No abstract

Retroelements are usually considered to be eukaryotic elements because of the large number and variety in eukaryotic genomes. By comparison, reverse transcriptases (RTs) are rare in bacteria, with only three characterized classes: retrons, group II introns and diversity-generating retroelements (DGRs). Here, we present the results of a bioinformatic survey that aims to define the landscape of RTs across eubacterial, archaeal and phage genomes. We identify and categorize 1021 RTs, of which the majority are group II introns (73%). Surprisingly, a plethora of novel RTs are found that do not belong to characterized classes. The RTs have 11 domain architectures and are classified into 20 groupings based on sequence similarity, phylogenetic analyses and open reading frame domain structures. Interestingly, group II introns are the only bacterial RTs to exhibit clear evidence for independent mobility, while five other groups have putative functions in defense against phage infection or promotion of phage infection. These examples suggest that additional beneficial functions will be discovered among uncharacterized RTs. The study lays the groundwork for experimental characterization of these highly diverse sequences and has implications for the evolution of retroelements.

Peroxisome proliferator-activated receptor (PPAR)-gamma is a member of the nuclear hormone receptor superfamily that can promote cellular differentiation and organ development. PPARgamma expression has been reported in a number of pulmonary cell types, including inflammatory, mesenchymal, and epithelial cells. We find that PPARgamma is prominently expressed in the airway epithelium in the mouse lung. In an effort to define the physiological role of PPARgamma within the lung, we have ablated PPARgamma using a novel line of mice capable of specifically targeting the airway epithelium. Airway epithelial cell PPARgamma-targeted mice display enlarged airspaces resulting from insufficient postnatal lung maturation. The increase in airspace size is accompanied by alterations in lung physiology, including increased lung volumes and decreased tissue resistance. Genome-wide expression profiling reveals a reduction in structural extracellular matrix (ECM) gene expression in conditionally targeted mice, suggesting a disruption in epithelial-mesenchymal interactions necessary for the establishment of normal lung structure. Expression profiling of airway epithelial cells isolated from conditionally targeted mice indicates PPARgamma regulates genes encoding known PPARgamma targets, additional lipid metabolism enzymes, and markers of cellular differentiation. These data reveal airway epithelial cell PPARgamma is necessary for normal lung structure and function.

Group II introns are a major class of ribozymes found in bacteria, mitochondria, and plastids. Many introns contain reverse transcriptase open reading frames (ORFs) that confer mobility to the introns and allow them to persist as selfish DNAs. Here, we report an updated compilation of group II introns in Eubacteria and Archaea comprising 234 introns. One new phylogenetic class is identified, as well as several specialized lineages. In addition, we undertake a detailed search for ORF-less group II introns in bacterial genomes in order to find undiscovered introns that either entirely lack an ORF or encode a novel ORF. Unlike organellar group II introns, we find only a handful of ORF-less introns in bacteria, suggesting that if a substantial number exist, they must be divergent from known introns. Together, these results highlight the retroelement character of bacterial group II introns, and suggest that their long-term survival is dependent upon retromobility.

BackgroundPuf proteins have important roles in controlling gene expression at the post-transcriptional level by promoting RNA decay and repressing translation. The Pumilio homology domain (PUM-HD) is a conserved region within Puf proteins that binds to RNA with sequence specificity. Although Puf proteins have been well characterized in animal and fungal systems, little is known about the structural and functional characteristics of Puf-like proteins in plants.ResultsThe Arabidopsis and rice genomes code for 26 and 19 Puf-like proteins, respectively, each possessing eight or fewer Puf repeats in their PUM-HD. Key amino acids in the PUM-HD of several of these proteins are conserved with those of animal and fungal homologs, whereas other plant Puf proteins demonstrate extensive variability in these amino acids. Three-dimensional modeling revealed that the predicted structure of this domain in plant Puf proteins provides a suitable surface for binding RNA. Electrophoretic gel mobility shift experiments showed that the Arabidopsis AtPum2 PUM-HD binds with high affinity to BoxB of the Drosophila Nanos Response Element I (NRE1) RNA, whereas a point mutation in the core of the NRE1 resulted in a significant reduction in binding affinity. Transient expression of several of the Arabidopsis Puf proteins as fluorescent protein fusions revealed a dynamic, punctate cytoplasmic pattern of localization for most of these proteins. The presence of predicted nuclear export signals and accumulation of AtPuf proteins in the nucleus after treatment of cells with leptomycin B demonstrated that shuttling of these proteins between the cytosol and nucleus is common among these proteins. In addition to the cytoplasmically enriched AtPum proteins, two AtPum proteins showed nuclear targeting with enrichment in the nucleolus.ConclusionsThe Puf family of RNA-binding proteins in plants consists of a greater number of members than any other model species studied to date. This, along with the amino acid variability observed within their PUM-HDs, suggests that these proteins may be involved in a wide range of post-transcriptional regulatory events that are important in providing plants with the ability to respond rapidly to changes in environmental conditions and throughout development.

