Layered Yttrium Hydroxide <i>l</i>‑Y(OH)₃ Luminescent Adsorbent for Detection and Recovery of Phosphate from Water over a Wide pH Range

Layered yttrium hydroxide, <i>l</i>-Y­(OH)₃, has been explored as a representative member of the layered rare earth hydroxide family (<i>l</i>-RE­(OH)₃; RE = rare earths) for removal and recovery of phosphate from aqueous solution. Compared to the hexagonal form, <i>h</i>-Y­(OH)₃, which has a weakly positive surface charge only at low pH, the layered polymorph composed of hydroxocation layers exhibited a high point of zero charge (pH_pzc ∼ 11) and significantly enhanced adsorptive ability for anions over a wide pH range. The Langmuir isotherm model and pseudo-second-order kinetic model were adopted to explain the phosphate adsorption on <i>l</i>-Y­(OH)₃. This new adsorbent revealed high capacity, efficiency, stability, selectivity, and reusability in adsorption of phosphate from a single electrolyte as well as natural waters containing competing anions. Essentially complete phosphate recovery from aqueous solutions at low phosphate concentrations (2.0 mg of P/L) was demonstrated with an adsorbent dosage of 0.025–0.5 g/L. The adsorption of phosphate was accompanied by an increase in the solution pH, suggesting a release of OH^– ions during the adsorption reaction. In particular, when Ce³⁺ and Tb³⁺ were co-doped (<i>l</i>-Y­(OH)₃:Ce,Tb), phosphate adsorption led to the characteristic ⁵D₄ → ⁷F_<i>J</i> (<i>J</i> = 6, 5, and 4) emissions of Tb³⁺ under commercial 312 nm UV irradiation. The photoluminescence of phosphate-adsorbed <i>l</i>-Y­(OH)₃:Ce,Tb provided evidence of the inner-sphere complexing mechanism involving the formation of Y­(Ce,Tb)–O–P bonds through which the energy transfer can occur. The “luminescence-on” behavior of <i>l</i>-Y­(OH)₃:Ce,Tb by phosphate adsorption was employed to detect and recover phosphorus at low concentrations in deionized water, mineral water, tap water, and river water

An alternatively spliced form affecting the Marked Box domain of <i>Drosophila</i> E2F1 is required for proper cell cycle regulation

<div><p>Across metazoans, cell cycle progression is regulated by E2F family transcription factors that can function as either transcriptional activators or repressors. For decades, the <i>Drosophila</i> E2F family has been viewed as a streamlined RB/E2F network, consisting of one activator (dE2F1) and one repressor (dE2F2). Here, we report that an uncharacterized isoform of dE2F1, hereon called dE2F1b, plays an important function during development and is functionally distinct from the widely-studied dE2F1 isoform, dE2F1a. dE2F1b contains an additional exon that inserts 16 amino acids to the evolutionarily conserved Marked Box domain. Analysis of <i>de2f1b</i>-specific mutants generated via CRISPR/Cas9 indicates that dE2F1b is a critical regulator of the cell cycle during development. This is particularly evident in endocycling salivary glands in which a tight control of dE2F1 activity is required. Interestingly, close examination of mitotic tissues such as eye and wing imaginal discs suggests that dE2F1b plays a repressive function as cells exit from the cell cycle. We also provide evidence demonstrating that dE2F1b differentially interacts with RBF1 and alters the recruitment of RBF1 and dE2F1 to promoters. Collectively, our data suggest that dE2F1b is a novel member of the E2F family, revealing a previously unappreciated complexity in the <i>Drosophila</i> RB/E2F network.</p></div

<i>de2f1b</i> mutant salivary glands display disrupted endocycle progression.

