Elucidating cell-penetrating peptide mechanisms of action for membrane interaction,cellular uptake,and translocation utilizing the hydrophobic counter-anion pyrenebutyrate

Cell-penetrating peptides (CPPs) are membrane permeable vectors recognized for their intrinsic ability to gain access to the cell interior. The hydrophobic counter-anion, pyrenebutyrate, enhances cellular uptake of oligoarginine CPPs. To elucidate CPP uptake mechanisms, the effect of pyrenebutyrate on well-recognized CPPs with varying hydrophobicity and arginine content is investigated. The cellular CPP uptake and CPP-mediated oligonucleotide delivery is analyzed by fluorescence activated cell sorting, confocal microscopy, and a cell-based splice-switching assay. The splice-switching oligonucleotide is a mixmer of 2′-O-methyl RNA and locked nucleic acids delivered as a non-covalent complex with 10-fold molar CPP excess. CPP-induced membrane perturbation on large unilamellar vesicles is investigated in calcein release experiments. We observed that pyrenebutyrate facilitates cellular uptake and translocation of oligonucleotide mediated by oligoarginine nonamer while limited effect of pyrenebutyrate on more hydrophobic CPPs was observed. By combining the different experimental results we conclude that the pathway for cellular uptake of oligoarginine is dominated by direct membrane translocation, whereas the pathway for oligoarginine-mediated oligonucleotide translocation is dominated by endocytosis. Both mechanisms are promoted by pyrenebutyrate and we suggest that pyrenebutyrate has different sites of action for the two uptake and translocation mechanisms.     

Impact of mineral fertilizers on mineral nutrients in the ginger rhizome and on soil enzymes activities and soil properties

Ginger is used as one of the important ingredients in traditional as well as modern medicine besides as a spice. It boosts immunity and is a rich source of many biologically active substances and minerals. Although it is a medicinally important crop, its productivity is, however, affected due to poor nutrient management and therefore it requires an adequate supply of nutrients in the form of inorganic fertilizers or organic manuring, or a mixture of both. In this context, the present study was aimed to investigate the effect of mineral fertilizers on the content of mineral elements in the ginger rhizome, on soil enzyme activity, and soil properties. Lysimeter experiments were conducted at the Institute of Genetics and Plant Experimental Biology, Kibray, Tashkent region, Uzbekistan. The experiment comprised of four treatments T1 – Control, T2 - N₇₅P₅₀K₅₀ kg/ha, T3 - and T4 - N₁₀₀P₇₅K₇₅ + B₃Zn₆Fe₆ kg/ha. The results showed that the application of N₁₂₅P₁₀₀K₁₀₀ kg/ha increased rhizome K content by 49%, P content by 20%, and Na content by 58% as compared to control without fertilizer. While the application of N₁₀₀P₇₅K₇₅ + B₃Zn₆Fe₆ kg/ha showed a significant enhancement in rhizome K, Ca, P, Mg, Na, Fe, Mn, Zn, Cu, Cr, Mo, and Si contents over the control. This treatment also improved active P content by 29%, total P content by 80%, total K content 16%, and N content by 33% content, and the activities of urease, invertase, and catalase activities as compared to control of without mineral fertilizer and control respectively. Thus the application of NPK + BZnFe at the rate of 100:75:75:3:6:6 kg/ha helps in improving macroelements and microelements in the ginger rhizome and activities of soil enzymes that helps in mineral nutrition of the rhizome.     

The majority of the genetic risk for Paget’s disease of bone is explained by genetic variants close to the CSF1, OPTN, TM7SF4, and TNFRSF11A genes