Medical and feeding therapy should be the first modality of treatment in patients with laryngeal cleft type I and type II. Factors supporting surgical repair include: 1) clinically apparent aspiration with feeding, 2) severity of pulmonary status, 3) findings on modified barium swallow and chest x-ray, 4) absence of significant comorbid conditions predisposing to aspiration, 5) findings on upper aerodigestive endoscopy, and 6) poor response to medical management and feeding therapy.

Spliceosomal (pre-mRNA) introns have previously been found in eukaryotic protein-coding genes, in the small nuclear RNAs of some fungi, and in the small- and large-subunit ribosomal DNA genes of a limited number of ascomycetes. How the majority of these introns originate remains an open question because few proven cases of recent and pervasive intron origin have been documented. We report here the widespread occurrence of spliceosomal introns (69 introns at 27 different sites) in the small- and large-subunit nuclear-encoded rDNA of lichen-forming and free-living members of the Ascomycota. Our analyses suggest that these spliceosomal introns are of relatively recent origin, i.e., within the Euascomycetes, and have arisen through aberrant reverse-splicing (in trans) of free pre-mRNA introns into rRNAs. The spliceosome itself, and not an external agent (e.g., transposable elements, group II introns), may have given rise to these introns. A nonrandom sequence pattern was found at sites flanking the rRNA spliceosomal introns. This pattern (AG-intron-G) closely resembles the proto-splice site (MAG-intron-R) postulated for intron insertions in pre-mRNA genes. The clustered positions of spliceosomal introns on secondary structures suggest that particular rRNA regions are preferred sites for insertion through reverse-splicing.

Rationale: The mechanisms contributing to alveolar formation are poorly understood. A better understanding of these processes will improve efforts to ameliorate lung disease of the newborn and promote alveolar repair in the adult. Previous studies have identified impaired alveogenesis in mice bearing compound mutations of fibroblast growth factor (FGF) receptors (FGFRs) 3 and 4, indicating that these receptors cooperatively promote postnatal alveolar formation. Objectives: To determine the molecular and cellular mechanisms of FGF-mediated alveolar formation. Methods: Compound FGFR3/FGFR4-deficient mice were assessed for temporal changes in lung growth, airspace morphometry, and genome-wide expression. Observed gene expression changes were validated using quantitative real-time RT-PCR, tissue biochemistry, histochemistry, and ELISA. Autocrine and paracrine regulatory mechanisms were investigated using isolated lung mesenchymal cells and type II pneumocytes. Measurements and Main Results: Quantitative analysis of airspace ontogeny confirmed a failure of secondary crest elongation in compound mutant mice. Genome-wide expression profiling identified molecular alterations in these mice involving aberrant expression of numerous extracellular matrix molecules. Biochemical and histochemical analysis confirmed changes in elastic fiber gene expression resulted in temporal increases in elastin deposition with the loss of typical spatial restriction. No abnormalities in elastic fiber gene expression were observed in isolated mesenchymal cells, indicating that abnormal elastogenesis in compound mutant mice is not cell autonomous. Increased expression of paracrine factors, including insulin-like growth factor21, in freshly-isolated type II pneumocytes indicated that these cells contribute to the observed pathology. Conclusions: Epithelial/mesenchymal signaling mechanisms appear to contribute to FGFR-dependent alveolar elastogenesis and proper airspace formation.