vivo in brain nuclei of genetically hypertensive rats of the Lyon strain

Summary The rate of tryptophan hydroxylation in vivo is unaltered in brain areas of 5, 9 and 21 week-old Lyon genetically Hypertensive (LH) rats as compared to both Lyon Normotensive (LN) and Low Blood Pressure (LL) rats, except for a decrease in the C1 area of the medulla oblongata in 9 week-old animals.Acknowledgment. The authors wish to thank Dr M.F. Belin, Dr J.F. Pujol and Mrs J. Sacquet for their help during this study. This work was supported by the Fondation pour la Recherche Médicale Française and the C.N.R.S.     

Genotypic differences between the mouse adenovirus strains FL and K87

Summary Restriction endonuclease analysis was used to compare the genome of mouse adenovirus (MAd) strains FL and K87. Large differences were found between theKpn I,PaeR7,Pvu I,Sal I andSma I restriction profiles of the prototype strains. MAd-FI and MAd-K87 thus represent two distinct species of mouse adenovirus.This work was supported by grants from the Fonds pour la formation de chercheurs et l'aide à la recherche and the Natural Sciences and Engineering Research Council of Canada.     

Proteomic interaction profiling reveals KIFC1 as a factor involved in early targeting of F508del-CFTR to degradation

Misfolded F508del-CFTR, the main molecular cause of the recessive disorder cystic fibrosis, is recognized by the endoplasmic reticulum (ER) quality control (ERQC) resulting in its retention and early degradation. The ERQC mechanisms rely mainly on molecular chaperones and on sorting motifs, whose presence and exposure determine CFTR retention or exit through the secretory pathway. Arginine-framed tripeptides (AFTs) are ER retention motifs shown to modulate CFTR retention. However, the interactions and regulatory pathways involved in this process are still largely unknown. Here, we used proteomic interaction profiling and global bioinformatic analysis to identify factors that interact differentially with F508del-CFTR and F508del-CFTR without AFTs (F508del-4RK-CFTR) as putative regulators of this specific ERQC checkpoint. Using LC–MS/MS, we identified kinesin family member C1 (KIFC1) as a stronger interactor with F508del-CFTR versus F508del-4RK-CFTR. We further validated this interaction showing that decreasing KIFC1 levels or activity stabilizes the immature form of F508del-CFTR by reducing its degradation. We conclude that the current approach is able to identify novel putative therapeutic targets that can be ultimately used to the benefit of CF patients.     

Epidermal sensitivity to hypoxia in the lugworm

Summary Physiological and anatomical data support the idea that in the caudal epidermis of the lugworm,Arenicola marina (L.), there are chemoreceptors detecting variations of oxygen partial pressure in the ambient water.This research was partly supported by a C.N.R.S. grant, AI No. 582 085. We thank Dr P. Dejours and Dr J. Taxi for helpful comments and discussion, Mrs S. Dejours for editing the English of this paper, M.F. Baucher, A. Dorme, F. Kleinbauer, the staff of the Station Biologique de Roscoff and the Service de Microscopie Electronique, C.N.R.S., boulevard Raspail, Paris, for technical assistance.     

RACK1 depletion in a mouse model causes lethality, pigmentation deficits and reduction in protein synthesis efficiency

The receptor for activated C-kinase 1 (RACK1) is a conserved structural protein of 40S ribosomes. Strikingly, deletion of RACK1 in yeast homolog Asc1 is not lethal. Mammalian RACK1 also interacts with many nonribosomal proteins, hinting at several extraribosomal functions. A knockout mouse for RACK1 has not previously been described. We produced the first RACK1 mutant mouse, in which both alleles of RACK1 gene are defective in RACK1 expression (ΔF/ΔF), in a pure C57 Black/6 background. In a sample of 287 pups, we observed no ΔF/ΔF mice (72 expected). Dissection and genotyping of embryos at various stages showed that lethality occurs at gastrulation. Heterozygotes (ΔF/+) have skin pigmentation defects with a white belly spot and hypopigmented tail and paws. ΔF/+ have a transient growth deficit (shown by measuring pup size at P11). The pigmentation deficit is partly reverted by p53 deletion, whereas the lethality is not. ΔF/+ livers have mild accumulation of inactive 80S ribosomal subunits by polysomal profile analysis. In ΔF/+ fibroblasts, protein synthesis response to extracellular and pharmacological stimuli is reduced. These results highlight the role of RACK1 as a ribosomal protein converging signaling to the translational apparatus.