Paget’s disease of bone (PDB) is one of the most frequent metabolic bone disorders (1–5%), next to osteoporosis, affecting individuals above age 55. Sequestosome1 mutations explain a part of the PDB patients, but still the disease pathogenesis in the remaining PDB patients is largely unknown. Therefore, association studies investigating the relationship between genetic polymorphisms and sporadic PDB have been performed to find the genetic risk variants. Previously such studies indicated a role of the OPG and RANK gene. The latter was recently confirmed in a genome-wide association study (GWAS) which also indicated the involvement of chromosomal regions harbouring the CSF1 and OPTN gene. In this study, we sought to replicate these findings in a Belgian and a Dutch population. Similar significant results were obtained for the single nucleotide polymorphisms and the haplotypes. The most significant results are found in the CSF1 gene region, followed by the OPTN and TNFRSF11A gene region (p values ranging from 1.3 × 10^?4 to 3.8 × 10^?8, OR = 1.523–1.858). We next obtained significant association with a polymorphism from the chromosomal region around the TM7SF4 gene (p = 2.7 × 10^?3, OR = 1.427), encoding DC-STAMP which did not reach genome-wide significance in the GWAS, but based on its function in osteoclasts it can be considered a strong candidate gene. After meta-analysis with the GWAS data, p values ranged between 2.6 × 10^?4 and 8.8 × 10^?32. The calculated cumulative population attributable risk of these four loci turned out to be about 67% in our two populations, indicating that most of the genetic risk for PDB is coming from genetic variants close to these four genes.     

Leaf temperature of desert sand dune plants: perspectives on the adaptability of leaf morphology

The leaf temperature of six annual and six perennial plant species was monitored during spring and summer on a sand dune ecosystem in the delta Mediterranean coast of Egypt. During winter, leaves of all tested perennial species attained temperatures higher than the air temperature at night and shortly after sunrise, with maximum leaf–air temperature differences reaching up to 8°C. The lowest differences were less than 1°C. Around noon, the leaves of several species attained temperatures lower than that of the air whereas others showed higher temperatures. The opposite was true during summer, when leaf temperatures were lower than air temperature. The maximum leaf–air temperature differences occurred after midnight towards sunrise and reached up to 10°C. The lowest differences were found around noon and were of less than 5°C. The annual plant species have more pronounced variations than perennials in their leaf temperatures during the night and for most of the day. The leaves were heated or cooled a few degrees above or below the air temperature. The results are discussed in relation to the morphological characters of the leaves. The variation in leaf temperature at different times of the day was significantly related to leaf morphology, specific leaf area, thickness, volume, leaf area index and the surrounding environment.     

Congenital diaphragmatic hernia and chromosome 15q26: determination of a candidate region by use of fluorescent in situ hybridization and array-based comparative genomic hybridization

Congenital diaphragmatic hernia (CDH) has an incidence of 1 in 3,000 births and a high mortality rate (33%-58%). Multifactorial inheritance, teratogenic agents, and genetic abnormalities have all been suggested as possible etiologic factors. To define candidate regions for CDH, we analyzed cytogenetic data collected on 200 CDH cases, of which 7% and 5% showed numerical and structural abnormalities, respectively. This study focused on the most frequent structural anomaly found: a deletion on chromosome 15q. We analyzed material from three of our patients and from four previously published patients with CDH and a 15q deletion. By using array-based comparative genomic hybridization and fluorescent in situ hybridization to determine the boundaries of the deletions and by including data from two individuals with terminal 15q deletions but without CDH, we were able to exclude a substantial portion of the telomeric region from the genetic etiology of this disorder. Moreover, one patient with CDH harbored a small interstitial deletion. Together, these findings allowed us to define a minimal deletion region of approximately 5 Mb at chromosome 15q26.1-26.2. The region contains four known genes, of which two--NR2F2 and CHD2--are particularly intriguing gene candidates for CDH.     

Histochemical Study of Photoperiodic and Low Temperature Effects on Floral Induction of Spinacia oleracea

Shoot apices of Spinacia oleracea plants have been induced toflower either by: (a) subjecting leaves to 24 h long day, or(b) exposure to a short photoperiod but displaced by 8 h (displacedshort day) in the usual 24 h short-day cycle, or (c) exposureto low temperature (5 °C) during the dark period of thenormal short day. A quantitative cytochemical assay of pentosephosphate pathway activity during floral induction indicatesan approximate doubling of the rate of activity when comparedto that of vegetative apices (short day) (21 °C). Exposure to either low temperature, or a displaced short photoperiodstimulates pentose phosphate pathway activity in the shoot apexin a manner similar to that seen by long-day induction. Thischange in metabolic activity is accompanied by changes in theshape of the shoot apex which resembles that seen at an earlystage during floral induction. Spinacia oleracea, pentose phosphate pathway, shoot apex, glucose-6-phosphate dehydrogenase, floral induction, chilling, displaced short day