Assessment of a model for intron RNA secondary structure relevant to RNA self-splicing--a review

A widespread class of introns is characterized by a particular RNA secondary structure, based upon four conserved nucleotide sequences. Among such "class I" introns are found the majority of introns in fungal mitochondrial genes and the self-splicing intron of the large ribosomal RNA of several species of Tetrahymena. A model of the RNA secondary structure, which must underlie the self-splicing activity, is here evaluated in the light of data on 16 further introns. The main body or "core structure" of the intron always consists of the base-paired regions P3 to P9 with the associated single-stranded loops, with P2 present also in most cases. Two minority sub-classes of core structure occur, one of which is typical of introns in fungal ribosomal RNA. Introns in which the core structure is close to the 5' splice site all have an internal guide sequence (IGS) which can pair with exon sequences adjacent to the 5' and 3' splice sites to align them precisely, as proposed by Davies et al. [Nature 300 (1982) 719-724]. In these cases, the internal guide model allows us to predict correctly the exact location of splice sites. All other introns probably use other mechanisms of alignment. This analysis provides strong support for the RNA splicing model which we have developed.     

Magnetic separation in biotechnology

New developments in magnetic labelling techniques for cells and microspheres have extended the useful range of magnetic separation, particularly high gradient magnetic separation, into biotechnical areas. The basic magnetic principles involved are reviewed and representative samples of labelling techniques and results drawn from the past three years are presented. Illustrative examples of large scale operation in other industries are also presented, demonstrating the potential of the biotechnological applications.     

Studies on bone marrow histogenesis: Morphometric and autoradiographic studies of regenerating marrow stroma in extramedullary autoimplants and after evacuation of marrow cavity

Summary The marrow cavity of the rat tibia was mechanically evacuated and autoimplanted to the subcutaneous tissue. The regenerative process which restored the integrity of marrow stroma and hemopoiesis, was morphometrically evaluated in whole mount of tibia. Following evacuation, the clot filled the cavity. The granulation tissue then appeared and expanded, penetrating and replacing the clot. The fibroblasts of the granulation tissue differentiated into osteoblasts forming osteoid bone. Within its interstices, the primordial marrow consisting of loose connective tissue and vascular sinuses appeared and hemopoiesis resumed. Expansion of hemopoiesis resulted in the resorption of bone and within three weeks the tibial cavity was restored to the pre-evacuation state.Autoradiography indicated that the labeling index was initially high in fibroblasts and osteoblasts but was subsequently reduced while it increased in osteocytes, cells of Haversian canals, stromal and hemopoietic cells of marrow. The finding is in disagreement with the view that the regenerative process originates from the Haversian canal. When the label was introduced on day 4 post-operatively, it subsequently appeared in osteocytes, cells of Haversian canal, stromal elements of the marrow, but not in the hemopoietic cells. This indicates complete dissociation of marrow stroma and hemopoietic stem cell.Supported by NASA Contract NSG 9061. Mehdi Tavassoli is the recipient of a CRD Award AM-70551     

Characterization of human blood platelet membrane proteins and glycoproteins by their isoelectric point (pI) and apparent molecular weight using two-dimensional electrophoresis and surface-labelling techniques

Intact human blood platelets were radioactively labelled at the surface by techniques specific for proteins or glycoproteins. Labelled platelet samples were analyzed by a high-resolution two-dimensional separation system involving isoelectric focusing in the first dimension and discontinuous sodium dodecyl sulphate-polyacrylamide gel electrophoresis in the second. The major platelet membrane glycoprotein (GP) bands (Ib, IIb, IIIa and IIIb) were found to be highly heterogeneous even after removal of terminal sialic acid residues. Lactoperoxidase-catalyzed iodination of platelets showed that the major labelled proteins (Ib, IIb, IIIa and IIIb) had altered isoelectric points ($\text{p}\text{I}$) and molecular weights after neuraminidase treatment. A number of membrane glycoproteins previously undetected by one-dimensional gel electrophoresis were demonstrated and good evidence provided that the major platelet surface proteins are glycosylated.     

Intracellular distribution and origin of sterols in Calendula officinalis leaves

In Calendula officinalis leaves 66% of all steryl forms are present in the ‘microsomal fraction’ (IV), 24% in the mitochondrial and Golgi membranes (III), 5% in the ‘chloroplast’ (II), 4% in the ‘cell wall and membrane’ (I) fraction and 1%. in the cytosol. Free sterols, their esters, glycosides and acylated glycosides are present in varying proportions in all cellular subtractions. Mevalonate-[2¹⁴C] labelling of sterols derived from various steryl forms showed that free sterols and all their derivatives, i.e. steryl esters and glucosides, are formed in fraction IV and are then translocated to other organelles. Fraction III is the main site of glycosylation of transported sterols as well as of acylation of steryl glycosides.     

Phytochrome control of amino acid synthesis in cotyledons of Sinapis alba

Continuous far red light, acting through phytochrome, stimulates the rate of incorporation of density label into amino acids in the cotyledons of Sinapis alba. It is shown that such stimulation leads to increased incorporation of label into proteins. This has important consequences for experiments in which rates of enzyme synthesis in light treated and dark grown plants are compared by labelling methods. The results of some such experiments are re-evaluated.     

Acute abdominal vagotomy reduces drinking to peripheral but not central angiotensin II

Rats were tested two or three days after bilateral abdominal vagotomy or a laparotomy control procedure for their drinking responses to subcutaneous (1 mg-kg-1) or intracerebroventricular (100 ng) injections of angiotensin II. Vagotomy delayed the initiation of drinking and decreased 60-min water intake after subcutaneous, but not after intracerebroventricular, angiotensin II. This is the shortest postoperative interval in which the decrease in drinking after systemic injection of angiotensin II by abdominal vagotomy has been observed. The failure of vagotomy to decrease the response to intracerebroventricular angiotensin II demonstrates that the deficit after subcutaneous injection was not a nonspecific effect of recent vagotomy. These results, therefore, suggest that the abdominal vagus is necessary for normal drinking in response to circulating angiotensin II. Furthermore, the selective and acute onset of the deficit is consistent with the loss of a specific, rather than tonic facilitatory, vagal mechanism for drinking after elevation of circulating angiotensin II levels. Finally, the results imply that the physiological mechanisms which mediate the drinking responses to central and peripheral angiotensin are not identical.     

Labelling the blueberry leaftier, Croesia curvalana with foliar sprays of rubidium chloride

The feasibility of labelling blueberry leaftier by rearing the larvae on blueberry plants treated with foliar sprays of rubidium chloride (RbCl) at concentrations of 1000, 5000, 10 000 and 20 000 ppm were assessed. RbCl sprays above 5000 ppm significantly reduced survivorship to adult stage. The adult longevity, fecundity and mating were not affected when the larvae were reared on foliage treated with 5000 ppm RbCl solution. Reciprocal matings of 5000 ppm treated moths with untreated moths revealed transfer of label above the 0.1 µg Rb/insect threshold level from treated males to untreated females (in 8 out of 13 pairs) and vice versa (in 1 out of 9 pairs). Considerable loss of Rb (56–64%) occurred from the leaves over a 15 day period. All of the moths and pupae collected from the RbCl treated plots in 1989 and 1990 respectively, had a Rb content higher than the threshold level. In a preliminary dispersal study, marked male and female moths were found in sweepnet samples collected 20 and 60 m from the centre of the treated field. Labelled male moths were also captured in pheromone traps arranged in a circle, 40 m from the treated plot.