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Estrogen is known for its positive stimulatory effects on surfactant proteins. ErbB4 receptor and its ligand neuregulin (NRG) positively stimulate lung development. ErbB receptors interact with nuclear receptors and ErbB4 co-regulates estrogen receptor (ER) α expression in breast cells. ERβ is highly expressed in pneumocytes and its deletion leads to fewer alveoli and reduced elastic recoil. A similar picture was seen in ErbB4-deleted lungs. We hypothesized that estrogen signals its effect on surfactant protein B (Sftpb) expression through interactions of ERβ and ErbB4. Estrogen and NRG treatment decreased cell numbers and stimulated Sftpb expression in type II cells. Estrogen and NRG both stimulated phosphorylation of ERβ and co-localization of both receptors. Overexpression of ERβ increased the cell number and Sftpb expression, which was further augmented by estrogen and NRG. Finally, estrogen and NRG stimulated ERβ and ErbB4 binding to the Sftpb promoter and the overexpression of these receptors stimulated Sftpb promoter activation, which was further enhanced by estrogen and NRG. The stimulatory effect of estrogen and NRG was abolished in ErbB4 deletion and reconstituted by re-expression of full-length ErbB4 in fetal ErbB4-deleted type II cells. Estrogen-induced nuclear translocation of ErbB4 required the cleavage site but not the nuclear localization signal of the ErbB4 receptor, suggesting that ERβ might function as a nuclear chaperone for ErbB4. These studies demonstrate that estrogen effects on Sftpb expression require an interaction of ERβ and ErbB4. We speculate that the stimulatory effects of estrogen on Sftpb are under transcriptional control of ErbB4.

BackgroundEnterococcus faecalis is a multifaceted microorganism known to act as a beneficial intestinal commensal bacterium. It is also a dreaded nosocomial pathogen causing life-threatening infections in hospitalised patients. Isolates of a distinct MLST type ST40 represent the most frequent strain type of this species, distributed worldwide and originating from various sources (animal, human, environmental) and different conditions (colonisation/infection). Since enterococci are known to be highly recombinogenic we determined to analyse the microevolution and niche adaptation of this highly distributed clonal type.ResultsWe compared a set of 42 ST40 isolates by assessing key molecular determinants, performing whole genome sequencing (WGS) and a number of phenotypic assays including resistance profiling, formation of biofilm and utilisation of carbon sources. We generated the first circular closed reference genome of an E. faecalis isolate D32 of animal origin and compared it with the genomes of other reference strains. D32 was used as a template for detailed WGS comparisons of high-quality draft genomes of 14 ST40 isolates. Genomic and phylogenetic analyses suggest a high level of similarity regarding the core genome, also demonstrated by similar carbon utilisation patterns. Distribution of known and putative virulence-associated genes did not differentiate between ST40 strains from a commensal and clinical background or an animal or human source. Further analyses of mobile genetic elements (MGE) revealed genomic diversity owed to: (1) a modularly structured pathogenicity island; (2) a site-specifically integrated and previously unknown genomic island of 138 kb in two strains putatively involved in exopolysaccharide synthesis; and (3) isolate-specific plasmid and phage patterns. Moreover, we used different cell-biological and animal experiments to compare the isolate D32 with a closely related ST40 endocarditis isolate whose draft genome sequence was also generated. D32 generally showed a greater capacity of adherence to human cell lines and an increased pathogenic potential in various animal models in combination with an even faster growth in vivo (not in vitro).ConclusionMolecular, genomic and phenotypic analysis of representative isolates of a major clone of E. faecalis MLST ST40 revealed new insights into the microbiology of a commensal bacterium which can turn into a conditional pathogen.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1367-x) contains supplementary material, which is available to authorized users.

The complete and annotated genome sequence of Enterococcus faecalis D32, a commensal strain isolated from a Danish pig, suggests putative adaptation to the porcine host and absence of distinct virulence-associated traits. Enterococcus faecalis is a common colonizer of human and animal intestinal tracts (6). Certain E. faecalis strains are also used in the food industry and as a probiotic mixture component (5). In contrast, E. faecalis is one of the leading causes of health careassociated infections, especially among patients in intensive care.E. faecalis strain D32 was isolated from pig feces in 2001 as part of the Danish Integrated Antimicrobial Resistance Monitoring and Research Programme (15) and belongs to multilocus sequence type 40 (ST40), the most frequent ST in a large and diverse E. faecalis strain collection (10). E. faecalis strains exhibiting ST40 did not show any ecological preference and were isolated from infections and colonizations of animals and humans, as well as from food (10,17).De novo genome sequencing of D32 was performed by standard 454 pyrosequencing on a GS-FLX system (Roche/454 Life Sciences). It resulted in 125,952 reads with an average length of 392 bp and 71 contigs. Additional sequencing of an 8-kb-long paired-end library (Roche/454 Life Sciences; performed at Eurofins MWG-Operon) generated 97,660 reads with an average read length of 346 bp. Using Celera Assembler software, both data sets could be assembled into three scaffolds. The contig orientation was compared to that of reference strain V583 (12) by progressiveMauve alignment (3). Remaining gaps and assembly ambiguities ("indel" errors) were corrected by sequencing of PCR amplicons. Annotation was performed with the GenDB and RAST annotation services (1, 11). For subsequent manual annotation, we used NCBI ORF Finder and BLAST comparisons against available reference genomes (13).The completed E. faecalis D32 circular chromosome contains 2,987,450 bp with a GC content of 37.49%. Annotation created a total of 2,890 coding sequences (CDS), 62 tRNA-encoding genes, and four entire rRNA-encoding operons. The web application Prophage Finder (2) identified four putative prophages integrated into the chromosome. High-level resistance to streptomycin is chromosomally encoded by an aminoglycoside 6-adenyltransferase gene (aadK). The pathogenicity island (PAI) in D32 contains a bile acid hydrolase (cbh) and lactose metabolic pathway genes (lacABCDEFG) but lacks common markers such as the enterococcal surface protein gene (esp) and the cytolysin operon (8, 15) present in the original PAI in MMH594 (9, 14). Additionally, a previously novel and uncharacterized genomic island (138 kb) is integrated at the attachment site of the conjugative vanB transposon in V583 (12).Contig assembly revealed two plasmid scaffolds, EFD32pA and EFD32pB. Plasmid EFD32pA (12,893 bp; 14 CDS) represents an Inc18 broad-host-range plasmid deduced from the similarity of its putative replicase to rep family 1 genes (7,16). It also contained an rRNA adenine N-6-methylt...