Non-mendelian inheritance of mitochondrial DNA and ribosomal DNA in the myxomycete, Didymium iridis

Summary The inheritance of both the mitochondrial DNA (mtDNA) and the nuclear-encoded extrachromosomal ribosomal DNA (rDNA) has been studied in the myxomycete, Didymium iridis, by DNA-DNA hybridization of labeled probes to total DNA at various stage of the life cycle. Both the mtDNA and rDNA populations rapidly become homogeneous in individuals, but there is a qualitative difference in the patterns of inheritance of these two molecules. One parental rDNA type was preferentially inherited in all crosses; selective replication of this molecule is tentatively proposed as the mechanism of inheritance. In contrast, either parental mtDNA type could be inherited. Since the inherited population of parental mtDNA molecules are not partitioned into cells in this coenocytic organism, no known mechanism of inheritance can explain the rapid and apparently random loss of one parental mtDNA type in individuals.     

Identification of procalcitonin in a rat medullary thyroid carcinoma cell line

As a first step in studying the biosynthesis of the peptide hormone calcitonin, we have identified procalcitonin species in CA-77 cells, a newly developed rat medullary thyroid carcinoma cell line. mRNA extracted from the cells directed the synthesis of a putative procalcitonin in a reticulocyte lysate translation system containing microsomal membranes. Both this species and a radiolabeled form of immunoreactive calcitonin from intact cells had the same retention time during reverse phase high performance liquid chromatography. The putative cellular procalcitonin was also immunoprecipitated by antiserum to a synthetic peptide whose sequence constitutes the COOH-terminal 16 residues of preprocalcitonin. The polypeptide had a Mr = 13,400, as estimated by gel filtration chromatography under denaturing conditions. Microsequencing of the [35S]methionine-labeled polypeptide indicated that residues 13, 32, and 34 of procalcitonin were methionine. Similar analysis of the peptide labeled with [3H]proline indicated that residues 2 and 11 of the precursor were proline. The positions of methionine and proline could be aligned in a unique manner with the NH2-terminal half of the preprocalcitonin sequence inferred from cDNA analyses. These results indicate that procalcitonin consists of 111 amino acids and suggest that a 25-residue signal sequence is cotranslationally cleaved from preprocalcitonin. From the procalcitonin sequence we can now predict the sequence of likely biosynthetic intermediates and mature secretory products derived from the NH2-terminal as well as COOH-terminal regions of the precursor.     

The innervation of the pyloric region of the crab,Cancer borealis: Homologous muscles in decapod species are differently innervated

Summary The muscles of the pyloric region of the stomach of the crab,Cancer borealis, are innervated by motorneurons found in the stomatogastric ganglion (STG). Electrophysiological recording and stimulating techniques were used to study the detailed pattern of innervation of the pyloric region muscles. Although there are two Pyloric Dilator (PD) motorneurons in lobsters, previous work reported four PD motorneurons in the crab STG (Dando et al. 1974; Hermann 1979a, b). We now find that only two of the crab PD neurons innervate muscles homologous to those innervated by the PD neurons in the lobster,Panulirus interrruptus. The remaining two PD neurons innervate muscles that are innervated by pyloric (PY) neurons inP. interruptus. The innervation patterns of the Lateral Pyloric (LP), Ventricular Dilator (VD), Inferior Cardiac (IC), and PY neurons were also determined and compared with those previously reported in lobsters. Responses of the muscles of the pyloric region to the neurotransmitters, acetylcholine (ACh) and glutamate, were determined by application of exogenous cholinergic agonists and glutamate. The effect of the cholinergic antagonist, curare, on the amplitude of the excitatory junctional potentials (EJPs) evoked by stimulation of the pyloric motor nerves was measured. These experiments suggest that the differences in innervation pattern of the pyloric muscles seen in crab and lobsters are also associated with a change in the neurotransmitter active on these muscles. Possible implications of these findings for phylogenetic relations of decapod crustaceans and for the evolution of neural circuits are discussed.Abbreviations ACh acetylcholine - Carb carbamylcholine - cpv muscles of the cardio-pyloric valve - cpv7n nerve innervating muscle cpv7 - cv muscles of the ventral cardiac ossicles - cv1n nerve innervating muscle cvl - cv2n nerve innervating muscle cv2 - EJP excitatory junctional potential - IC inferior cardiac neuron - IV inferior ventricular neuron - IVN inferior ventricular nerve - LP lateral pyloric neuron - LPG lateral posterior gastric neuron - lvn lateral ventricular nerve - mvn medial ventricular nerve - p muscles of the pylorus - PD pyloric dilator neuron - PD _in intrinsic PD neuron - PD _ex extrinsic PD neuron - pdn pyloric dilator nerve - PY pyloric neuron - pyn pyloric nerve - STG stomatogastric ganglion - VD ventricular dilator neuron     

2-Aminoethylphosphonic acid concentrations in some rumen ciliate protozoa.

