Do capuchin monkeys, Cebus apella, know what conspecifics do and do not see?

Capuchin monkeys were tested in five experiments in which two individuals competed over food. When given a choice between retrieving a piece of food that was visible or hidden from the dominant, subordinate animals preferred to retrieve hidden food. This preference is consistent with the hypotheses that either (1) the subordinate knew what the dominant could and could not see or (2) the subordinate was monitoring the behaviour of the dominant and avoiding the piece of food that it approached. To test between these alternatives, we released subordinates with a slight head start forcing them to make their choice (between a piece of food hidden or visible to the dominant) before the dominant entered the area. Unlike chimpanzees, Pan troglodytes, subordinates that were given a head start did not preferentially approach hidden pieces of food first. Therefore, our experiments provide little support for the hypothesis that capuchin monkeys are sensitive to what another individual does or does not see. We compare our results with those obtained with chimpanzees in the same paradigm and discuss the evolution of primate social cognition. Copyright 2003 The Association for the Study of Animal Behaviour. Published by Elsevier Science Ltd. All rights reserved.      

Measuring dynamics of caspase-8 activation in a single living HeLa cell during TNFalpha-induced apoptosis

In this study, we reported the first measurement of the dynamics of activation of caspase-8 in a single living cell. This measurement was conducted using a specially developed molecular sensor based on the FRET (fluorescence resonance energy transfer) technique. This sensor was constructed by fusing a CFP (cyan fluorescent protein) and a YFP (yellow fluorescent protein) with a linker containing a tandem caspase-8-specific cleavage site. The change of the FRET ratio upon cleavage was larger than 4-fold. Using this sensor, we found that during TNFalpha-induced apoptosis, the activation of caspase-8 was a slower process than that of caspase-3, and it was initiated much earlier than the caspase-3 activation. Inhibition of caspase-9 delayed the full activation of caspase-3 but did not affect the dynamics of caspase-8. Results of these single-cell measurements suggested that caspase-3 was activated by caspase-8 through two parallel pathways during TNFalpha-induced apoptosis in HeLa cells.     

A receptor linked to a Gi-family G-protein functions in initiating oocyte maturation in starfish but not frogs

The stimulation of oocyte maturation by 1-methyladenine in starfish, and by a steroid in frogs, has been proposed to involve G-protein-coupled receptors. To examine whether activation of receptors linked to G(i) or G(z) was sufficient to cause oocyte maturation, we expressed mammalian G(i)- and G(z)-linked receptors in starfish and frog oocytes. Application of the corresponding agonists caused meiosis to resume in the starfish but not the frog oocytes. We confirmed that the receptors were effectively expressed in the frog oocytes by using a chimeric G-protein, G(qi), that converts input from G(i)- and G(z)-linked receptors to a G(q) output and results in a contraction of the oocyte's pigment. These results argue against G(i) or G(z) functioning to cause maturation in frog oocytes. Consistently, maturation-inducing steroids did not cause pigment contraction in frog oocytes expressing G(qi), and G(z) protein was not detectable in frog oocytes. For starfish oocytes, however, our results support the conclusion that G(i) functions in 1-methyladenine signaling and suggest the possibility of using frog oocyte pigment contraction as an assay to identify the 1-methyladenine receptor. To test this concept, we coexpressed G(qi) and a starfish adenosine receptor in frog oocytes and showed that applying adenosine caused pigment contraction.     

Development and application of computer software for cell culture laboratory management

Prototype computer software for a Cell Culture Laboratory Management System (CCLMS) has been developed to relieve cell culture specialists of the burden of manual recordkeeping. Conventional data archives in cell culture laboratories are prone to error and expensive to maintain. The reliance upon cell culture to provide models for biochemical and molecular biological research serves to magnify errors at great expense. The CCLMS prototype encapsulates a modular software application that manages the many aspects of cell culture laboratory recordkeeping. A transaction-based database stores detailed information on subcultures, freezes and thaws, prints waterproof labels for culture vessels, and provides for immediate historical trace-back of any cultured cell line. Linked database files store information specific to an individual culture flask while removing redundancy between similar groups of flasks. A frozen cell log maintains locations of all vials within any type of cryogenic storage unit, locates spaces for newly frozen cell lines, and generates alphabetical or numerical reports. Finally, modules for maintaining cell counts, user records, and culture vessel specifications to support a comprehensive automation process are incorporated within this software. The developed CCLMS prototype has been demonstrated to be an adaptable, reliable tool for improving training, efficiency, and historical rigor for two independent cell culture facilities.     

Functional and molecular evidence for Na+-HCO3- cotransporter in porcine vas deferens epithelia

This study focused on the role ofsodium-bicarbonate cotransporter (NBC1) in cAMP-stimulated iontransport in porcine vas deferens epithelium. Ion substitutionexperiments in modified Ussing chambers revealed that cAMP-mediatedstimulation was dependent on the presence of Na⁺,HCO, and Cl for a full response.HCO-dependent current was unaffected byacetazolamide, bumetanide, or amiloride but was inhibited bybasolateral 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid.Na⁺-driven, HCO-dependent,stilbene-inhibitable anion flux was observed across the basolateralmembrane of selectively permeabilized monolayers. Results ofradiotracer flux studies suggest a4,4'-dinitrostilbene-2,2'-disulfonate-sensitive stoichiometry of 2 baseequivalents per Na⁺. Antibodies raised against rat kidneyNBC epitopes (rkNBC; amino acids 338-391 and 928-1035)identified a single band of ~145 kDa. RT-PCR detected NBC1 message inporcine vas deferens epithelia. These results demonstrate that vasdeferens epithelial cells possess the proteins necessary for thevectoral transport of HCO and that these mechanismsare maintained in primary culture. Taken together, the results indicatethat vas deferens epithelia play an active role in male fertility andhave implications for our understanding of the relationship betweencystic fibrosis and congenital bilateral absence of the vas deferens.

