Cloning and gene map assignment of the Xiphophorus DNA ligase 1 gene

Fishes represent the stem vertebrate condition and have maintained several gene arrangements common to mammalian genomes throughout the 450 Myr of divergence from a common ancestor. One such syntenic arrangement includes the GPI-PEPD enzyme association on Xiphophorus linkage group IV and human chromosome 19. Previously we assigned the Xiphophorus homologue of the human ERCC2 gene to linkage group U5 in tight association with the CKM locus. CKM is also tightly linked to the ERCC2 locus on human chromosome 19, leading to speculation that human chromosome 19 may have arisen by fusion of two ancestral linkage groups which have been maintained in fishes. To investigate this hypothesis further, we isolated and sequenced Xiphophorus fish genomic regions exhibiting considerable sequence similarity to the human DNA ligase 1 amino acid sequence. Comparison of the fish DNA ligase sequence with those of other species suggests several modes of amino acid conservation in this gene. A 2.2-kb restriction fragment containing part of an X. maculatus DNA ligase 1 exon was used in backcross hybrid mapping with 12 enzyme or RFLP loci. Significant linkage was observed between the nucleoside phosphorylase (NP2) and the DNA ligase (LIG1) loci on Xiphophorus linkage group VI. This assignment suggests that the association of four DNA repair-related genes on human chromosome 19 may be the result of chance chromosomal rearrangements.      

Marine Heterotrophic Amoebae, Flagellates and Heliozoa From Belize (Central America) and Tenerife (Canary Islands), With Descriptions of New Species, Luffisphaera Bulbochaete N. Sp., L. Longihastis N. Sp., L. Turriformis N. Sp. and Paulinella Intermedia N. Sp.

ABSTRACT. Thirty four taxa of heterotrophic protists (amoebae, flagellates and heliozoa) were encountered in cultures established from marine samples from Belize (Central America) and Tenerife (Canary Islands). Most species are flagellates drawn from the choanoflagellates, the cryptophyceans, the euglenids, the kinetoplastids, the bicosoecids, the chromulinids, the pedinellids and a variety of laxa of uncertain affinities (Protista incertae sedis). the identity of the thecate choanoflagellates Salpingoeca ringens Kent, 1880, and S. tuba Kent, 1880, is discussed, and four new species of heterotrophic protists are described: one new species of the amoeba genus Paulinella (Paulinella intermedia n. sp.) and three new species of the incertae sedis genus Luffisphaera Belcher & Swale, 1975 ( Luffisphaera bulbochaete n. sp.; L. longihastis n. sp.; L. turriformis n. sp.).     

Sulfation of fucoidin in focus embryos: III. Required for localization in the rhizoid wall

Zygotes of the brown alga Fucus distichus L. Powell accumulate a sulfated polysaccharide (fucoidin) in the cell wall at the site of rhizoid formation. Previous work indicated that zygotes grown in seawater minus sulfate do not sulfate the preformed fucan (an unsulfated fucoidin) but form rhizoids. Under these conditions, we determined whether sulfation of the fucan is required for its localization in the rhizoid wall. This was accomplished by developing a specific stain for both the fucan and fucoidin. Using a precipitin assay, we demonstrated in vitro that the lectin ricin (RCA(I)) specifically complexes with both the sulfated and desulfated polysaccharide. No precipitate is observed when either is incubated in 0.1 M D-galactose or when RCA(I) is mixed with laminarin or alginic acid, the other major polysaccharides in Fucus. RCA(I) conjugated with fluorescein isothiocyanate (FITC) is also shown to bind specifically to fucoidin using a filter paper (DE81) assay. When added to zygotes, RCA(I)-FITC binds only to the site of fucoidin localization, i.e., the rhizoid cell wall. However, RCA(I)-FITC is not observed in the rhizoid wall of zygotes grown in the absence of sulfate. This observation is not due to inability of RCA(I)-FITC to bind to the fucan in vivo. Chemically desulfated cell walls that contained fucoidin in the rhizoid wall bind RCA(I)-FITC only in the rhizoid region. Also, the concentration of fucose-containing polymers and polysaccharides that form precipitates with RCA(I) is the same in embryos grown in the presence or absence of sulfate. If sulfate is added back to cultures of zygotes grown without sulfate, fucoidin is detected at the rhizoid tip by RCA(I)-FITC several hours later. These results support the conclusion that the enzymatic sulfation of the fucan is a modification of the polysaccharide required for its localization and/or assembly into a specific region of the cell wall.     

