An increasing number of studies have shown the key role that RNA polymerase II (RNA Pol II) elongation plays in gene regulation. We systematically examine how various enhancers, promoters, and gene ...body composition influence the RNA Pol II elongation rate through a single-cell-resolution live imaging assay. By using reporter constructs containing 5′ MS2 and 3′ PP7 repeating stem loops, we quantify the rate of RNA Pol II elongation in live Drosophila embryos. We find that promoters and exonic gene lengths have no effect on elongation rate, while enhancers and the presence of long introns may significantly change how quickly RNA Pol II moves across a gene. Furthermore, we observe in multiple constructs that the RNA Pol II elongation rate accelerates after the transcriptional onset of nuclear cycle 14 in Drosophila embryos. Our study provides a single-cell view of various mechanisms that affect the dynamic RNA Pol II elongation rate, ultimately affecting the rate of mRNA production.
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•Enhancer strength is inversely correlated with RNA Pol II elongation rate•Exonic gene length has no impact on the elongation rate•Long-intron-containing genes show a faster elongation rate than exon-only genes•RNA Pol II elongation rate is dynamic during nuclear cycle 14
Keller et al. explore how enhancers, promoters, and the exon/intron ratio affect the rate of RNA Pol II elongation using live imaging in Drosophila embryos. Promoters and exonic gene lengths do not affect elongation rates, while enhancers and long-intron-containing genes have a faster rate. The elongation rate is dynamic during nuclear cycle 14.
The regulatory specificity of a gene is determined by the structure of its enhancers, which contain multiple transcription factor binding sites. A unique combination of transcription factor binding ...sites in an enhancer determines the boundary of target gene expression, and their disruption often leads to developmental defects. Despite extensive characterization of binding motifs in an enhancer, it is still unclear how each binding site contributes to overall transcriptional activity. Using live imaging, quantitative analysis, and mathematical modeling, we measured the contribution of individual binding sites in transcriptional regulation. We show that binding site arrangement within the Rho-GTPase component t48 enhancer mediates the expression boundary by mainly regulating the timing of transcriptional activation along the dorsoventral axis of Drosophila embryos. By tuning the binding affinity of the Dorsal (Dl) and Zelda (Zld) sites, we show that single site modulations are sufficient to induce significant changes in transcription. Yet, no one site seems to have a dominant role; rather, multiple sites synergistically drive increases in transcriptional activity. Interestingly, Dl and Zld demonstrate distinct roles in transcriptional regulation. Dl site modulations change spatial boundaries of t48, mostly by affecting the timing of activation and bursting frequency rather than transcriptional amplitude or bursting duration. However, modulating the binding site for the pioneer factor Zld affects both the timing of activation and amplitude, suggesting that Zld may potentiate higher Dl recruitment to target DNAs. We propose that such finetuning of dynamic gene control via enhancer structure may play an important role in ensuring normal development.
Heterochromatin, typically marked by histone H3 trimethylation at lysine 9 (H3K9me3) or lysine 27 (H3K27me3), represses different protein-coding genes in different cells, as well as repetitive ...elements. The basis for locus specificity is unclear. Previously, we identified 172 proteins that are embedded in sonication-resistant heterochromatin (srHC) harbouring H3K9me3. Here, we investigate in humans how 97 of the H3K9me3-srHC proteins repress heterochromatic genes. We reveal four groups of srHC proteins that each repress many common genes and repeat elements. Two groups repress H3K9me3-embedded genes with different extents of flanking srHC, one group is specific for srHC genes with H3K9me3 and H3K27me3, and one group is specific for genes with srHC as the primary feature. We find that the enhancer of rudimentary homologue (ERH) is conserved from Schizosaccharomyces pombe in repressing meiotic genes and, in humans, now represses other lineage-specific genes and repeat elements. The study greatly expands our understanding of H3K9me3-based gene repression in vertebrates.
