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Abstract Background Pseudomonas syringae pv. phaseolicola, the causal agent of halo blight disease in beans, produces a toxin known as phaseolotoxin, in whose synthesis participate a group of genes organized within the genome in a region known as the "Pht cluster". This region, which is thought to have been acquired by horizontal gene transfer, includes 5 transcriptional units, two monocistronic (argK, phtL) and three polycistronic (phtA, phtD, phtM), whose expression is temperature dependent. So far, the regulatory mechanisms involved in phaseolotoxin synthesis have not been elucidated and the only well-established fact is the requirement of low temperatures for its synthesis. In this work, we searched for regulatory proteins that could be involved in phaseolotoxin synthesis, focusing on the regulation of the phtD operon. Results In this study we identified the global regulator IHF (Integration Host Factor), which binds to the promoter region of the phtD operon, exerting a negative effect on the expression of this operon. This is the first regulatory protein identified as part of the phaseolotoxin synthesis system. Our findings suggest that the Pht cluster was similarly regulated in the ancestral cluster by IHF or similar protein, and integrated into the global regulatory mechanism of P. syringae pv. phaseolicola, after the horizontal gene transfer event by using the host IHF protein. Conclusion This study identifies the IHF protein as one element involved in the regulation of phaseolotoxin synthesis in P. syringae pv. phaseolicola NPS3121 and provides new insights into the regulatory mechanisms involved in phaseolotoxin production.

Background: Pseudomonas syringae pv. phaseolicola, the causal agent of halo blight disease in beans, produces a toxin known as phaseolotoxin, in whose synthesis participate a group of genes organized within the genome in a region known as the "Pht cluster". This region, which is thought to have been acquired by horizontal gene transfer, includes 5 transcriptional units, two monocistronic (argK, phtL) and three polycistronic (phtA, phtD, phtM), whose expression is temperature dependent. So far, the regulatory mechanisms involved in phaseolotoxin synthesis have not been elucidated and the only well-established fact is the requirement of low temperatures for its synthesis. In this work, we searched for regulatory proteins that could be involved in phaseolotoxin synthesis, focusing on the regulation of the phtD operon. Results: In this study we identified the global regulator IHF (Integration Host Factor), which binds to the promoter region of the phtD operon, exerting a negative effect on the expression of this operon. This is the first regulatory protein identified as part of the phaseolotoxin synthesis system. Our findings suggest that the Pht cluster was similarly regulated in the ancestral cluster by IHF or similar protein, and integrated into the global regulatory mechanism of P. syringae pv. phaseolicola, after the horizontal gene transfer event by using the host IHF protein. Conclusion: This study identifies the IHF protein as one element involved in the regulation of phaseolotoxin synthesis in P. syringae pv. phaseolicola NPS3121 and provides new insights into the regulatory mechanisms involved in phaseolotoxin production. [ABSTRACT FROM AUTHOR]

Entamoeba histolytica is a unicellular eukaryotic organism that was located in theaamitochondriate branch. However, recent experiments reveal that presence of organelles and mitochondrial proteins in E. histoytica. Several protozoa parasites like Trypanosome cruzy and Plasmodium falciparium contain family A DNA polymerases that replicate their organelles. We have performed a "in silico" search for Family A DNA polymerases in the genome data base of E. histolytica using as bait the arquetype of family A DNA polymerases: Klenow Fragment. We identified an open reading frame that corresponds to a protein of 67 kDa with high homology to Klenow Fragment, bacteriophage T7 DNA polymerase and mitochondrial polymerases. We amplified this ORF using PCR reaction and subcloned it in a pCOLD vector. The expression of the recombinant polymerase was carried out at 16 C and its purification was facilitated by the addition of N-terminal histidine tag. The recombinant polymerase is active in primer extension experiments and displays very low processivity. We are in the process to obtain antibodies against this polymerase to observe its cellular localization. We hypothesize that this DNA polymerase may target to the organelles present in E. histolytica. This would be evidence that E. histolytica may have a primitive mitochondrial-like organelle. [ABSTRACT FROM AUTHOR]