Divergent calcium signaling in RBCs from <Emphasis Type="Italic">Tropidurus torquatus</Emphasis> (Squamata – Tropiduridae) strengthen classification in lizard evolution

Background  

We have previously reported that a Teiid lizard red blood cells (RBCs) such as Ameiva ameiva and Tupinambis merianae controls intracellular calcium levels by displaying multiple mechanisms. In these cells, calcium stores could be discharged not only by: thapsigargin, but also by the Na⁺/H⁺ ionophore monensin, K⁺/H⁺ ionophore nigericin and the H⁺ pump inhibitor bafilomycin as well as ionomycin. Moreover, these lizards possess a P2Y-type purinoceptors that mobilize Ca²⁺ from intracellular stores upon ATP addition.     

Botryoid odontogenic cyst. Exploration of proliferative activity,apoptosis and expression of TP53 and BCL2 compared to the histologically identical lateral periodontal and gingival cysts

The botryoid odontogenic cyst (BOC) is a rare, locally more aggressive variant of the usually indolent lateral periodontal cyst (LPC) and gingival cyst (GC). A recent case of BOC provided an opportunity for an exploratory study on the causes of its more aggressive behavior. The limited objective was to see if the BOC was sufficiently different from the other cysts to warrant an investment in a large study. Sections of neutral buffered formalin fixed, paraffin-embedded tissues from the BOC and archival specimens of four GCs, four LPCs and three odontogenic keratocysts (OKCs) were stained using immunohistochemistry for Ki-67, a marker of proliferating cells, caspase-3, a marker of cells undergoing apoptosis, tumor suppressor p53, and the apoptosis inhibitor BCL2. The mean labeling index (LI) of immunoreactive cyst epithelial cells was computed for each antibody and type of cyst. Compared to the LPCs and GCs, the BOC exhibited a moderately larger Ki-67/caspase-3 LI difference, which indicates that the BOC had a net higher rate of growth. We found a much higher level of LI, therefore likely dysregulation of p53. We also found a much higher LI of BCL2. The LIs of p53 and BCL2 in the BOC were similar to and more than twice that of the OKCs, respectively. Although meaningful statistical analysis was precluded by our use of only one case of BOC and a small number of the other cysts, the high p53 and very high BCL2 labeling indices of the BOC offer a potential explanation for its reportedly more aggressive behavior that clearly is worthy of further investigation.     

Effects of high arsenic and fluoride soil concentrations on soybean plants

Arsenic (As) and Fluoride (F) are present in many soils, affecting crops and posing risks in the food chain. We performed pot experiments on spiked soils enriched in these elements either individually or simultaneously, over a wide range of concentrations. Soybean biomass production, grain yield, As and F accumulation and distribution within the plant, and the antioxidant response to these stresses were analyzed. Arsenic was more toxic than F. At As levels >35 mg/kg and F levels >375 mg/kg, yield loss reached 60% and 30%, respectively. At the highest dose of As plants died within 2 weeks, whereas F showed no lethality. When they were applied simultaneously, detrimental effects were more important. As and F in plants increased in all soybean organs although grains presented the lowest concentrations. Antioxidant enzymes were enhanced in plants but this increase was not high enough to cope with the oxidative damage.     

PKA and Epac cooperate to augment bradykinin-induced interleukin-8 release from human airway smooth muscle cells

Background

Platelet-derived growth factor A (PDGF-A) signals solely through PDGF-Rα, and is required for fibroblast proliferation and transdifferentiation (fibroblast to myofibroblast conversion) during alveolar development, because pdgfa-null mice lack both myofibroblasts and alveoli. However, these PDGF-A-mediated mechanisms remain incompletely defined. At postnatal days 4 and 12 (P4 and P12), using mouse lung fibroblasts, we examined (a) how PDGF-Rα correlates with ki67 (proliferation marker) or alpha-smooth muscle actin (αSMA, myofibroblast marker) expression, and (b) whether PDGF-A directly affects αSMA or modifies stimulation by transforming growth factor beta (TGFβ).

Methods

Using flow cytometry we examined PDGF-Rα, αSMA and Ki67 in mice which express green fluorescent protein (GFP) as a marker for PDGF-Rα expression. Using real-time RT-PCR we quantified αSMA mRNA in cultured Mlg neonatal mouse lung fibroblasts after treatment with PDGF-A, and/or TGFβ.

