Effects of Season and Vegetation Type on Community Organization of Dung Beetles in a Tropical Dry Forest1

Dung beetles are important components of most terrestrial ecosystems. In tropical rain forests, dung beetle communities can be very rich in number of species and individuals, and they are known to be useful bioindicators of habitat disturbance. In contrast, very little is known about the organization of dung beetle communities in tropical dry forests. The aim of this study was to describe in detail the dung beetle community of a Mexican tropical dry forest and to assess the relative importance of rainfall seasonality and forest structure in affecting the temporal and spatial dynamics of this community. Dung beetles were captured with pitfall traps at the beginning of the rainy season, the middle of the rainy season, and the middle of the dry season, in two distinct forest types: deciduous forest (DF) and semideciduous forest (SDF) at the Estación de Biología Chamela. Both rainfall seasonality and forest structure affected the community organization of dung beetles. During both rainy periods, 14 species were captured, but only three during the dry season. Dung beetles captured during the dry season were only found in the SDF. When comparing the beginning and the middle of the rainy season, differences in abundance and guild structure were also observed between both periods and between forest types, but these differences were much less pronounced.     

Dispersión Primaria de Semillas por Primates y Dispersión Secundaria por Escarabajos Coprófagos en Tikal,Guatemala1

We linked primary dispersal by spider monkeys (Ateles geoffroyi) and howler monkeys (Alouatta pigra) to post‐dispersal seed fate by studying the effects of dung type and defecation pattern on secondary seed dispersal by dung beetles. First, we described the defecation patterns for both primate species. Howler monkeys generally defecated in groups (88% of observed defecations), with each individual producing on average 31 g of dung, resulting in a large area of the forest floor (31 m²) covered by large amounts of dung (clumped spatial pattern). Spider monkeys generally (96% of observed defecations) defecated individually, each individual producing an average of 11 g of dung, resulting in a small area of the forest floor (2 m²) covered by small amounts of dung (scattered spatial pattern). Secondly, we captured dung beetles using as bait the dung of both primate species, to detect differences in the assemblages of these secondary seed dispersers attracted to the dung of both primates. More individual dung beetles, but not more species, were attracted to howler monkey dung than to spider monkey dung. Finally, we assessed experimentally (using plastic beads as seed mimics) how dung type (Ateles vs. Alouatta) and defecation pattern (scattered vs. clumped) affect secondary seed dispersal by dung beetles. We found that post‐dispersal seed fate was affected by dung type, with more seeds being buried when present in howler monkey dung, than in spider monkey dung, but was not affected by defecation pattern. It is important to consider post‐dispersal processes, such as secondary seed dispersal by dung beetles, when comparing species of primary dispersers.     

Differentiation-associated modulation of heparan sulfate structure and function in CaCo-2 colon carcinoma cells

Heparan sulfate species expressed by different cell and tissue types differ in their structural and functional properties. Limited information is available on differences in regulation of heparan sulfate biosynthesis within a single tissue or cell population under different conditions. We have approached this question by studying the effect of cell differentiation on the biosynthesis and function of heparan sulfate in human colon carcinoma cells (CaCo-2). These cells undergo spontaneous differentiation in culture when grown on semipermeable supports; the differentiated cells show phenotypic similarity to small intestine enterocytes. Metabolically labeled heparan sulfate was isolated from the apical and basolateral media from cultures of differentiated and undifferentiated cells. Compositional analysis of disaccharides, derived from the contiguous N-sulfated regions of heparan sulfate, indicated a greater proportion of 2-O- sulfated iduronic acid units and a smaller amount of 6-O-sulfated glucosamine units in differentiated than in undifferentiated cells. By contrast, the overall degree of sulfation, the chain length and the size distribution of the N-acetylated regions were similar regardless the differentiation status of the cells. The structural changes were found to affect the binding of heparan sulfate to the long isoform of platelet-derived growth factor A chain but not to fibroblast growth factor 2. These findings show that heparan sulfate structures change during cell differentiation and that heparan sulfate-growth factor interactions may be affected by such changes.      

Short tetracysteine tags to beta-tubulin demonstrate the significance of small labels for live cell imaging

Genetically encoded tags are of fundamental importance for live cell imaging. We show that small tetracysteine (TetCys) tags can be highly advantageous for the functionality of the host protein compared with large fluorescent protein tags. One to three concatenated small TetCys tags as well as the large green fluorescent protein (GFP) were fused by integrative epitope tagging to the C terminus of beta-tubulin (Tub2) in the budding yeast Saccharomyces cerevisiae. The increasing tag size correlated with functional interference to the host protein. Tub2 tagged with either 1 x TetCys (10 amino acids [aa]) or 2 x TetCys (20 aa) was able to substitute Tub2 in haploid cells. In contrast, C-terminal tagging of Tub2 with 3 x TetCys (29 aa) or with GFP (244 aa) resulted in nonviable haploid cells. Cells expressing Tub2-1 x TetCys or Tub2-2 x TetCys were stained with FlAsH, which selectively binds to the TetCys-tag. The stained cells displayed dynamic FlAsH-labeled microtubules and low cellular background fluorescence. The presented approach to tag open reading frames (ORFs) at their native loci with very small TetCys-tags and the subsequent visualization of the tagged proteins in vivo can be extended in principle to any ORF in S. cerevisiae.     