The concentration of 2-aminoethylphosphonic acid has been measured in seven genera of rumen ciliate protozoa. Expressed as milligrams per gram of total nitrogen, 2-aminoethylphosphonic acid concentrations ranged from 17.2 in Ophryoscolex spp. to 72.4 in Eremoplastron spp.     

Mutational analysis of the histidine operon promoter of Salmonella typhimurium.

We isolated a collection of 67 independent, spontaneous Salmonella typhimurium his operon promoter mutants with decreased his expression. The mutants were isolated by selecting for resistance to the toxic lactose analog o-nitrophenyl-beta-D-thiogalactoside in a his-lac fusion strain. The collection included base pair substitutions. small insertions, a deletion, and one large insertion identified as IS30 (IS121), which is resident on the Mu d1 cts(Apr lac) phage used to construct the his-lac fusion. Of the 37 mutations that were sequenced, 14 were unique. Six of the 14 were isolated more than once, with the IS30 insertion occurring 16 times. The mutations were located throughout the his promoter region, with two in the conserved - 35 hexamer sequence, four in the conserved - 10 hexamer sequence (Pribnow box), seven in the spacer between the - 10 and -35 hexamer sequences, and the IS30 insertions just upstream of the -35 hexamer sequence. Four of the five substitution mutations changed a consensus base pair recognized by E sigma 70 RNA polymerase in the -10 or -35 hexamer. Decreased his expression caused by the 14 different his promoter mutations was measured in vivo. Relative to the wild-type promoter, the mutations resulted in as little as a 4-fold decrease to as much as a 357-fold decrease in his expression, with the largest decreases resulting from changes in the most highly conserved features of E sigma 70 promoters.     

Uniform 15N labeling of a fungal peptide: the structure and dynamics of an alamethicin by 15N and 1H NMR spectroscopy.

An alamethicin, secreted by the fungus Trichoderma viride and containing a glutamine at position 18 instead of the usual glutamic acid, has been uniformly labeled with 15N and purified by HPLC. The extent of 15N incorporation at individual backbone and side-chain sites was found to vary from 85% to 92%, as measured by spin-echo difference spectroscopy. The proton NMR spectrum of the peptide dissolved in methanol was assigned using correlation spectroscopies and nuclear Overhauser enhancements (NOE) measured in the rotating frame. The 15N resonances were assigned by the 2D 1H-15N correlation via heteronuclear multiple-quantum coherence experiment. NOEs and 3JNHC alpha H coupling constants strongly suggest that, in methanol, from Aib-3 to Gly-11, the peptide adopts a predominantly helical conformation, in agreement with previous 1H NMR studies [Esposito, G., Carver, J.A, Boyd, J., & Campbell, I.D. (1987) Biochemistry 26, 1043-1050; Banerjee, U., Tsui, F.-P., Balasubramanian, T.N., Marshall, G.R., & Chan, S I. (1983) J. Mol. Biol. 165, 757-775]. The conformation of the carboxyl terminus (12-20) is less well determined, partly because the amino acid composition reduces the number of NOEs and coupling constants which can be determined by 1H NMR spectroscopy. The 3JNHC alpha H in the C-terminus suggest the possibility of conformational averaging at Leu-12, Val-15, and Gln-19, an interpretation which is supported by a recent molecular dynamics simulation of the peptide [Fraternalli, F. (1990) Biopolymers 30, 1083-1099].(ABSTRACT TRUNCATED AT 250 WORDS)     

Death of the Escherichia coli K-12 strain W3110 in soil and water.