<p>(A) Late stage (105–110 hour After Egg Laying, AEL) salivary glands from a control, <i>dDP</i> and <i>de2f1b</i> mutant larvae are stained with DAPI to visualize nuclei. Yellow arrows show an example of nuclei with different sizes. (B) A box and whisker graph showing the distribution of nuclear area (μm²) in control, <i>dDP</i> and <i>de2f1b</i> mutant salivary glands is presented. Three salivary glands are used for each genotype. (C) Average of the relative standard deviation of nuclear area of indicated genotype is shown. Values represent the mean of triplicated biological replicates and error bars represent s.d. One-way ANOVA was used to calculate statistical significance, where ns = p>0.05; *** = p≤0.001. (D) Scatter plot representing DAPI intensity from 105–110 hour AEL salivary glands from indicated genotypes. The values in the y-axis represent fluorescence intensity of each nuclei. Average mean fluorescence intensity was compared between control group (1–3) and <i>de2f1b</i> mutant group (1–3). ** = p ≤ 0.01 using two-tailed t-test. (E) Salivary glands of indicated genotype are labeled with EdU to visualize S-phase cells. Scale bars for all salivary glands indicate 100 μm. P: proximal end. D: distal end. FB: Fat body.</p

Two alternatively spliced forms of <i>de2f1</i>, <i>def1a</i> and <i>de2f1b</i>, are required to rescue larval lethality of <i>de2f1</i> mutant flies.

<p>(A) A schematic of the coding region of the <i>de2f1</i> gene showing the two alternatively spliced forms of <i>de2f1</i>, <i>de2f1a</i> and <i>de2f1b</i>. The exon highlighted in red indicates the <i>de2f1b</i>-specific exon, exon 3b. Primer locations for absolute quantification of total <i>de2f1</i> and <i>de2f1b</i> are indicated in blue and red, respectively. (B) RT-qPCR is performed to estimate the absolute levels of the total <i>de2f1</i> RNA and <i>de2f1b-</i>specific RNA at different developmental stages (E: embryonic, L: larval, P: pupal and A: adult). The y-axis indicates the copy number per 1 unit of cDNA (1 unit of cDNA represents 25 ng of RNA). The error bars indicate standard error of the mean (s.e.m.) of triplicated independent biological replicates. (C) The copy numbers of the total <i>de2f1</i> RNA and <i>de2f1b-</i>specific RNA in indicated third instar larval tissues are determined. The graph shows the percentage contribution of the <i>de2f1b-</i>specific transcript to the total <i>de2f1</i>. Error bars indicate s.e.m. (D) Molecular complementation tests are performed in <i>de2f1</i> mutants by expressing either <i>de2f1a</i> or <i>de2f1b</i> alone or by expressing both. A survival curve from third instar larval stage (L3) to adult of indicated genotypes is presented. The percent survival is determined by comparing the observed frequency of survival to the expected frequency based on Mendelian ratio (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1007204#sec004" target="_blank">materials and methods</a>). (E) A control (<i>yw</i>), <i>de2f1</i> mutant (<i>de2f1</i>^<i>729</i><i>/Df</i>) and rescued (<i>de2f1</i>^<i>729</i><i>/Df</i>, <i>UbiG4>dE2F1+dE2Fb</i>) larvae are shown.</p

The effect of dE2F1b deregulation is mediated by cyclin E.

<p>(A) A schematic of different domains of RBF1 protein and the GST-fusion constructs used in the GST pull-down assay, “Large Pocket” (LP), “Small Pocket” (SP) and C-term are shown. These GST-fusion proteins are used to pull down HA-tagged dE2F1a or dE2F1b that are transiently expressed in S2 tissue culture cells. Anti-HA is used to visualize the transfected protein. A GST alone construct is used as a negative control and 10% of the input is used a loading control. (B) Myc-tagged dE2F1a or dE2F1b is expressed the Drosophila eye using an eye-specific GMR-Gal4 driver (GMRG4). Control eye discs are shown in upper panel. The effects of dE2F1a or dE2F1b overexpression alone (second panel), together with RBF1 (third panel), and together with cycERNAi (bottom panel) on the PCNA-GFP reporter activity and CycE expression are shown. (C) The adult eye morphology is visualized by the nail polish imprinting technique (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1007204#sec004" target="_blank">Materials and Methods</a>). dE2F1a or dE2F1b is either expressed alone (upper panel) or together with an RNAi construct targeting cycE (lower panel).</p

Oscillation of Cyclin E and E2F target gene expression is deregulated in <i>de2f1b</i> mutant salivary glands.