Synthesis and membrane behavior of a new class of unnatural phospholipid analogs useful as phospholipase A2 degradable liposomal drug carriers

A new and unnatural type of lipid analogs with the phosphocholine and phosphoglycerol head groups linked to the C-2 position of the glycerol moiety have been synthesized and the thermodynamic lipid membrane behavior has been investigated using differential scanning calorimetry. From the heat capacity measurements, it was observed that the pre-transition was abolished most likely due to the central position of the head groups providing better packing properties in the low temperature ordered gel phase. Activity measurements of secretory phospholipase A2 (PLA2) on unilamellar liposomal membranes revealed that the unnatural phospholipids are excellent substrates for PLA2 catalyzed hydrolysis. This was manifested as a minimum in the PLA2 lag time in the main phase transition temperature regime and a high degree of lipid hydrolysis over a broad temperature range. The obtained results provide new information about the interplay between the molecular structure of phospholipids and the lipid membrane packing constrains that govern the pre-transition. In addition, the PLA2 activity measurements are useful for obtaining deeper insight into the molecular details of the catalytic site of PLA2. The combined results also suggest new approaches to rationally design liposomal drug carries that can undergo a triggered activation in diseased tissue by overexpressed PLA2.     

Clinical results of TRAM flap delay by selective embolization of the deep inferior epigastric arteries

Preoperative selective embolization of the deep inferior epigastric arteries constitutes a new technique in TRAM flap delay. Whereas surgical ligation of these vessels has proved to be an effective delay procedure in experimental and clinical settings, it requires an additional operative step under general anesthesia. Despite the introduction of the free TRAM leading to improved flap perfusion, this microsurgical technique is not always available because of the requirements of specialized equipment and staff, longer operating hours, and subsequently higher expenses. The search for a minimally invasive, easy, and inexpensive technique to improve perfusion of the pedicled TRAM flap led us to selective embolization of the deep inferior epigastric arteries by an angiographic procedure. After 4 years of experience with this technique, we now present the first clinical results. Breast reconstruction by a delayed pedicled TRAM flap was performed in 40 patients with a mean age of 48.4 years (range, 31 to 66 years). The mean interval between embolization and surgery was 3.6 months. Postoperative data concerning flap survival and complications were available for all patients. Embolization of the deep inferior epigastric arteries was performed bilaterally in 35 patients (87.5 percent) and unilaterally in 5 patients (12.5 percent). Radiotherapy had been applied in 21 patients (52.5 percent) before surgery. Postoperative flap complications consisted of partial necrosis in three (7.5 percent), fat necrosis in one (2.5 percent), impaired wound healing in five (12.5 percent), and postoperative bleeding in two patients (5 percent). Abdominal wound healing complications occurred in six patients (15 percent), abdominal wall weakness in eight (20 percent), and hernia formation in four (10 percent). Surgical corrections were performed at the breast (TRAM flap) in 22 patients (55 percent) and at the abdomen (donor site) in 9 (22.5 percent). Preoperative selective embolization of the deep inferior epigastric arteries constitutes an alternative delay procedure for the pedicled TRAM flap. It is superior to the conventional procedure without delay, offers several advantages compared with surgical ligation of these vessels, and represents an alternative to the free TRAM flap in selected cases.     

CCK inhibits the orexigenic effect of peripheral ghrelin

CCK and ghrelin exert antagonistic effects on ingestive behavior. The aim of the present study was to investigate the interaction between ghrelin and CCK administered peripherally on food intake and neuronal activity in specific hypothalamic and brain stem nuclei, as assessed by c-Fos-like immunoreactivity (c-FLI) in nonfasted rats. Ghrelin (13 microg/kg body wt) injected intraperitoneally significantly increased the cumulative food intake when measured at 30 min and 1 h after injection, compared with the vehicle group (2.9 +/- 1.0 g/kg body wt vs. 1.2 +/- 0.5 g/kg body wt, P < 0.028). Sulfated CCK octapeptide (CCK-8S) (2 or 25 microg/kg body wt) injected simultaneously blocked the orexigenic effect of ghrelin (0.22 +/- 0.13 g/kg body wt, P < 0.001 and 0.33 +/- 0.23 g/kg body wt, P < 0.0008), while injected alone, both doses of CCK-8S exerted a nonsignificant trend to reduce food intake. Ghrelin (13 microg/kg body wt ip) markedly increased the number of c-FLI-positive neurons per section in the arcuate nucleus (ARC) compared with vehicle (median: 31.35 vs. 9.86, P < 0.0001). CCK-8S (2 or 25 microg/kg body wt ip) had no effect on neuronal activity in the ARC, as assessed by c-FLI (median: 5.33 and 11.21 cells per section), but blocked the ghrelin-induced increase of c-fos expression in this area when both peptides were administered simultaneously (median: 13.33 and 12.86 cells per section, respectively). Ghrelin at this dose had no effect on CCK-induced stimulation of c-fos expression in the paraventricular nucleus of the hypothalamus and the nucleus of the solitary tract. These results suggest that CCK abolishes ghrelin-induced food intake through dampening increased ARC neuronal activity.     

