DNA-Binding Specificity Changes in the Evolution of Forkhead Transcription Factors

The evolution of transcriptional regulatory networks entails the expansion and diversification of transcription factor (TF) families. The forkhead family of TFs, defined by a highly conserved winged helix DNA-binding domain (DBD), has diverged into dozens of subfamilies in animals, fungi, and related protists. We have used a combination of maximum-likelihood phylogenetic inference and independent, comprehensive functional assays of DNA-binding capacity to explore the evolution of DNA-binding specificity within the forkhead family. We present converging evidence that similar alternative sequence preferences have arisen repeatedly and independently in the course of forkhead evolution. The vast majority of DNA-binding specificity changes we observed are not explained by alterations in the known DNA-contacting amino acid residues conferring specificity for canonical forkhead binding sites. Intriguingly, we have found forkhead DBDs that retain the ability to bind very specifically to two completely distinct DNA sequence motifs.We propose an alternate specificity-determining mechanism whereby conformational rearrangements of the DBD broaden the spectrum of sequence motifs that a TF can recognize. DNA-binding bispecificity suggests a previously undescribed source of modularity and flexibility in gene regulation and may play an important role in the evolution of transcriptional regulatory networks.Organismic and Evolutionary Biolog

Evolution of small putative group I introns in the SSU rRNA gene locus of Phialophora species

<p>Abstract</p> <p>Background</p> <p>Group I introns (specifically subgroup IC1) are common in the nuclear ribosomal RNA genes of fungi. While most range in length from more than 200 to nearly 1800 nucleotides (nt) in length, several small putative (or degenerate) group I introns have been described that are between 56 and 81 nt. Although small, previously we demonstrated that the <it>Pa</it>SSU intron in the rRNA small subunit gene of <it>Phialophora americana </it>isolate Wang 1046 is capable of <it>in vitro </it>splicing using a standard group I intron pathway, thus qualifying it as a functional ribozyme.</p> <p>Findings</p> <p>Here, we describe eight short putative group I introns, ranging in length from 63 to 75 nt, in the rRNA small subunit genes of <it>Phialophora </it>isolates, a fungal genus that ranges from saprobic to pathogenic on plants and animals. All contain putative pairing regions P1, P7, and P10, as well as a pairing region formed between the middle of the intron and part of the 3' exon. The other pairing regions common in the core of standard group I introns are absent. However, parts of the 3' exon may aid in the stabilization of these small introns. Although the eight putative group I introns were from at least three species of <it>Phialophora</it>, phylogenetic analysis indicated that the eight are monophyletic. They are also monophyletic with the small introns of two lichen-forming fungi, <it>Porpidia crustulata </it>and <it>Arthonia lapidicola</it>.</p> <p>Conclusions</p> <p>The small putative group I introns in <it>Phialophora </it>have common features that may represent group I introns at their minima. They appear to have a single origin as indicated by their monophyly in phylogenetic analyses.</p

Heterogeneity and organization of the ribosomal RNA genes of Cucurbita maxima

Thirty-six clones were recovered from Cucurbita maxima genomic DNA which had been enriched for rDNA and cleaved at the unique repeat unit Hin d III site. Twenty-nine of these, which contain complete rDNA units, were compared to a standard whose intergenic spacer (IGS) nucleotide sequence has been determined. Twenty-one are identical in length and restriction site pattern. Eight which differ from the standard in length do so because of addition or deletion of varying numbers of IGS subrepetitive units of two different classes, with four of the length variants being different in both of these classes. Seven clones were isolated which contain incomplete repeat units, six of which are composites of rDNA and non-rDNA material. They have been cleaved at the unique rDNA Hin d III site at one end and at a non-rDNA Hin d III site at the other. We consider it most likely that these are derived from the termini of repeat unit tandem arrays, although other explanations are possible. Twelve individual plants of two different cultivars were examined for heterogeneity of IGS length distribution. They all appear to be identical in this regard.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/43423/1/11103_2004_Article_BF00019390.pd

Variability in a dominant block to SIV early reverse transcription in rhesus monkey cells predicts in vivo viral replication and time to death