<p>(A) Salivary glands of control and <i>de2f1b</i> mutant early third instar larvae (80–85 hr AEL) are stained with anti-dE2F1 (green) and anti-Cyclin E (CycE, red). Bar graph indicates quantification of the percentages of CycE and dE2F1 expressing cells that co-express dE2F1 and CycE, respectively, in control and <i>de2f1b</i> mutant salivary glands. **** = p≤ 0.0001 using two-tailed t-test. (B) Salivary glands of control and <i>de2f1b</i> mutant early (80–85 hr AEL) third instar larvae expressing PCNA-GFP (green) are stained with anti-dE2F1 (red). The region where high PCNA-GFP is observed with low dE2F1 is marked by an asterisk. (C) Salivary glands of control and <i>de2f1b</i> mutant late third instar larvae (105–110 hr AEL) are stained with anti-Cyclin E (CycE). In addition, dE2F1 activity is monitored using PCNA-GFP, which is a GFP reporter of a well-known E2F target gene, PCNA. Cytoplasmic accumulation of dE2F1 in <i>de2f1b</i> mutants is marked by an arrowhead. (D) Relative expression levels of E2F target genes between control and <i>de2f1b</i> mutant late (105–110 hr AEL) third instar larval salivary glands are determined by RT-qPCR. Values represent the mean of triplicated biological replicates and error bars represent s.d. * = p ≤ 0.05; ** = p ≤ 0.01; **** = p≤ 0.0001 using two-tailed t-test. (E) Averages of the relative standard deviation of nuclear area from control, <i>de2f1b</i> mutant, and <i>de2f1b</i> mutant knocking down <i>cycE</i> using <i>heatshock-Gal4</i> and <i>UAS-cycE</i>^<i>RNAi</i>. Values represent the mean of triplicated biological replicates and error bars represent s.d. P values represent: ns = p>0.05; *** = p≤0.001; **** = p≤0.0001 calculated using one-way ANOVA. Scale bars for all salivary glands represent 50 μm. P: proximal end. D: distal end. FB: Fat body.</p

The expression pattern of dE2F1 and its target gene expression is altered in mitotic tissues of <i>de2f1b</i> mutants.

<p>(A) The expression patterns of dE2F1 (red) in third instar eye imaginal discs of control and <i>de2f1b</i> mutant larvae are shown. Eye absent (Eya, green), a nuclear protein, is also visualized in the same eye discs to control for the focal planes of the images. (B) PCNA-GFP activity and the expression pattern of an E2F target gene in third instar eye imaginal discs of the indicated genotypes are determined. For the E2F target gene, a <i>rnrS</i> antisense probe is used. (C) Eye imaginal discs of the indicated genotypes are labeled with EdU to visualize S-phase cells (EdU). CycE expression pattern was also determined by anti-CycE antibody. Yellow arrows indicate the position of the morphogenetic furrow. The asterisks indicate the location where ectopic S-phase cells and CycE expression were observed. (D) The expression patterns of dE2F1 and PCNA-GFP in third instar wing imaginal discs of the indicated genotypes are determined. Yellow arrowheads indicate the position of the zone of non-proliferating cell (ZNC) region.</p

<i>de2f1b</i>-specific mutant flies have reduced viability.

