BUSCADOR RECOLECTA

The biofilm matrix, composed of exopolysaccharides, proteins, nucleic acids and lipids, plays a well-known role as a defence
structure, protecting bacteria from the host immune system and antimicrobial therapy. However, little is known about its
responsibility in the interaction of biofilm cells with host tissues. Staphylococcus aureus, a leading cause of biofilmassociated
chronic infections, is able to develop a biofilm built on a proteinaceous Bap-mediated matrix. Here, we used the
Bap protein as a model to investigate the role that components of the biofilm matrix play in the interaction of S. aureus with
host cells. The results show that Bap promotes the adhesion but prevents the entry of S. aureus into epithelial cells. A broad
analysis of potential interaction partners for Bap using ligand overlayer immunoblotting, immunoprecipitation with purified
Bap and pull down with intact bacteria, identified a direct binding between Bap and Gp96/GRP94/Hsp90 protein. The
interaction of Bap with Gp96 provokes a significant reduction in the capacity of S. aureus to invade epithelial cells by
interfering with the fibronectin binding protein invasion pathway. Consistent with these results, Bap deficient bacteria
displayed an enhanced capacity to invade mammary gland epithelial cells in a lactating mice mastitis model. Our
observations begin to elucidate the mechanisms by which components of the biofilm matrix can facilitate the colonization
of host tissues and the establishment of persistent infections., J.V. and A.T-A were supported by Spanish Ministry of Science and Innovation ‘‘Ramón y Cajal’’ contract. This work was supported by Spanish Ministry of Science and Innovation Grants BIO2008-05284-C02-01, AGL2011-23954 and the grant of Dpto. de Innovación from Gobierno de Navarra IIQ14066.RI1.

UPNa. Instituto de Agrobiotecnología. Laboratorio de Biofilms Microbianos., Incluye 10 ficheros de datos, The presence of regulatory sequences in the 39 untranslated region (39-UTR) of eukaryotic mRNAs controlling RNA stability
and translation efficiency is widely recognized. In contrast, the relevance of 39-UTRs in bacterial mRNA functionality has
been disregarded. Here, we report evidences showing that around one-third of the mapped mRNAs of the major human
pathogen Staphylococcus aureus carry 39-UTRs longer than 100-nt and thus, potential regulatory functions. We selected the
long 39-UTR of icaR, which codes for the repressor of the main exopolysaccharidic compound of the S. aureus biofilm matrix,
to evaluate the role that 39-UTRs may play in controlling mRNA expression. We showed that base pairing between the 39-
UTR and the Shine-Dalgarno (SD) region of icaR mRNA interferes with the translation initiation complex and generates a
double-stranded substrate for RNase III. Deletion or substitution of the motif (UCCCCUG) within icaR 39-UTR was sufficient to
abolish this interaction and resulted in the accumulation of IcaR repressor and inhibition of biofilm development. Our
findings provide a singular example of a new potential post-transcriptional regulatory mechanism to modulate bacterial
gene expression through the interaction of a 39-UTR with the 59-UTR of the same mRNA., This work was supported by grants from Spanish Ministry of Economy and Competitiveness (BFU2011-23222, BIO2008-05284-C02-01 and ERA-NET Pathogenomics PIM2010EPA-00606) and from Fundação para a Ciência e Tecnologia - Portugal (ERA-PTG/0002/2010 and PEst-OE/EQB/LA0004/2011). IRdlM and JV were supported by F.P.I. (BES-2009-017410) and Ramón y Cajal (RYC-2009-03948) contracts, respectively, from the Spanish Ministry of Economy and Competitiveness.

Incluye 1 fichero de datos, The Rcs phosphorelay pathway is a complex signaling pathway involved in the regulation of many cell surface structures in enteric
bacteria. In response to environmental stimuli, the sensor histidine kinase (RcsC) autophosphorylates and then transfers
the phosphate through intermediary steps to the response regulator (RcsB), which, once phosphorylated, regulates gene expression.
Here, we show that Salmonella biofilm development depends on the phosphorylation status of RcsB. Thus, unphosphorylated
RcsB, hitherto assumed to be inactive, is essential to activate the expression of the biofilm matrix compounds. The prevention
of RcsB phosphorylation either by the disruption of the phosphorelay at the RcsC or RcsD level or by the production of a
nonphosphorylatable RcsB allele induces biofilm development. On the contrary, the phosphorylation of RcsB by the constitutive
activation of the Rcs pathway inhibits biofilm development, an effect that can be counteracted by the introduction of a nonphosphorylatable
RcsB allele. The inhibition of biofilm development by phosphorylated RcsB is due to the repression of CsgD expression,
through a mechanism dependent on the accumulation of the small noncoding RNA RprA. Our results indicate that unphosphorylated
RcsB plays an active role for integrating environmental signals and, more broadly, that RcsB phosphorylation
acts as a key switch between planktonic and sessile life-styles in Salmonella enterica serovar Typhimurium., A.T.-A. and J.V. were supported by Ramón y Cajal contracts from the
Ministerio de Ciencia e Innovación, Spain. This research was supported
by grants BIO2008-05284-C02-01 and BIO2010-18885 from the Spanish
Ministerio de Ciencia e Innovación and IIM13329.RI1 from the Gobierno
de Navarra (Spain).