The effects of granulocyte colony-stimulating factor and neutrophil recruitment on the pulmonary chemokine response to intratracheal endotoxin

Although G-CSF has been shown to increase neutrophil (polymorphonuclear leukocyte, PMN) recruitment into the lung during pulmonary infection, relatively little is known about the local chemokine profiles associated with this enhanced PMN delivery. We investigated the effects of G-CSF and PMN recruitment on the pulmonary chemokine response to intratracheal LPS. Rats pretreated twice daily for 2 days with an s.c. injection of G-CSF (50 microg/kg) were sacrificed at either 90 min or 4 h after intratracheal LPS (100 microg) challenge. Pulmonary recruitment of PMNs was not observed at 90 min post LPS challenge. Macrophage inflammatory protein-2 (MIP-2) and cytokine-induced neutrophil chemoattractant (CINC) concentrations in bronchoalveolar lavage (BAL) fluid were similar in animals pretreated with or without G-CSF at this time. G-CSF pretreatment enhanced pulmonary recruitment of PMNs (5-fold) and greatly reduced MIP-2 and CINC levels in BAL fluid at 4 h after LPS challenge. In vitro, the presence of MIP-2 and CINC after LPS stimulation of alveolar macrophages was decreased by coculturing with circulating PMNs but not G-CSF. G-CSF had no direct effect on LPS-induced MIP-2 and CINC mRNA expression by alveolar macrophages. Pulmonary recruited PMNs showed a significant increase in cell-associated MIP-2 and CINC. Cell-associated MIP-2 and CINC of circulating PMNs were markedly increased after exposure of these cells to the BAL fluid of LPS-challenged lungs. These data suggest that recruited PMNs are important cells in modulating the local chemokine response. G-CSF augments PMN recruitment and, thereby, lowers local chemokine levels, which may be one mechanism resulting in the subsidence of the host proinflammatory response.     

Human and mouse mitochondrial orthologs of bacterial ClpX

We have determined the cDNA sequence and exon/intron structure of the human CLPX gene encoding a human ortholog of the E. coli ClpX chaperone and protease subunit. The CLPX gene comprises 14 exons and encodes a 633-amino acid-long precursor polypeptide. The polypeptide contains an N-terminal putative mitochondrial transit peptide, and expression of a full-length ClpX cDNA tagged at its C-terminus (Myc-His) shows that the polypeptide is transported into mitochondria. FISH analysis localized the CLPX gene to human Chromosome (Chr) 15q22.1-22.32. This localization was refined by radiation hybrid mapping placing the CLPX gene 4.6 cR distal to D15S159. Murine ClpX cDNA was sequenced, and the mouse Clpx locus was mapped to a position between 31 and 42 cM offset from the centromere on mouse Chr 9. Experimental observations indicate the presence of a pseudogene in the mouse genome and sequence variability between mouse ClpX cDNAs from different strains. Alignment of the human and mouse ClpX amino acid sequences with ClpX sequences from other organisms shows that they display the typical modular organization of domains with one AAA⁺ domain common to a large group of ATPases and several other domains conserved in ClpX orthologs linked by non-conserved sequences. Notably, a C-4 zinc finger type motif is recognized in human and mouse ClpX. This motif of so far unknown function is present only in a subset of the known ClpX sequences. Received: 5 April 2000 / Accepted: 14 June 2000     

Characterization of mouse Clpp protease cDNA, gene, and protein

Mutations that cause accumulation or rapid degradation owing to protein misfolding are a frequent cause of inherited disease in humans. In Escherichia coli, Clpp protease is one of the components of the protein quality control system that handles misfolded proteins. In the present study, we have characterized the mouse Clpp cDNA sequence, the organization of the mouse gene, the chromosomal localization, and the tissue-specific expression pattern. Moreover, the cellular localization and processing of mouse Clpp was studied by overexpression in transfected eukaryotic cells. Our results indicate that mouse and human Clpp have similar roles, and they provide the molecular basis for establishing a Clpp knockout mouse and to study its phenotype, thereby shedding light on a possible role of Clpp in human disease. Received: 25 June 1999 / Accepted: 9 December 1999