Restriction analysis of spacers in ribosomal DNA of Drosophila melanogaster.

The sequence arrangement of ribosomal DNA (rDNA) spacers in Drosophila melanogaster was analyzed with restriction endonucleases. Spacers, derived from cloned rDNA repeats and from uncloned purified rDNA, are internally repetitive, as demonstrated by the regular 250 base pairs interval between sites recognized by the enzyme Alu I. Length heterogeneity of spacers is due at least in part to varying numbers of repeated sequence elements. All spacers and analyzed, whether derived from X or from Y chromosomal rDNA, have a very similar sequence organization. The distance separating the repeated nontranscribed spacer sequences from the 5' end of the transcribed region is conserved in all ten cloned fragments examined, and is probably less than 150 base pairs, as measured by electron microscopy.     

A reinvestigation of 5′→3′ polarity in 40S ribosomal RNA precursor of xenopus laevis

The 5′→3′ polarity of the 40S precursor rRNA molecule relative to the location of the 18S and 28S RNA regions in the precursor has been reinvestigated. Fragments of rDNA derived by the restriction endonuclease EcoRI and cloned in E. coli were partially digested with the exonuclease induced by bacteriophage λ and with exonuclease III from E. coli. The resulting rDNA fragments with single-stranded tails were hybridized separately with 18S and 28S rRNA, and the formation of the hybrid was monitored by determination of radioactivity and by electron microscopy. Since the location of the EcoRI sites in rDNA is known, and the specificity of the two exonucleases for 5′ and 3′ ends of DNA strands has been established, the hybridization of the different partially digested rDNA fragments with either 18S or 28S rRNA could be interpreted in terms of polarity of the coding strand of rDNA, and consequently of the RNA (see models in Figure 1). The results support the following model for the rRNA precursor molecule: 5′ end-transcribed spacer-18S gene-transcribed spacer-28S gene-3′ end.     

Lrig3 regulates neural crest formation in Xenopus by modulating Fgf and Wnt signaling pathways

Leucine-rich repeats and immunoglobulin-like domains 3 (Lrig3) was identified by microarray analysis among genes that show differential expression during gastrulation in Xenopus laevis. Lrig3 was expressed in the neural plate and neural crest (NC) at neurula stages, and in NC derivatives and other dorsal structures during tailbud stages. A prominent consequence of the morpholino-induced inhibition of Lrig3 expression was impaired NC formation, as revealed by the suppression of marker genes, including Slug, Sox9 and Foxd3. In the NC induction assay involving Chordin plus Wnt3a-injected animal caps, Lrig3 morpholino inhibited expression of Slug, Sox9 and Foxd3, but not of Pax3 and Zic1. In line with this, Lrig3 knockdown prevented NC marker induction by Pax3 and Zic1, suggesting that Lrig3 acts downstream of these two genes in NC formation. Injection of Lrig3 and Wnt3a led to low-level induction of NC markers and enhanced induction of Fgf3, Fgf4 and Fgf8 in animal caps, suggesting a positive role for Lrig3 in Wnt signaling. Lrig3 could attenuate Fgf signaling in animal caps, did interact with Fgf receptor 1 in cultured cells and, according to context, decreased or increased the induction of NC markers by Fgf. We suggest that Lrig3 functions in NC formation in Xenopus by modulating the Wnt and Fgf signaling pathways.     

Coordinated Bacteriocin Expression and Competence in Streptococcus pneumoniae Contributes to Genetic Adaptation through Neighbor Predation

Streptococcus pneumoniae (pneumococcus) has remained a persistent cause of invasive and mucosal disease in humans despite the widespread use of antibiotics and vaccines. The resilience of this organism is due to its capacity for adaptation through the uptake and incorporation of new genetic material from the surrounding microbial community. DNA uptake and recombination is controlled by a tightly regulated quorum sensing system that is triggered by the extracellular accumulation of competence stimulating peptide (CSP). In this study, we demonstrate that CSP can stimulate the production of a diverse array of blp bacteriocins. This cross stimulation occurs through increased production and secretion of the bacteriocin pheromone, BlpC, and requires a functional competence regulatory system. We show that a highly conserved motif in the promoter of the operon encoding BlpC and its transporter mediates the upregulation by CSP. The accumulation of BlpC following CSP stimulation results in augmented activation of the entire blp locus. Using biofilm-grown organisms as a model for competition and genetic exchange on the mucosal surface, we demonstrate that DNA exchange is enhanced by bacteriocin secretion suggesting that co-stimulation of bacteriocins with competence provides an adaptive advantage. The blp and com regulatory pathways are believed to have diverged and specialized in a remote ancestor of pneumococcus. Despite this, the two systems have maintained a regulatory connection that promotes competition and adaptation by targeting for lysis a wide array of potential competitors while simultaneously providing the means for incorporation of their DNA.     

Vacuum-assisted core-needle biopsy as a diagnostic and therapeutic method in lesions radiologically suspicious of breast fibroadenoma

Background

Treatment of breast fibroadenoma remains a subject of clinical discussion. Recommended methods include clinical observation or surgical excision of the lesion. The procedure involves hospitalisation and anaesthesia, leaving a scar on the breast.