The great hammerhead (Sphyrna mokarran) is a highly mobile, large‐bodied shark primarily found in coastal‐pelagic and semi‐oceanic waters across a circumtropical range. It is a target or by‐catch ...species in multiple fisheries, and as a result, rapid population declines have occurred in many regions. These declines have contributed to the species being assessed as globally critically endangered on the IUCN Red List. Although conservation and management measures have yielded promising results in some regions, such as the United States, high levels of at‐vessel and post‐release mortality remain a major concern to the species population recovery. This examined the vertical space use and thermal range of pop‐off archival satellite–tagged S. mokarran in the western North Atlantic Ocean, expanding the understanding of the ecological niche of this species and providing insight into by‐catch mitigation strategies for fisheries managers. The results showed that S. mokarran predominantly used shallow depths (75% of records <30 m) and had a narrow temperature range (89% of records between 23 and 28°C). Individual differences in depth use were apparent, and a strong diel cycle was observed, with sharks occupying significantly deeper depths during the daytime. Furthermore, two individuals were confirmed pregnant with one migrating from the Bahamas to South Carolina, U.S.A., providing further evidence of regional connectivity and parturition off the U.S. East Coast. The findings suggest that S. mokarran may be vulnerable to incidental capture in the western North Atlantic commercial longline fisheries due to substantial vertical overlap between the species and the gear. The results can be incorporated into conservation and management efforts to develop and/or refine mitigation measures focused on reducing the by‐catch and associated mortality of this species, which can ultimately aide S. mokarran population recovery in areas with poor conservation status.
Glucagon-like peptide-1 (GLP-1) is a natural agonist for GLP-1R, a G protein-coupled receptor (GPCR) on the surface of pancreatic β cells. GLP-1R agoinsts are attractive for treatment of type 2 ...diabetes, but GLP-1 itself is rapidly degraded by peptidases in vivo. We describe a design strategy for retaining GLP-1-like activity while engendering prolonged activity in vivo, based on strategic replacement of native α residues with conformationally constrained β-amino acid residues. This backbone-modification approach may be useful for developing stabilized analogues of other peptide hormones.
Invasive fungal diseases are generally difficult to treat and often fatal. The therapeutic agents available to treat fungi are limited, and there is a critical need for new agents to combat these ...deadly infections. Antifungal compound development has been hindered by the challenge of creating agents that are highly active against fungal pathogens but not toxic to the host. Host defense peptides (HDPs) are produced by eukaryotes as a component of the innate immune response to pathogens and have served as inspiration for the development of many new antibacterial compounds. HDP mimics, however, have largely failed to exhibit potent and selective antifungal activity. Here, we present an HDP-like nylon-3 copolymer that is effective against diverse fungi while displaying only mild to moderate toxicity toward mammalian cells. This polymer is active on its own and in synergy with existing antifungal drugs against multiple species of
and
, reaching levels of efficacy comparable to those of the clinical agents amphotericin B and fluconazole in some cases. In addition, the polymer acts synergistically with azoles against different species of
, including some azole-resistant strains. These findings indicate that nylon-3 polymers are a promising lead for development of new antifungal therapeutic strategies.
Diabetic kidney disease (DKD), the most common cause of kidney failure, is a frequent complication of diabetes and obesity, and yet to date, treatments to halt its progression are lacking. We analyze ...kidney single-cell transcriptomic profiles from DKD patients and two DKD mouse models at multiple time points along disease progression—high-fat diet (HFD)-fed mice aged to 90–100 weeks and BTBR ob/ob mice (a genetic model)—and report an expanding population of macrophages with high expression of triggering receptor expressed on myeloid cells 2 (TREM2) in HFD-fed mice. TREM2high macrophages are enriched in obese and diabetic patients, in contrast to hypertensive patients or healthy controls in an independent validation cohort. Trem2 knockout mice on an HFD have worsening kidney filter damage and increased tubular epithelial cell injury, all signs of worsening DKD. Together, our studies suggest that strategies to enhance kidney TREM2high macrophages may provide therapeutic benefits for DKD.
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•TREM2high macrophages reside in the human adult kidney•Trem2high macrophages increase in frequency in a high-fat-diet mouse model of diabetic kidney disease•Human adult diabetic and obese kidney tissue has increased frequency of TREM2high macrophages•Trem2 deletion promotes kidney injury after exposure to a high-fat diet but not a regular diet
Subramanian et al. identify a macrophage population expressing a TREM2high transcriptional program in the human adult kidney, matching a homologous population in other tissues (adipose, heart, and liver). Trem2high macrophages expand in diabetic kidney disease in both a high-fat-diet-fed mouse model and an independent cohort of patients. Trem2 deletion results in hastened kidney injury in the presence of a high-fat diet, suggesting a role of these macrophages as responders to local tissue injury in obesity- and diabetes-driven kidney injury.
