Replication slippage may cause parallel evolution in the secondary structures of mitochondrial transfer RNAs

Presence of the dihydrouridine (D) stem in the mitochondrial cysteine tRNA is unusually variable among lepidosaurian reptiles. Phylogenetic and comparative analyses of cysteine tRNA gene sequences identify eight parallel losses of the D-stem, resulting in D-arm replacement loops. Sampling within the monophyletic Acrodonta provides no evidence for reversal. Slipped-strand mispairing of noncontiguous repeated sequences during replication or direct replication slippage can explain repeats observed within cysteine tRNAs that contain a D-arm replacement loop. These two mechanisms involving replication slippage can account for the loss of the cysteine tRNA D-stem in several lepidosaurian lineages, and may represent general mechanisms by which the secondary structures of mitochondrial tRNAs are altered.      

Expression, Purification, and Inhibitory Properties of Human Proteinase Inhibitor 8

In a previous report, the cDNA for human proteinase inhibitor 8 (PI8) was first identified, isolated, and subcloned into a mammalian expression vector and expressed in baby hamster kidney cells. Initial studies indicated that PI8 was able to inhibit the amidolytic activity of trypsin and form an SDS-stable approximately 67-kDa complex with human thrombin [Sprecher, C. A., et al. (1995) J. Biol Chem. 270, 29854-29861]. In the present study, we have expressed recombinant PI8 in the methylotropic yeast Pichia pastoris, purified the inhibitor to homogeneity, and investigated its ability to inhibit a variety of proteinases. PI8 inhibited the amidolytic activities of porcine trypsin, human thrombin, human coagulation factor Xa, and the Bacillus subtilis dibasic endoproteinase subtilisin A through different mechanisms but failed to inhibit the Staphylococcus aureus endoproteinase Glu-C. PI8 inhibited trypsin in a purely competitive manner, with an equilibrium inhibition constant (Ki) of less than 3.8 nM. The interaction between PI8 and thrombin occurred with a second-order association rate constant (kassoc) of 1.0 x 10(5) M-1 s-1 and a Ki of 350 pM. A slow-binding kinetics approach was used to determine the kinetic constants for the interactions of PI8 with factor Xa and subtilisin A. PI8 inhibited factor Xa via a two-step mechanism with a kassoc of 7.5 x 10(4) M-1 s-1 and an overall Ki of 272 pM. PI8 was a potent inhibitor of subtilisin A via a single-step mechanism with a kassoc of 1.16 x 10(6) M-1 s-1 and an overall Ki of 8.4 pM. The interaction between PI8 and subtilisin A may be of physiological significance, since subtilisin A is an evolutionary precursor to the intracellular mammalian dibasic processing endoproteinases.     

An Ethnographic Filmflam: Giving Gifts, Doing Research, and Videotaping the Native Subject/Object

Using the discussion of self-reflexivity as an organizing principle, this article examines how mobilizing digital video technology during fieldwork opens up empirical and theoretical space for reconceptualizing the relationship between anthropologists and informants. Placing the field of visual anthropology into critical conversation with long-standing theoretical arguments about the objectivist limitations of native anthropologists, I argue that the slipperiness of nativity as an anthropological designation helps to provide analytical tools for examining filmmaking as a kind of gift-giving process between native ethnographic filmmakers and the subjects of their films. This article highlights some of the ways in which my own filmic and videographic exploits in Harlem, New York, mark integral connections between seeing and being the proverbial other, probing social exchanges predicated on the usefulness of low-budget digital technology as a means of fostering politically and epistemologically valuable ethnographic collaborations.     

Pathways that control cortical F-actin dynamics during secretion

Chromaffin cells possess a mesh of filamentous actin underneath the plasma membrane which acts as a barrier to the chromaffin vesicles access to exocytotic sites. Disassembly of cortical F-actin in response to stimulation allows the movement of vesicles from the reserve pool to the release-ready vesicle pool and, therefore, to exocytotic sites. The dynamics of cortical F-actin is controlled by two mechanisms: a) stimulation-induced Ca²⁺ entry and scinderin activation and b) protein kinase C (PKC) activation and MARCKS phosphorylation as demonstrated here by experiments with recombinant proteins, antisense olygodeoxynucleotides and vector mediated transient expressions. Under physiological conditions (i.e., cholinergic receptor stimulation followed by Ca²⁺ entry), mechanism (a) is the most important for the control of cortical F-actin network whereas when Ca²⁺ is released from intracellular stores (i.e., histamine stimulation) cortical F-actin is regulated mainly by mechanism b.     