Results

The intensity of GFP-fluorescence enabled us to distinguish three groups of fibroblasts which exhibited absent, lower, or higher levels of PDGF-Rα. At P4, more of the higher than lower PDGF-Rα + fibroblasts contained Ki67 (Ki67+), and Ki67+ fibroblasts predominated in the αSMA + but not the αSMA- population. By P12, Ki67+ fibroblasts comprised a minority in both the PDGF-Rα + and αSMA+ populations. At P4, most Ki67+ fibroblasts were PDGF-Rα + and αSMA- whereas at P12, most Ki67+ fibroblasts were PDGF-Rα- and αSMA-. More of the PDGF-Rα + than - fibroblasts contained αSMA at both P4 and P12. In the lung, proximate αSMA was more abundant around nuclei in cells expressing high than low levels of PDGF-Rα at both P4 and P12. Nuclear SMAD 2/3 declined from P4 to P12 in PDGF-Rα-, but not in PDGF-Rα + cells. In Mlg fibroblasts, αSMA mRNA increased after exposure to TGFβ, but declined after treatment with PDGF-A.

Conclusion

During both septal eruption (P4) and elongation (P12), alveolar PDGF-Rα may enhance the propensity of fibroblasts to transdifferentiate rather than directly stimulate αSMA, which preferentially localizes to non-proliferating fibroblasts. In accordance, PDGF-Rα more dominantly influences fibroblast proliferation at P4 than at P12. In the lung, TGFβ may overshadow the antagonistic effects of PDGF-A/PDGF-Rα signaling, enhancing αSMA-abundance in PDGF-Rα-expressing fibroblasts.     

Lateralization of hand skill in bipolar affective disorder

Diverse strands of evidence suggest that schizophrenia is associated with an excess of left and mixed handedness, reflecting anomalous cerebral lateralization. Genetic studies have indicated a degree of overlap between bipolar disorder (BPD) and schizophrenia. Nevertheless, pattern of handedness and degree of lateralization have not been explicitly tested in BPD. We measured handedness, footedness and relative manual dexterity in a sample of 47 families comprising BPD probands and their bipolar-spectrum and unaffected relatives (N = 240). The BPD I sample (N = 55) was significantly more lateralized on handedness, footedness and relative manual dexterity than their unaffected relatives (N = 66). They were also more lateralized than their relatives with other psychiatric diagnoses. No evidence of excess mixed handedness or footedness was observed in the BPD I sample. We raise the possibility that schizophrenia and BPD I differ in that disproportionate left-hemisphere dominance in BPD I is associated with right-hemisphere dysfunction leading to deficits in emotional regulation. Given our results, we hypothesized that degree of lateralization may be a phenotypic marker or endophenotype for BPD I. We therefore conducted a family-based genetic association analysis with this quantitative trait. Relative hand skill was significantly associated with a functional variant in the catechol-O-methyltransferase gene. We speculate that this polymorphism may influence brain lateralization.     

A Genomic Portrait of Haplotype Diversity and Signatures of Selection in Indigenous Southern African Populations

We report a study of genome-wide, dense SNP (∼900K) and copy number polymorphism data of indigenous southern Africans. We demonstrate the genetic contribution to southern and eastern African populations, which involved admixture between indigenous San, Niger-Congo-speaking and populations of Eurasian ancestry. This finding illustrates the need to account for stratification in genome-wide association studies, and that admixture mapping would likely be a successful approach in these populations. We developed a strategy to detect the signature of selection prior to and following putative admixture events. Several genomic regions show an unusual excess of Niger-Kordofanian, and unusual deficiency of both San and Eurasian ancestry, which were considered the footprints of selection after population admixture. Several SNPs with strong allele frequency differences were observed predominantly between the admixed indigenous southern African populations, and their ancestral Eurasian populations. Interestingly, many candidate genes, which were identified within the genomic regions showing signals for selection, were associated with southern African-specific high-risk, mostly communicable diseases, such as malaria, influenza, tuberculosis, and human immunodeficiency virus/AIDs. This observation suggests a potentially important role that these genes might have played in adapting to the environment. Additionally, our analyses of haplotype structure, linkage disequilibrium, recombination, copy number variation and genome-wide admixture highlight, and support the unique position of San relative to both African and non-African populations. This study contributes to a better understanding of population ancestry and selection in south-eastern African populations; and the data and results obtained will support research into the genetic contributions to infectious as well as non-communicable diseases in the region.