Aim

The aim of this study was to present the Centre''s experience in removing lesions radiologically suspicious of fibroadenoma by means of an ultrasound-guided vacuum-assisted core-needle biopsy as an alternative to a classical surgery.

Materials and methods

Between March 2007 and April 2010, 196 ultrasound-guided vacuum-assisted biopsies were performed in the Mammotome Biopsy Laboratory of the 1st Surgical Oncology and General Surgery Department at the Greater Poland Cancer Centre in Poznań. The procedure was delivered to female patients aged 17–91 years (mean 40.8, median 39). Qualified for removal were ultrasound identified lesions described as fibroadenomas.

Results

The average size of excised lesions according to pre-biopsy ultrasound image was 13.53 ± 8.92 mm (median 11 mm, range 4–60 mm). In 184 cases (93.9%), benign lesions were found in the final histopathologic examination. Pre-cancer lesions were found in 10 cases, and invasive lesions in two cases. Overall, after follow-up ultrasound examination, four patients were qualified for subsequent surgical resection of lesions that had been left behind.

Conclusion

Vacuum core-needle biopsy is an effective tool enabling removal of breast fibroadenomas. It combines features of a lesion resection and histopathologic material collection providing an access with minimum invasiveness.     

Isolation and characterization of calmodulin genes from Xenopus laevis.

Two cDNAs derived from Xenopus laevis calmodulin mRNA have been cloned. Both cDNAs contain the complete protein-coding region and various lengths of untranslated segments. The two cDNAs encode an identical protein but differ from each other by 5% nucleotide substitutions. The 5' and 3' untranslated regions, to the extent available, are highly homologous between the two cDNAs. The predicted sequence of X. laevis calmodulin is identical to that of vertebrate calmodulins from mammals and chickens and shows one substitution compared with electric eel calmodulin. Genomic DNA sequences homologous to each of the two cDNA clones have been isolated and were shown to account for the major calmodulin-coding DNA sequences in X. laevis. These data suggest that X. laevis carries two active, nonallelic calmodulin genes. Although no complete analysis has been carried out, it appears that the X. laevis calmodulin genes are interrupted by at least four introns. The relative concentrations of calmodulin mRNA have been estimated in different embryonic stages and adult tissues and found to vary by up to a factor of 10. The highest levels of calmodulin mRNA were found in ovaries, testes, and brains. In these three tissues, the two calmodulin genes appear to be expressed at approximately equal levels.     

Isolation and sequence organization of human ribosomal DNA.

The genes coding for 28 S and 18 S ribosomal RNA have been purified from leukemic leukocytes of one human individual by density gradient centrifugation. The purified ribosomal DNA was analyzed by restriction endonuclease digestion and electron microscopy. The location of cleavage sites for the restriction endonuclease EcoRI was established by R-loop mapping of restriction fragments by electron microscopy. The results are in agreement with gel analysis and gel transfer hybridization. One type of ribosomal DNA repeating unit contains four cleavage sites for EcoRI. Two of these cuts are located in the genes coding for 28 S and 18 S rRNA, while the other two are in the non-transcribed spacer. Thus, one of the restriction fragments generated contains non-transcribed spacer sequences only and is not detected by gel transfer hybridization if labeled rRNA is used as the hybridization probe. A second type of repeating unit lacks one of the EcoRI cleavage sites within the non-transcribed spacer. This indicates that sequence heterogeneity exists in human rDNA spacers. R-loop mapping of high molecular weight rDNA in the electron microscope reveals that the majority of repeats are rather uniform in length. The average size of 22 repeats was 43.65(±1.27) kb. Two repeats were found with lengths of 28.6 and 53.9 kb, respectively. This, and additional evidence from gels, indicates that some length heterogeneity does exist in the non-transcribed spacer. The structure of the human rDNA repeat is summarized in Figure 10.     

Drill Holes and Predation Traces versus Abrasion-Induced Artifacts Revealed by Tumbling Experiments

Drill holes made by predators in prey shells are widely considered to be the most unambiguous bodies of evidence of predator-prey interactions in the fossil record. However, recognition of traces of predatory origin from those formed by abiotic factors still waits for a rigorous evaluation as a prerequisite to ascertain predation intensity through geologic time and to test macroevolutionary patterns. New experimental data from tumbling various extant shells demonstrate that abrasion may leave holes strongly resembling the traces produced by drilling predators. They typically represent singular, circular to oval penetrations perpendicular to the shell surface. These data provide an alternative explanation to the drilling predation hypothesis for the origin of holes recorded in fossil shells. Although various non-morphological criteria (evaluation of holes for non-random distribution) and morphometric studies (quantification of the drill hole shape) have been employed to separate biological from abiotic traces, these are probably insufficient to exclude abrasion artifacts, consequently leading to overestimate predation intensity. As a result, from now on, we must adopt more rigorous criteria to appropriately distinguish abrasion artifacts from drill holes, such as microstructural identification of micro-rasping traces.