Comparison of the effect of hormones on the hormone synthesis of Tetrahymena in medium or salt solution

Tetrahymena pyriformis was maintained in TYM (tryptone‐yeast medium) as well as in Losina salt solution. One hour treatment of 10^?15 M histamine, serotonin or insulin was given before the histamine, serotonin, triiodothyronine and adrenocorticotropin contents of the cells were measured by flow cytometry after immunocytochemical staining. Maintenance in salt solution increased the hormone level in the cells, and use of the treatment hormone treatments further increased the endogenous hormone content relative to that in medium. The cells in salt mimic better the natural conditions, which means that the effects of hormones under more natural conditions are expressed to a greater extent than the exogenously given hormones in TYM typically used under laboratory conditions. Intercellular hormonal communication between the cells of a Tetrahymena population might assist in the survival of the individual cells.     

A three-species biofilm model for the evaluation of enamel and dentin demineralization

Although Streptococcus mutans biofilms have been useful for evaluating the cariogenic potential of dietary carbohydrates and the effects of fluoride on dental demineralization, a more appropriate biofilm should be developed to demonstrate the influence of other oral bacteria on cariogenic biofilms. This study describes the development and validation of a three-species biofilm model comprising Streptococcus mutans, Actinomyces naeslundii, and Streptococcus gordonii for the evaluation of enamel and dentin demineralization after cariogenic challenges and fluoride exposure. Single- or three-species biofilms were developed on dental substrata for 96?h, and biofilms were exposed to feast and famine episodes. The three-species biofilm model produced a large biomass, mostly comprising S. mutans (41%) and S. gordonii (44%), and produced significant demineralization in the dental substrata, although enamel demineralization was decreased by fluoride treatment. The findings indicate that the three-species biofilm model may be useful for evaluating the cariogenic potential of dietary carbohydrates other than sucrose and determining the effects of fluoride on dental substrata.     

Towards a framework for the study of the neural correlates of aesthetic preference

Aiming to provide a tentative framework for the study of the neural correlates of aesthetic preference, we review three recent neuroimaging studies carried out with the purpose of locating brain activity associated with decisions about the beauty of visual stimuli (Cela-Conde et al., 2004; Kawabata and Zeki, 2004; Vartanian and Goel, 2004). We find that the results of the three studies are not in line with previous neuropsychological data. Moreover, there are no coincidences among their results. However, when they are mapped on to Chatterjee's (2003) neuropsychological model of aesthetic preference it becomes clear that neuroimaging data are not contradictory, but complementary, and their interpretation is enriched. The results of these studies suggest that affective processes have an important role in aesthetic preference, and that they are integrated with cognitive processes to reach a decision regarding the beauty of visual stimuli. Future studies must aim to clarify whether certain methodological procedures are better suited to study any of the particular cognitive operations involved in aesthetic preference, and ascertain the extent to which the proposed framework is compatible with the aesthetic appreciation of musical stimuli.     

On the Trail of Atmospheric Mutagens and Carcinogens: A Combined Chemical/Microbiological Approach

Major ecological problems of our polluted troposphere includeairborne toxic chemicals, acid rain and photochemical smog,all three of which are now recognized as being closely relatedchemical phenomena. We also recognize that inorder to developcost-effective strategies for their control, which protect publichealth and the environment, there must be close scientific interactionsbetween chemists and biological scientists. For example, ofrapidly emerging importance is the development of risk assessmentevaluations for specific aspects of each of these problem areas.In preparing such assessments, chemists must define the "exposure,"and biological scientists the "effects." In this paper, I discuss an example of how such close interactionsproved indispensible in our search for atmospheric mutagensand carcinogens. Thus, an integrated chemical/ microbiologicalprocedure for the isolation and identificationof particulatechemical mutagens in respirable diesel soot and ambient particlesis described. Emphasis is placed on our use of the short-term,Ames Salmonella typhimurium bacterial mutagenicity test as arapid, and relatively inexpensive, means of following the biologicalactivities of these environmental mutagens through the chemicalsteps of their separation, isolation and identification fromhighly complex environmental samples. Possible mechanisms offormation of these particulate mutagens are discussed. Theyinclude the reactions of polycyclic aromatic hydrocarbons presenton the surfaces of combustion-generated particles with gaseousco-pollutants such as nitrogen dioxide plus nitric acid, andozone. In discussing this research on a societally "relevant" problem,we illustrate the importance of "Science as a Way of Knowing."We further suggest that this integrated approach to scientificproblem solving by chemical and biological scientists mightserve as an example of a discussion topic on human ecology forundergraduate courses in the natural sciences.