While it has long been appreciated that there is considerable variability in host containment of HIV/SIV replication, the determinants of that variability are not fully understood. Previous studies demonstrated that the degree of permissivity of a macaque's peripheral blood mononuclear cells (PBMC) for infection with simian immunodeficiency virus (SIV) in vitro predicted that animal's peak plasma virus RNA levels following SIV infection in vivo. The present study was conducted to define the mechanisms underlying the variable intrinsic susceptibility of rhesus monkey PBMC to SIVsmE660 infection. In a cohort of 15 unrelated Indian-origin rhesus monkeys, infectability of PBMC of individual animals with SIVsmE660, as defined by tissue culture infectious dose (TCID50), varied by more than 3 logs and was a stable phenotype over time. Susceptibility of a monkey's PBMC to wild type SIVsmE660 infection correlated with the susceptibility of that monkey's PBMC to infection with VSV-G pseudotyped SIVsm543-GFP. Moreover, the permissivity of an individual monkey's PBMC for infection with this construct correlated with the permissivity of a B-lymphoblastoid cell line (B-LCL) generated from PBMC of the same animal. We found that the degree of intrinsic resistance of monkey B-LCL correlated with the copy number of early reverse transcription (ERT) SIV DNA. The resistance of monkey B-LCL to SIVsmE660 replication could be abrogated by preincubation of cells with the SIV virus-like particles (VLPs) and SIV resistance phenotype could be transferred to a SIV susceptible B-LCL through cell fusion. Finally, we observed a positive correlation between susceptibility of monkey B-LCL to SIV infection with a VSV-G pseudotyped SIV-GFP construct in vitro and both the peak plasma virus RNA levels in vivo and time to death following wild type SIV infection. These findings suggest that a dominant early RT restricting factor that can be saturated by SIV capsid may contribute to the variable resistance to SIV infection in rhesus monkey B-LCL and that this differential intrinsic susceptibility contributes to the clinical outcome of an SIV infection

Setting the standard: multidisciplinary hallmarks for structural, equitable and tracked antibiotic policy

There is increasing concern globally about the enormity of the threats posed by antimicrobial resistance (AMR) to human, animal, plant and environmental health. A proliferation of international, national and institutional reports on the problems posed by AMR and the need for antibiotic stewardship have galvanised attention on the global stage. However, the AMR community increasingly laments a lack of action, often identified as an ‘implementation gap’. At a policy level, the design of internationally salient solutions that are able to address AMR’s interconnected biological and social (historical, political, economic and cultural) dimensions is not straightforward. This multidisciplinary paper responds by asking two basic questions: (A) Is a universal approach to AMR policy and antibiotic stewardship possible? (B) If yes, what hallmarks characterise ‘good’ antibiotic policy? Our multistage analysis revealed four central challenges facing current international antibiotic policy: metrics, prioritisation, implementation and inequality. In response to this diagnosis, we propose three hallmarks that can support robust international antibiotic policy. Emerging hallmarks for good antibiotic policies are: Structural, Equitable and Tracked. We describe these hallmarks and propose their consideration should aid the design and evaluation of international antibiotic policies with maximal benefit at both local and international scale

High frequencies of elevated alkaline phosphatase activity and rickets exist in extremely low birth weight infants despite current nutritional support

<p>Abstract</p> <p>Background</p> <p>Osteopenia and rickets are common among extremely low birth weight infants (ELBW, <1000 g birth weight) despite current practices of vitamin and mineral supplementation. Few data are available evaluating the usual course of markers of mineral status in this population. Our objectives in this study were to determine the relationship between birth weight (BW) and peak serum alkaline phosphatase activity (P-APA) in ELBW infants and evaluate our experience with the diagnosis of rickets in these infants.</p> <p>Methods</p> <p>We evaluated all ELBW infants admitted to Texas Children's Hospital NICU in 2006 and 2007. Of 211 admissions, we excluded 98 patients who were admitted at >30 days of age or did not survive/stay for >6 weeks. Bone radiographs obtained in 32 infants were reviewed by a radiologist masked to laboratory values.</p> <p>Results</p> <p>In this cohort of 113 infants, P-APA was found to have a significant inverse relationship with BW, gestational age and serum phosphorus. In paired comparisons, P-APA of infants <600 g (957 ± 346 IU/L, n = 20) and infants 600–800 g (808 ± 323 IU/L, n = 43) were both significantly higher than P-APA of infants 800–1000 g (615 ± 252 IU/L, n = 50), p < 0.01. Thirty-two patients had radiographic evaluation for evidence of rickets, based on P-APA greater than 800 IU/L, parenteral nutrition greater than 3 to 4 weeks, or clinical suspicion. Of these, 18 showed radiologic rickets and 14 showed osteopenia without rickets. Infants with BW <600 g were more likely to have radiologic rickets (10/20 infants) compared to those with BW 600–800 g (6/43 infants) and BW 800–1000 g (2/50 infants), p < 0.01 for each. P-APA was not significantly higher in infants with radiologic rickets (1078 ± 356 IU/L) compared to those without radiologic evidence of rickets (943 ± 346, p = 0.18).</p> <p>Conclusion</p> <p>Elevation of P-APA >600 IU/L was very common in ELBW infants. BW was significantly inversely related to both P-APA and radiologic rickets. No single value of P-APA was related to radiological findings of rickets. Given the very high risk of osteopenia and rickets among ELBW infants, we recommend consideration of early screening and early mineral supplementation, especially among infants <600 g BW.</p

Bone Marrow Transplantation Results in Human Donor Blood Cells Acquiring and Displaying Mouse Recipient Class I MHC and CD45 Antigens on Their Surface

