The envelope of Micrococcus radiodurans: isolation, purification, and preliminary analysis of the wall layers

Two methods are presented that separate the complex envelope of Micrococcus radiodurans, strain Sark, into its constituent layers. The first involved treating whole cells with 0.025 M Tris buffer (pH 7.5) containing 2 mM of calcium and 3 mM of magnesium, resulting in the degradation of an intermediate ('compartmentalized') layer and consequent sloughing of the outer subunit and interior layers to form vesicles. This treatment also appears to show that the interior layer may be connected with the peptidoglycan-containing 'holey' layer. The second method involves treating whole cells with benzene followed by sonication; the results suggested that this treatment only released the outer layers from the 'compartmentalized' layer and did not degrade layers. Following benzene treatment, digestion of the 'compartmentalized' layer with cold sodium dodecyl sulfate (SDS) released the 'holey' layer. Electrophoretic analysis of some of the isolated layer preparations suggested that the subunit layer consisted of three major proteins of 90 000, 92 000, and 94 000 molecular weight, one minor protein of 100 000, a small amount of carbohydrate associated with the 94 000 protein, and a small amount of a 55 000 lipoprotein. The interior layer contained at least 10 proteins and may be attached to the peptidoglycan-containing 'holey' layer by means of the 55 000 lipoprotein.     

Effect of polyamines on cyclic AMP-dependent and independent protein kinases from mouse epidermis.

Soluble extracts from mouse epidermis contained both cyclic AMP-dependent and independent protein kinases which could be separated by DEAE-Sephadex chromatography. The cyclic AMP-dependent histone kinase activity was inhibited by millimolar concentrations of the polyamines putrescine, spermidine and spermine. Similar concentrations of polyamines stimulated the cyclic AMP-independent phosphorylation of casein. The polyamines did not inhibit cyclic AMP binding by soluble epidermal extracts.     

The conversion of benzo(alpha)pyrene 4,5-oxide into 4-hydroxybenzo(alpha)pyrene in the presence of polyriboguanylic acid.

Incubation of benzo[alpha] pyrene 4,5-oxide with poly(G) in neutral aqueous ethanol resulted in the formation of covalent adducts and in the production of free 4-hydroxybenzo[alpha]pyrene. This phenol, which was identified by its UV spectral properties and by its chromatographic characteristics, was also formed but at a much slower rate when the epoxide was incubated with DNA or with GMP. Phenol formation was not detected when benzo[alpha]-pyrene 4,5-oxide was incubated for prolonged periods in the presence of poly(A), poly(C) or poly(U) or in the absence of nucleic acid. Formation of 4-hydroxybenzo[alpha] pyrene from the epoxide in the presence of poly(G) was not accompanied by detectable base modifications or by breakage of phosphodiester linkages.     

Supergenes in polymorphic land snails. I. Partula taeniata.

The general colour of the shell in Partula taeniata is controlled by at least two loci. One of these (C) has a series of six alleles which determine the yellow (Y) and neutral brown (N) series of colours. Alleles for darker colours are dominant to those for lighter colours, but dominance is not always complete. The pink (P) colours are determined by a second locus (P) which modified the expression of the lighter alleles of the C locus. Orangeshell colour segregates with yellow but its allelic relationship is unknown. Colour of the lip is controlled by a locus (L) with pink lip dominant to white lip. The colour of the spire is determined by a locus (S) with dark (N4) spire dominant to light spire. An intermediate spire colour shows the same pattern of inheritance and may represent the effect of another allele. Banding of the shell is dominant to absence of bands, with two loci (B1 and B2) determining the type of banding. An allele at B1 produces the frenata pattern; an allele at B2 produces zonata; together they produce lyra. All the loci for which linkage data are available are linked so strongly that the whole array may be considered a supergene. Self-fertilisation takes place primarily during early reproductive life. About 20 per cent of the young of the first mating of an individual are produced by selfing, but over the whole reproductive span the frequency is only about 2-5 per cent. There is inconclusive evidence for heterozygote advantage of banded individuals.     

Quantification of microtubule stutters: dynamic instability behaviors that are strongly associated with catastrophe

Microtubules (MTs) are cytoskeletal fibers that undergo dynamic instability (DI), a remarkable process involving phases of growth and shortening separated by stochastic transitions called catastrophe and rescue. Dissecting DI mechanism(s) requires first characterizing and quantifying these dynamics, a subjective process that often ignores complexity in MT behavior. We present a Statistical Tool for Automated Dynamic Instability Analysis (STADIA) that identifies and quantifies not only growth and shortening, but also a category of intermediate behaviors that we term “stutters.” During stutters, the rate of MT length change tends to be smaller in magnitude than during typical growth or shortening phases. Quantifying stutters and other behaviors with STADIA demonstrates that stutters precede most catastrophes in our in vitro experiments and dimer-scale MT simulations, suggesting that stutters are mechanistically involved in catastrophes. Related to this idea, we show that the anticatastrophe factor CLASP2γ works by promoting the return of stuttering MTs to growth. STADIA enables more comprehensive and data-driven analysis of MT dynamics compared with previous methods. The treatment of stutters as distinct and quantifiable DI behaviors provides new opportunities for analyzing mechanisms of MT dynamics and their regulation by binding proteins.     

Microbiota within the perennial ice cover of Lake Vida, Antarctica

Lake Vida, located in the McMurdo Dry Valleys, Antarctica, is an 'ice-sealed' lake with approximately 19 m of ice covering a highly saline water column (approximately 245 ppt). The lower portions of the ice cover and the lake beneath have been isolated from the atmosphere and land for circa 2800 years. Analysis of microbial assemblages within the perennial ice cover of the lake revealed a diverse array of bacteria and eukarya. Bacterial and eukaryal denaturing gradient gel electrophoresis phylotype profile similarities were low (<59%) between all of the depths compared (five depths spanning 11 m of the ice cover), with the greatest differences occurring between surface and deep ice. The majority of bacterial 16S rRNA gene sequences in the surface ice were related to Actinobacteria (42%) while Gammaproteobacteria (52%) dominated the deep ice community. Comparisons of assemblage composition suggest differences in ice habitability and organismal origin in the upper and lower portions of ice cover. Specifically, the upper ice cover microbiota likely reflect the modern day transport and colonization of biota from the terrestrial landscape, whereas assemblages in the deeper ice are more likely to be persistent remnant biota that originated from the ancient liquid water column of the lake that froze.     

siRNA-directed inhibition of HIV-1 infection

RNA interference silences gene expression through short interfering 21 23-mer double-strand RNA segments that guide mRNA degradation in a sequence-specific fashion. Here we report that siRNAs inhibit virus production by targeting the mRNAs for either the HIV-1 cellular receptor CD4, the viral structural Gag protein or green fluorescence protein substituted for the Nef regulatory protein. siRNAs effectively inhibit pre- and/or post-integration infection events in the HIV-1 life cycle. Thus, siRNAs may have potential for therapeutic intervention in HIV-1 and other viral infections.