“Unblocking” Serum Activity <Emphasis Type="Italic">in vitro</Emphasis> in the Polyoma System may Correlate with Antitumour Effects of Antiserum <Emphasis Type="Italic">in vivo</Emphasis>

In a variety of tumour systems, individuals carrying progressively growing neoplasms have lymphoid cells with a specific cytotoxic effect on cultured tumour cells from the same individual^1–4. Since the sera of tumour-bearing individuals have been shown to prevent tumour cell destruction by immune lymphocytes in vitro^2,5–8 and since this serum blocking activity appears early in primary and transplant tumour development^5,7, it has been suggested that the appearance of this serum blocking activity might be responsible for the progressive growth of tumours in individuals having cytotoxic lymphocytes. Counteraction of this blocking activity would thus be of primary importance in facilitating the function of an already existing or bolstered cell-mediated immunity. The serum blocking activity might be inhibited in various ways, by preventing the formation of blocking antibody or by interfering with its action (“unblocking”), as demonstrated in Moloney sarcoma regressor sera⁹. This type of serum also has a therapeutic effect on Moloney sarcomas in vivo^10,11, which has been tentatively attributed to its unblocking activity^8,9 or, possibly, to a complement-dependent cytotoxicity¹⁰. Tumour growth in the Moloney sarcoma system, however, might be due in part to continuous recruitment of neoplastic cells by virus-induced transformation and so the therapeutic effect could be due to a virus-neutralizing serum activity^9,10.     

Examination of the 1,4-disubstituted azetidinone ring system as a template for combretastatin A-4 conformationally restricted analogue design

A series of novel 1,4-diaryl-2-azetidinones was prepared by stereospecific Staudinger reaction as conformationally restricted analogues of combretastatin A-4 because molecular modeling studies suggested close geometric similarities. They were evaluated for cytotoxicity against a number of human tumor and normal cell lines. Strong potencies were observed, with the best compounds exhibiting IC(50)'s of 25-74 nM against human neuroblastoma IMR 32 cell growth and a variety of other cell lines. Compounds inhibited tubulin polymerization with potencies commensurate with their cytotoxic activity and a more soluble anilino-containing analogue was very effective in inhibiting the growth of AR42J rat pancreatic tumors transplanted into in nude mice. Further studies on this interesting group of compounds as anti-cancer agents appear warranted.     

On the mechanism of ATP-induced shape changes in the human erythrocyte membranes: the role of ATP

In the preceding paper (Sheetz, M. and S.J. Singer. 1977. J Cell Biol. 73:638-646) it was shown that erythrocyte ghosts undergo pronounced shape changes in the presence of mg-ATP. The biochemical effects of the action of ATP are herein examined. The biochemical effects of the action of ATP are herein examined. Phosphorylation by ATP of spectrin component 2 of the erythrocyte membrane is known to occur. We have shown that it is only membrane protein that is significantly phosphorylated under the conditions where the shape changes are produced. The extent of this phosphorylation rises with increasing ATP concentration, reaching nearly 1 mol phosphoryle group per mole of component 2 at 8mM ATP. Most of this phosphorylation appears to occur at a single site on the protein molecule, according to cyanogen bromide peptide cleavage experiments. The degree of phosphorylation of component 2 is apparently also regulated by a membrane-bound protein phosphatase. This activity can be demonstrated in erythrocyte ghosts prepared from intact cells prelabeled with [(32)P]phosphate. In addition to the phosphorylation of component 2, some phosphorylation of lipids, mainly of phosphatidylinositol, is also known to occur. The ghost shape changes are, however, shown to be correlated with the degree of phosphorylation of component 2. In such experiment, the incorporation of exogenous phosphatases into ghosts reversed the shape changes produced by ATP, or by the membrane-intercalating drug chlorpromazine. The results obtained in this and the preceding paper are consistent with the proposal that the erythrocyte membrane possesses kinase and phosphates activities which produce phosphorylation and dephosphorylation of a specific site on spectrin component 2 molecules; the steady-state level of this phosphorylation regulates the structural state of the spectrin complex on the cytoplasmic surface of the membrane, which in turn exerts an important control on the shape of the cell.     

Using the longest significance run to estimate region-specific p-values in genetic association mapping studies

Background  

Association testing is a powerful tool for identifying disease susceptibility genes underlying complex diseases. Technological advances have yielded a dramatic increase in the density of available genetic markers, necessitating an increase in the number of association tests required for the analysis of disease susceptibility genes. As such, multiple-tests corrections have become a critical issue. However the conventional statistical corrections on locus-specific multiple tests usually result in lower power as the number of markers increases. Alternatively, we propose here the application of the longest significant run (LSR) method to estimate a region-specific p-value to provide an index for the most likely candidate region.     

