Multi-tissue microarray analysis identifies a molecular signature of regeneration.

The inability to functionally repair tissues that are lost as a consequence of disease or injury remains a significant challenge for regenerative medicine. The molecular and cellular processes involved in complete restoration of tissue architecture and function are expected to be complex and remain largely unknown. Unlike humans, certain salamanders can completely regenerate injured tissues and lost appendages without scar formation. A parsimonious hypothesis would predict that all of these regenerative activities are regulated, at least in part, by a common set of genes. To test this hypothesis and identify genes that might control conserved regenerative processes, we performed a comprehensive microarray analysis of the early regenerative response in five regeneration-competent tissues from the newt Notophthalmus viridescens. Consistent with this hypothesis, we established a molecular signature for regeneration that consists of common genes or gene family members that exhibit dynamic differential regulation during regeneration in multiple tissue types. These genes include members of the matrix metalloproteinase family and its regulators, extracellular matrix components, genes involved in controlling cytoskeleton dynamics, and a variety of immune response factors. Gene Ontology term enrichment analysis validated and supported their functional activities in conserved regenerative processes. Surprisingly, dendrogram clustering and RadViz classification also revealed that each regenerative tissue had its own unique temporal expression profile, pointing to an inherent tissue-specific regenerative gene program. These new findings demand a reconsideration of how we conceptualize regenerative processes and how we devise new strategies for regenerative medicine.

Methanosarcina acetivorans flap endonuclease 1 activity is inhibited by a cognate single-stranded-DNA-binding protein.

The oligonucleotide/oligosaccharide-binding (OB) fold is central to the architecture of single-stranded- DNA-binding proteins, which are polypeptides essential for diverse cellular processes, including DNA replication, repair, and recombination. In archaea, single-stranded DNA-binding proteins composed of multiple OB folds and a zinc finger domain, in a single polypeptide, have been described. The OB folds of these proteins were more similar to their eukaryotic counterparts than to their bacterial ones. Thus, the archaeal protein is called replication protein A (RPA), as in eukaryotes. Unlike most organisms, Methanosarcina acetivorans harbors multiple functional RPA proteins, and it was our interest to determine whether the different proteins play different roles in DNA transactions. Of particular interest was lagging-strand DNA synthesis, where recently RPA has been shown to regulate the size of the 5' region cleaved during Okazaki fragment processing. We report here that M. acetivorans RPA1 (MacRPA1), a protein composed of four OB folds in a single polypeptide, inhibits cleavage of a long flap (20 nucleotides) by M. acetivorans flap endonuclease 1 (MacFEN1). To gain a further insight into the requirement of the different regions of MacRPA1 on its inhibition of MacFEN1 endonuclease activity, N-terminal and C-terminal truncated derivatives of the protein were made and were biochemically and biophysically analyzed. Our results suggested that MacRPA1 derivatives with at least three OB folds maintained the properties required for inhibition of MacFEN1 endonuclease activity. Despite these interesting observations, further biochemical and genetic analyses are required to gain a deeper understanding of the physiological implications of our findings.

The euryarchaeota, nature's medium for engineering of single-stranded DNA-binding proteins.

The architecture of single-stranded DNA-binding proteins, which play key roles in DNA metabolism, is based on different combinations of the oligonucleotide/oligosaccharide binding (OB) fold. Whereas the polypeptide serving this function in bacteria contains one OB fold, the eukaryotic functional homolog comprises a complex of three proteins, each harboring at least one OB fold. Here we show that unlike these groups of organisms, the Euryarchaeota has exploited the potential in the OB fold to re-invent single-stranded DNA-binding proteins many times. However, the most common form is a protein with two OB folds and one zinc finger domain. We created several deletion mutants of this protein based on its conserved motifs, and from these structures functional chimeras were synthesized, supporting the hypothesis that gene duplication and recombination could lead to novel functional forms of single-stranded DNA-binding proteins. Biophysical studies showed that the orthologs of the two OB fold/one zinc finger replication protein A in Methanosarcina acetivorans and Methanopyrus kandleri exhibit two binding modes, wrapping and stretching of DNA. However, the ortholog in Ferroplasma acidarmanus possessed only the stretching mode. Most interestingly, a second single-stranded DNA-binding protein, FacRPA2, in this archaeon exhibited the wrapping mode. Domain analysis of this protein, which contains a single OB fold, showed that its architecture is similar to the functional homologs thought to be unique to the Crenarchaeotes. Most unexpectedly, genes coding for similar proteins were found in the genomes of eukaryotes, including humans. Although the diversity shown by archaeal single-stranded DNA-binding proteins is unparalleled, the presence of their simplest form in many organisms across all domains of life is of greater evolutionary consequence.

Diferenciación de Entamoeba histolytica y Entamoeba dispar en muestras de materia fecal por detección de adhesina de E. histolytica mediante ELISA / Entamoeba histolytica and Entamoeba dispar differentiation in faeces samples by E. histolytica adhesin detection through ELISA

La infecciones causadas por el complejo Entamoeba histolytica/Entamoeba dispar son de distribución cosmopolita. E. histolytica es la única especie patógena, pero la diferenciación con E. dispar se hace por métodos bioquímicos. Este trabajo tuvo como finalidad detectar rápidamente la adhesina específica de E. histolytica para diferenciarla de E. dispar en materia fecal, así como determinar la frecuencia del complejo E. histolytica/E. dispar en una población rural de Cundinamarca. En el puesto de salud de La Virgen, Quipile, Cundinamarca, se recolectaron 140 muestras de materia fecal de niños y adultos y se analizaron por el método de concentración (formol-éter) para determinar la frecuencia del complejo E. histolytica/E. dispar. Las 23 muestras positivas para este complejo (16,42 por ciento) se valoraron con la metodología de la prueba de ELISA para la detección de adhesina; se obtuvo una frecuenca de 8,69 por ciento (2/23) de E. histoytica. A 19 pacientes positivos para el complejo E. histolytica/E. dispar se les realizó ELISA para la detección de anticuerpos en sangre contra E. histolytica con resultado negativo. Extrapolando los resultados de la detección de adhesina a la población total de este estudio (2/140), se obtuvo una frecuencia de 1,42 por ciento de E. histolytica. La ventaja de la prueba de ELISA para la detección de adhesina es su fácil ejecución que permite hacer un diagnóstico rápido para instaurar un manejo adecuado. Se recomienda la realizació de estudios de prevalencia a nivel nacional