Background: Mouse models of human disease are invaluable for determining the differentiation ability and functional capacity of stem cells. The best example is bone marrow transplants for studies of hematopoietic stem cells. For organ studies, the interpretation of the data can be difficult as transdifferentiation, cell fusion or surface antigen transfer (trogocytosis) can be misinterpreted as differentiation. These events have not been investigated in hematopoietic stem cell transplant models. Methodology/Principal Findings: In this study we investigated fusion and trogocytosis involving blood cells during bone marrow transplantation using a xenograft model. We report that using a standard SCID repopulating assay almost 100 % of the human donor cells appear as hybrid blood cells containing both mouse and human surface antigens. Conclusion/Significance: Hybrid cells are not the result of cell-cell fusion events but appear to be due to efficient surface antigen transfer, a process referred to as trogocytosis. Antigen transfer appears to be non-random and includes all donor cells regardless of sub-type. We also demonstrate that irradiation preconditioning enhances the frequency of hybrid cell

A longitudinal survey of African animal trypanosomiasis in domestic cattle on the Jos Plateau, Nigeria:prevalence, distribution and risk factors

BACKGROUND: Trypanosomiasis is a widespread disease of livestock in Nigeria and a major constraint to the rural economy. The Jos Plateau, Nigeria was free from tsetse flies and the trypanosomes they transmit due to its high altitude and the absence of animal trypanosomiasis attracted large numbers of cattle-keeping pastoralists to inhabit the plateau. The Jos Plateau now plays a significant role in the national cattle industry, accommodating approximately 7% of the national herd and supporting 300,000 pastoralists and over one million cattle. However, during the past two decades tsetse flies have invaded the Jos Plateau and animal trypanosomiasis has become a significant problem for livestock keepers. METHODS: In 2008 a longitudinal two-stage cluster survey on the Jos Plateau. Cattle were sampled in the dry, early wet and late wet seasons. Parasite identification was undertaken using species-specific polymerase chain reactions to determine the prevalence and distribution bovine trypanosomiasis. Logistic regression was performed to determine risk factors for disease. RESULTS: The prevalence of bovine trypanosomiasis (Trypanosoma brucei brucei, Trypanosoma congolense savannah, Trypanosoma vivax) across the Jos Plateau was found to be high at 46.8% (39.0 – 54.5%) and significant, seasonal variation was observed between the dry season and the end of the wet season. T. b. brucei was observed at a prevalence of 3.2% (1% – 5.5%); T. congolense at 27.7% (21.8% - 33.6%) and T. vivax at 26.7% (18.2% - 35.3%). High individual variation was observed in trypanosomiasis prevalence between individual villages on the Plateau, ranging from 8.8% to 95.6%. Altitude was found to be a significant risk factor for trypanosomiasis whilst migration also influenced risk for animal trypanosomiasis. CONCLUSIONS: Trypanosomiasis is now endemic on the Jos Plateau showing high prevalence in cattle and is influenced by seasonality, altitude and migration practices. Attempts to successfully control animal trypanosomiasis on the Plateau will need to take into account the large variability in trypanosomiasis infection rates between villages, the influence of land use, and husbandry and management practices of the pastoralists, all of which affect the epidemiology of the disease

Non-Diversifiable Volatility Risk and Risk Premiums at Earnings Announcements

This study seeks to determine whether earnings announcements pose non-diversifiable volatility risk that commands a risk premium. We find that investors anticipate some earnings announcements to convey news that increases market return volatility and pay a premium to hedge this non-diversifiable risk. In particular, we find evidence of risk premiums embedded in prices of firms' traded options that are significantly positively associated with the extent to which the firms' earnings announcements pose non-diversifiable volatility risk. In addition, we find that volatility risk premiums are concentrated among bellwether firms and result in predictable variation in option straddle returns around earnings announcements. Taken together, our findings show that some earnings announcements pose non-diversifiable volatility risk that commands a risk premium

Non-Invasive In Vivo Imaging of Calcium Signaling in Mice

Rapid and transient elevations of Ca2+ within cellular microdomains play a critical role in the regulation of many signal transduction pathways. Described here is a genetic approach for non-invasive detection of localized Ca2+ concentration ([Ca2+]) rises in live animals using bioluminescence imaging (BLI). Transgenic mice conditionally expressing the Ca2+-sensitive bioluminescent reporter GFP-aequorin targeted to the mitochondrial matrix were studied in several experimental paradigms. Rapid [Ca2+] rises inside the mitochondrial matrix could be readily detected during single-twitch muscle contractions. Whole body patterns of [Ca2+] were monitored in freely moving mice and during epileptic seizures. Furthermore, variations in mitochondrial [Ca2+] correlated to behavioral components of the sleep/wake cycle were observed during prolonged whole body recordings of newborn mice. This non-invasive imaging technique opens new avenues for the analysis of Ca2+ signaling whenever whole body information in freely moving animals is desired, in particular during behavioral and developmental studies