Investigating individual preferences using Social Network Analysis (SNA) has contributed to understanding group formations, which hold significant evolutionary value, and the dynamics occurring ...within them across the animal kingdom. Groups of sharks have been observed for centuries, and understanding the mechanisms that facilitate their formation furthers our comprehension of the adaptive value of sociality. However, there is a dearth of quantitative analyses on these mechanisms. The preference for familiars can influence group formation and facilitate an increased level of fitness. This preference has been documented across many vertebrate classes, but has not been shown to have an overall effect on partner preference in chondrichthyan fishes. This deficiency is significant due to the taxa's ecological diversification and importance. In this study, we used behavioral experiments to assess the potential role of familiarity in group formation and on social behavior in a large marine vertebrate. Juvenile lemon sharks, Negaprion brevirostris, (N=23) in Bimini, Bahamas, were separated into holding pens according to their size and given 14days to familiarize with their cohort. Two pairs of sharks were selected from separate holding pens and introduced in a social network pen for a behavioral trial. SNA indicated that N. brevirostris showed a significant preference for familiar individuals, performing more social interactions, such as closely following, with individuals from their holding pen. We suggest that this preference for familiars is developed through repeated interactions and facilitates the advantages of maintaining stable behavioral interactions, which results in spatial isolation from unfamiliars. These findings advance our understanding of the mechanisms driving social structure in N. brevirostris and other large marine predators, which contribute to an animal's overall fitness and evolutionary adaptability.
•Lemon sharks showed a significant preference for familiar individuals.•Capture location and length did not significantly impact interactions.•Findings suggest individuals are potentially capable of individual recognition.•Findings offer explanatory mechanism for wild trials with repeated partnership.
Understanding the dimensions of fungal diversity has major implications for the control of diseases in humans, plants, and animals and in the overall health of ecosystems on the planet. One ancient ...evolutionary strategy organisms use to manage interactions with microbes, including fungi, is to produce host defense peptides (HDPs). HDPs and their synthetic analogs have been subjects of interest as potential therapeutic agents. Due to increases in fungal disease worldwide, there is great interest in developing novel antifungal agents. Here we describe activity of polymeric HDP analogs against fungi from 18 pathogenic genera composed of 41 species and 72 isolates. The synthetic polymers are members of the nylon-3 family (poly-β-amino acid materials). Three different nylon-3 polymers show high efficacy against surprisingly diverse fungi. Across the phylogenetic spectrum (with the exception of
species), yeasts, dermatophytes, dimorphic fungi, and molds were all sensitive to the effects of these polymers. Even fungi intrinsically resistant to current antifungal drugs, such as the causative agents of mucormycosis (
spp.) and those with acquired resistance to azole drugs, showed nylon-3 polymer sensitivity. In addition, the emerging pathogens
(cause of white nose syndrome in bats) and
(cause of nosocomial infections of humans) were also sensitive. The three nylon-3 polymers exhibited relatively low toxicity toward mammalian cells. These findings raise the possibility that nylon-3 polymers could be useful against fungi for which there are only limited and/or no antifungal agents available at present.
Fungi reside in all ecosystems on earth and impart both positive and negative effects on human, plant, and animal health. Fungal disease is on the rise worldwide, and there is a critical need for more effective and less toxic antifungal agents. Nylon-3 polymers are short, sequence random, poly-β-amino acid materials that can be designed to manifest antimicrobial properties. Here, we describe three nylon-3 polymers with potent activity against the most phylogenetically diverse set of fungi evaluated thus far in a single study. In contrast to traditional peptides, nylon-3 polymers are highly stable to proteolytic degradation and can be produced efficiently in large quantities at low cost. The ability to modify nylon-3 polymer composition easily creates an opportunity to tailor efficacy and toxicity, which makes these materials attractive as potential broad-spectrum antifungal therapeutics.
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