Cytokinin synthesis is higher in the Arabidopsis amp1 mutant

Cytokinins are involved in plant cell proliferation leading to plant growth and morphogenesis. Earlier we described a mutant of Arabidopsis thaliana, amp1, that had five times higher levels of cytokinin and had a number of pleiotropic phenotypes, including increased cell proliferation and de-etiolated growth in the dark. While these phenotypes were correlated with higher levels of cytokinin, the actual mechanism of how cytokinin is elevated was not elucidated before. In order to understand if the increased cytokinin is a result of increased biosynthesis or decreased degradation we have compared the synthesis of cytokinins from radiolabelled adenine and the degradation of zeatin ribosides and other cytokinins between amp1 and wild type plants. The degradation of the hormone is not affected in the mutant but there is a 4 to 6 fold increase in cytokinin synthesis compared to the wild type. Because the amp1 mutant is recessive we hypothesise that the AMP1 product negatively regulates cytokinin production.     

MPDA: Microarray pooled DNA analyzer

Background  

Microarray-based pooled DNA experiments that combine the merits of DNA pooling and gene chip technology constitute a pivotal advance in biotechnology. This new technique uses pooled DNA, thereby reducing costs associated with the typing of DNA from numerous individuals. Moreover, use of an oligonucleotide gene chip reduces costs related to processing various DNA segments (e.g., primers, reagents). Thus, the technique provides an overall cost-effective solution for large-scale genomic/genetic research. However, few publicly shared tools are available to systematically analyze the rapidly accumulating volume of whole-genome pooled DNA data.     

A web server to locate periodicities in a sequence

Summary : FT is a tool written in C++, which implements the Fourier analysis method to locate periodicities in aminoacid or DNA sequences. It is provided for free public use on a WWW server with a Java interface. Availability : The server address is http://o2.db. uoa.gr/FT Contact : shamodr@atlas.uoa.gr      

Genes encoding ADP-ribosylation factors in Arabidopsis thaliana L. Heyn.; genome analysis and antisense suppression

Vesicle trafficking delivers proteins to intracellular and extracellular compartments, cellulose synthase to the plasma membrane, and non-cellulosic polysaccharides to the cell wall. The Arabidopsis genome potentially encodes 19 proteins with sequence similarities to ARFs (ADP-ribosylation factors) and its relatives such as ARLs (ARF-like proteins). ARFs are essential for vesicle coating and uncoating in all eukaryotic cells, while ARLs play more diverse roles. Nine proteins, six of them highly similar, are possible ARFs, three are putative ARL orthologues and the remainder were designated ARF-related proteins. The functions of the six highly similar, putative ARFs in whole plant development were probed by suppressing their expression with antisense. Antisense plants were severely stunted because cell production rate and final cell size were both reduced. Changed time-to-flowering, apical dominance, and fertility may reflect alterations to hormonal and other signalling pathways with which ARFs may interact. No gross changes in targeting or compartmentalization were seen in antisense plants containing GFP targeted to the ER and Golgi and changes in cell wall composition were limited to increases in some non-cellulosic polysaccharides and a relatively small decrease in cellulose. The reasons why these effects are less drastic than the effects on endomembranes and wall composition that are seen in short-term experiments with brefeldin A and with dominant negative ARF mutants are discussed.     

Gas chromatography-mass spectrometry analysis of indoleacetic acid and tryptophan following aqueous chloroformate derivatisation of Rhizobium exudates.

A new method for preparing alkyl esters of indole-3-acetic acid (IAA) in aqueous solution is adapted from the chloroformate method originally described by Husek for the analysis of amino acids. This method has the significant advantage of avoiding the generation and use of diazomethane, and is done in aqueous solution without the need to dry the sample with concomitant non-specific losses of IAA. The effectiveness of this method is demonstrated by its use in an isotope dilution gas chromatography-mass spectrometry (GC-MS) assay of IAA and L-tryptophan (Trp) in the culture supernatant of a series of Sinorhizobium meliloti and Rhizobium leguminosarum bv. trifolii strains that can interact with rice to either enhance or inhibit rice plant growth. We were testing the hypothesis that the rice growth inhibition was related to the biosynthesis of IAA. It was found that S. meliloti and Rhizobium strains produced high amounts of IAA in Trp supplemented BIII minimal medium compared to BIII media. All the strains produced more than the minimum amount of IAA required to inhibit rice growth and thus IAA is not the major inhibitory factor of rice seedling growth from S. meliloti and Rhizobium strains.