<p>(A) A schematic of the <i>de2f1</i> gene region shows the deletion of the 3b exon in <i>de2f1b</i> mutant flies. (B) RT-PCR confirms the lack of <i>de2f1b</i> transcript in the trans-heterozygous flies between <i>de2f1b</i> mutant and a deficiency line covering the <i>de2f1b</i> locus (<i>de2f1b/Df</i>). <i>yw</i> flies were used as control. (C) Quantification of the eclosion rate of the <i>de2f1b</i> mutants is shown. <i>de2f1b</i> homozygous (<i>de2f1b/de2f1b)</i> and trans-heterozygous over a deficiency (<i>de2f1b/Df</i>) flies are compared to a control (<i>yw</i>). The error bars indicate standard deviation (s.d.) of three independent experiments and the number of total pupae examined is indicated. (D) A control (<i>yw</i>) and <i>de2f1b</i> (<i>de2f1b/Df</i>) third instar larvae are shown on the left and their salivary glands are shown on the right. The scale bar for larvae is 1mm and for salivary glands is 0.5 mm. (E) Relative levels of the total <i>de2f1</i>, <i>de2f1b</i>, <i>CycE and rnrS</i> transcripts are determined by RT-qPCR. RNA samples are isolated from control (<i>yw</i>) and <i>de2f1b</i> mutants (<i>de2f1b</i> and <i>de2f1b/Df</i>) third instar larvae. The error bars indicate s.d. of triplicated biological replicates. Statistical testing in 2C and 2E were performed using two-tailed t-tests where ns = p>0.5; * = p ≤ 0.05; ** = p ≤ 0.01; *** = p ≤ 0.001; **** = p≤ 0.0001.</p

Highly Moldable Electrospun Clay-Like Fluffy Nanofibers for Three-Dimensional Scaffolds

The development of three-dimensional polymeric systems capable of mimicking the extracellular matrix is critical for advancing tissue engineering. To achieve these objectives, three-dimensional fibrous scaffolds with “clay”-like properties were successfully developed by coaxially electrospinning polystyrene (PS) and poly­(ε-caprolactone) (PCL) and selective leaching. As PS is known to be nonbiodegradable and vulnerable to mechanical stress, PS layers present at the outer surface were removed using a “selective leaching” process. The fibrous PCL scaffolds that remained after the leaching step exhibited highly advantageous characteristics as a tissue engineering scaffold, including moldability (i.e., clay-like), flexibility, and three-dimensional structure (i.e., cotton-like). More so, the “clay-like” PCL fibrous scaffolds could be shaped into any desired form, and the microenvironment within the clay scaffolds was highly favorable for cell expansion both in vitro and in vivo. These “electrospun-clay” scaffolds overcome the current limitations of conventional electrospun, sheet-like scaffolds, which are structurally inflexible. Therefore, this work extends the scope of electrospun fibrous scaffolds toward a variety of tissue engineering applications

Compositional Effects on Leaching of Stain-Guarded (Perfluoroalkyl and Polyfluoroalkyl Substance-Treated) Carpet in Landfill Leachate

Perfluoroalkyl and polyfluoroalkyl substances (PFASs) from stain-guard treated carpets in landfills continue to be released into the environment. To understand the leaching of PFASs from carpets to landfill leachate as a function of environmental factors, leaching concentrations of ten perfluoroalkyl carboxylic acids and four perfluoroalkyl sulfonic acids were quantified for different pHs, contact times, mixing speeds, and temperatures. Partitioning from carpet to leachate and distilled water at different pHs showed negligible differences. The total concentration of leaching PFASs in distilled water was approximately 1 ng L^–1 higher than in landfill leachate, indicating that the presence of multivalent cations in leachate could have a negative effect on leaching of PFASs. For all PFASs monitored, leaching increased with increasing contact time and temperature. Perfluorohexanoic and perfluoro­heptanoic acids experienced the largest increases with contact time and temperature. Gibbs free energy (Δ<i>G</i> > 0), enthalpy (Δ<i>H</i> > 0), and entropy energy (Δ<i>S</i> < 0) indicated that PFAS leaching from carpet was dominantly controlled by entropy-driven processes and did not differ significantly among individual PFASs. PFAS concentrations in leachate with rotation of an end-over-end contactor were higher than under static conditions, but otherwise, varying the rotation speed had negligible influence. The results provide useful information for management of discarded stain-guard carpets in landfills