This is an accepted manuscript of an article published by Taylor & Francis in RNA Biology on August 1st 2012, available online: http://dx.doi.org/10.4161/rna.21167., Analysis of bacterial transcriptomes have shown the existence of a genome-wide process of overlapping transcription due to the presence of antisense RNAs, as well as mRNAs that overlapped in their entire length or in some portion of the 5′- and 3′-UTR regions. The biological advantages of such overlapping transcription are unclear but may play important regulatory roles at the level of transcription, RNA stability and translation. In a recent report, the human pathogen Staphylococcus aureus is observed to generate genome-wide overlapping transcription in the same bacterial cells leading to a collection of short RNA fragments generated by the endoribonuclease III, RNase III. This processing appears most prominently in Gram-positive bacteria. The implications of both the use of pervasive overlapping transcription and the processing of these double stranded templates into short RNAs are explored and the consequences discussed., A.T.-A. is recipient of “Ramon y Cajal” contracts from the Spanish Ministry of Science and Innovation. This research was supported by grants ERA-NET Pathogenomics (PIM2010EPA-00606), BIO2008-05284-C02, BIO2011-30503-C02 and BFU2011-23222 from Spanish Ministry of Economy and Competitiveness.

UPNa. Instituto de Agrobiotecnología. Laboratorio de Biofilms Microbianos, Incluye 7 ficheros de datos, Background: High-density oligonucleotide microarray is an appropriate technology for genomic analysis, and is particulary useful in the generation of transcriptional maps, ChIP-on-chip studies and re-sequencing of the genome. Transcriptome analysis of tiling microarray data facilitates the discovery of novel transcripts and the assessment of differential expression in diverse experimental conditions. Although new technologies such as next-generation sequencing have appeared, microarrays might still be useful for the study of small genomes or for the analysis of genomic regions with custom microarrays due to their lower price and good accuracy in expression quantification. Results: Here, we propose a novel wavelet-based method, named ZCL (zero-crossing lines), for the combined denoising and segmentation of tiling signals. The denoising is performed with the classical SUREshrink method and the detection of transcriptionally active regions is based on the computation of the Continuous Wavelet Transform (CWT). In particular, the detection of the transitions is implemented as the thresholding of the zero-crossing lines. The algorithm described has been applied to the public Saccharomyces cerevisiae dataset and it has been compared with two well-known algorithms: pseudo-median sliding window (PMSW) and the structural change model (SCM). As a proof-of-principle, we applied the ZCL algorithm to the analysis of the custom tiling microarray hybridization results of a S. aureus mutant deficient in the sigma B transcription factor. The challenge was to identify those transcripts whose expression decreases in the absence of sigma B. Conclusions: The proposed method archives the best performance in terms of positive predictive value (PPV) while its sensitivity is similar to the other algorithms used for the comparison. The computation time needed to process the transcriptional signals is low as compared with model-based methods and in the same range to those based on the use of filters. Automatic parameter selection has been incorporated and moreover, it can be easily adapted to a parallel implementation. We can conclude that the proposed method is well suited for the analysis of tiling signals, in which transcriptional activity is often hidden in the noise. Finally, the quantification and differential expression analysis of S. aureus dataset have demonstrated the valuable utility of this novel device to the biological analysis of the S. aureus transcriptome., This work was supported by the Spanish Torres-Quevedo fellowship [PTQ-08-03-07769] to VS. ATA and AMB were supported by Spanish Ministry of Science and Innovation ‘Ramón y Cajal’ contracts. This work was supported by the Spanish Ministry of Science and Innovation Grants BIO2008-05284-C02-01, BFU2011-23222, ERA-NET Pathogenomics PIM2010EPA-00606 and the agreement between ‘Fundación para la Investigación médica aplicada’ (FIMA) and the ’UTE project CIMA’.