     

Two genetic circuits repress the Caenorhabditis elegans heterochronic gene lin-28 after translation initiation

The heterochronic gene lin-28 of the nematode Caenorhabditis elegans controls the relative timing of diverse developmental events during the animal's larval stages. lin-28 is stage-specifically regulated by two genetic circuits: negatively by the 22-nt RNA lin-4 and positively by the heterochronic gene lin-14. Here, we show that lin-28 is repressed during normal development by a mechanism that acts on its mRNA after translation initiation. We provide evidence that lin-14 inhibits a negative regulation that is independent of the lin-4 RNA and involves the gene daf-12, which encodes a nuclear hormone receptor. The lin-4-independent repression does not affect the initiation of translation on the lin-28 mRNA, and like the lin-4-mediated repression, acts through the gene's 3'-untranslated region. In addition, we find that lin-4 is not sufficient to cause repression of lin-28 if the lin-4-independent circuit is inhibited. Therefore, the lin-4-independent circuit likely contributes substantially to the down-regulation of lin-28 that occurs during normal development. The role of lin-4 may be to initiate or potentiate the lin-4-independent circuit. We speculate that a parallel lin-4-independent regulatory mechanism regulates the expression of lin-14.     

Fingerprinting closely related xanthomonas pathovars with random nonamer oligonucleotide microarrays

Current bacterial DNA-typing methods are typically based on gel-based fingerprinting methods. As such, they access a limited complement of genetic information and many independent restriction enzymes or probes are required to achieve statistical rigor and confidence in the resulting pattern of DNA fragments. Furthermore, statistical comparison of gel-based fingerprints is complex and nonstandardized. To overcome these limitations of gel-based microbial DNA fingerprinting, we developed a prototype, 47-probe microarray consisting of randomly selected nonamer oligonucleotides. Custom image analysis algorithms and statistical tools were developed to automatically extract fingerprint profiles from microarray images. The prototype array and new image analysis algorithms were used to analyze 14 closely related Xanthomonas pathovars. Of the 47 probes on the prototype array, 10 had diagnostic value (based on a chi-squared test) and were used to construct statistically robust microarray fingerprints. Analysis of the microarray fingerprints showed clear differences between the 14 test organisms, including the separation of X. oryzae strains 43836 and 49072, which could not be resolved by traditional gel electrophoresis of REP-PCR amplification products. The proof-of-application study described here represents an important first step to high-resolution bacterial DNA fingerprinting with microarrays. The universal nature of the nonamer fingerprinting microarray and data analysis methods developed here also forms a basis for method standardization and application to the forensic identification of other closely related bacteria.     

The Salton Sea as critical habitat to migratory and resident waterbirds

Concern about the Salton Sea ecosystem, based on potential impacts of increasing salinity, contaminants, disease outbreaks, and large die-offs of birds, is heightened because of tremendous prior loss and degradation of wetland habitat in western North America. In 1999, we used a variety of survey methods to describe patterns of abundance of birds at the Salton Sea and in adjacent habitats. Our results further documented the great importance of the Salton Sea within the Pacific Flyway to wintering, migratory, and breeding waterbirds. Exclusive of Eared Grebes, we estimated about 187000 individual waterbirds at the Salton Sea in January, 88000 in April, 170000 in August, and 261000 in November. Additional surveys of Eared Grebes in November and December suggested the total population of all waterbirds was about 434000 to 583000 in those months, respectively. We also documented breeding by about 14000 pairs of colonial waterbirds. Waterbirds were particularly concentrated along the northern, southwestern, southern, and southeastern shorelines and river deltas. By contrast, some species of wading birds (Cattle Egret, White-faced Ibis, Sandhill Crane) and shorebirds (Mountain Plover, Whimbrel, Long-billed Curlew) were much more numerous in agricultural fields of the Imperial Valley than in wetland habitats at the Sea. Various studies indicate the Salton Sea is of regional or national importance to pelicans and cormorants, wading birds, waterfowl, shorebirds, and gulls and terns. Important taxa are the Eared Grebe, American White Pelican, Double-crested Cormorant, Cattle Egret, White-faced Ibis, Ruddy Duck, Yuma Clapper Rail, Snowy Plover, Mountain Plover, Gull-billed, Caspian, and Black terns, and Black Skimmer. Proposed restoration projects should be carefully assessed to ensure they do not have unintended impacts and are not placed where large numbers of breeding, roosting, or foraging birds concentrate. Similarly, plans to enhance opportunities for recreation or commerce at the Sea should aim to avoid or minimize disturbance to birds. Future research should focus on filling gaps in knowledge needed to effectively conserve birds at the Salton Sea.