Whether Escherichia coli K-12 strain W3110 can enter the "viable but nonculturable" state was studied with sterile and nonsterile water and soil at various temperatures. In nonsterile river water, the plate counts of added E. coli cells dropped to less than 10 CFU/ml in less than 10 days. Acridine orange direct counts, direct viable counts, most-probable-number estimates, and PCR analyses indicated that the added E. coli cells were disappearing from the water in parallel with the number of CFU. Similar results were obtained with nonsterile soil, although the decline of the added E. coli was slower. In sterile water or soil, the added E. coli persisted for much longer, often without any decline in the plate counts even after 50 days. In sterile river water at 37 degrees C and sterile artificial seawater at 20 and 37 degrees C, the plate counts declined by 3 to 5 orders of magnitude, while the acridine orange direct counts remained unchanged. However, direct viable counts and various resuscitation studies all indicated that the nonculturable cells were nonviable. Thus, in either sterile or nonsterile water and soil, the decline in plate counts of E. coli K-12 strain W3110 is not due to the cells entering the viable but nonculturable state, but is simply due to their death.     

Incomplete protection, but suppression of virus burden, elicited by subunit simian immunodeficiency virus vaccines.

We compared the efficacy of immunization with either simian immunodeficiency virus (SIV) Env glycoprotein (Env), Env plus Gag proteins (Gag-Env), or whole inactivated virus (WIV), with or without recombinant live vaccinia vector (VV) priming, in protecting 23 rhesus macaques (six vaccine and two control groups) from challenge with SIVmac251 clone BK28. Vaccination elicited high titers of syncytium-inhibiting and anti-Env (gp120/gp160) antibodies in all vaccinated macaques and anti-Gag (p27) antibodies in groups immunized with WIV or Gag-Env. Only WIV-immunized macaques developed anticell (HuT78) antibodies. After homologous low-dose intravenous virus challenge, we used frequency of virus isolation, provirus burden, and change in antibody titers to define four levels of resistance to SIV infection as follows. (i) No infection ("sterilizing" immunity) was induced only in WIV-immunized animals. (ii) Abortive infection (strong immunity) was defined when virus or provirus were detected early in the postchallenge period but not thereafter and no evidence of virus or provirus was detected in terminal tissues. This response was observed in two animals (one VV-Env and one Gag-Env). (iii) Suppression of infection (incomplete or partial immunity) described a gradient of virus suppression manifested by termination of viremia, declining postchallenge antibody titers, and low levels (composite mean = 9.1 copies per 10(6) cells) of provirus detectable in peripheral blood mononuclear cells or lymphoid tissues at termination (40 weeks postchallenge). This response occurred in the majority (8 of 12) of subunit-vaccinated animals. (iv) Active infection (no immunity) was characterized by persistent virus isolation from blood mononuclear cells, increasing viral antibody titers postchallenge, and high levels (composite mean = 198 copies per 10(6) cells) of provirus in terminal tissues and blood. Active infection developed in all controls and two of three VV-Gag-Env-immunized animals. The results of this study restate the protective effect of inactivated whole virus vaccines produced in heterologous cells but more importantly demonstrate that a gradient of suppression of challenge virus growth, reflecting partial resistance to SIV infection, is induced by subunit vaccination. The latter finding may be pertinent to studies with human immunodeficiency virus vaccines, in which it is plausible that vaccination may elicit significant suppression of virus infection and pathogenicity rather than sterilizing immunity.     

Soil P resources,plant growth and rooting characteristics in nutrient poor upland grasslands

A field study was undertaken to establish the demand for P by mixed herbage, manipulated by cutting regimes, and the extent to which orthophosphate alone in soil solution could meet this demand from three cambisols derived from different parent materials. Differences in soil types were sufficient to produce significantly different rooting patterns at each site. Yields for 7-and 10-cm treatments generally exceeded those for swards cut to 2-and 4-cm. The highest yields were from plots cut once at the end of the season, or when herbage was cut in June and October only. Yields fell in the second season by an average of 30%. Two cuts in the season resulted in almost twice the P uptake compared with other treatments, leading to the view that a silage cut stimulated root growth. Rooting was deepest in Tarves Association soil (Dystric cambisol), densest in Insch Association soil (Eutric cambisol) and intermediate in Foudland Association soil (Dystric cambisol) but herbage yield at each site was similar. Whole season mean P and N content in roots ranged from 1.0 to 3.4 and from 8.1 to 27.9 mg g^–1 dry weight, respectively. The lowest values were in once cut herbage and were half those in herbage cut in June and October only. Data for the total P resources of the soils, extractable P, and shoot and root P at each site are presented together with data for P in soil solution (principally organic) from an associated soil solution study. There was a disparity between daily uptake and orthophosphate in soil solution. These findings suggested that it was probable that soluble organic forms of P are important for P nutrition in these nutrient poor soils, and could account for the excess of observed P uptake (from soils low in P) over that predicted by mechanistic mathematical models.