DNA Condensation Research Articles

Targeted co-delivery of paclitaxel and anti P-gp shRNA by low molecular weight PEI decorated with L-3,4-dihydroxyphenylalanine.

Co-delivery of small chemotherapeutic molecules and nucleic acid materials via targeted carriers has attracted great attention for treatment of resistant tumors and reducing adverse effects. In this study, a targeted carrier for co-delivery was prepared based on low-molecular weight polyethylenimine (LMW PEI). Paclitaxel (PTX) was covalently conjugated onto PEI via a succinate linker. The PEI conjugate was decorated with L-DOPA in order to target large neutral amino acid transporter-1 (LAT-1) that is over-expressed on various cancer cells. This PEI conjugate was complexed with human ABCB1 shRNA plasmid to down-regulate the expression of P-glycoprotein, as one of the major efflux pumps inducing resistance against chemotherapeutics. The formation of PEI conjugate enhanced the solubility of PTX and resulted in the condensation and protection of plasmid DNA in nanosized polyplexes. The results of targeted delivery into the cells demonstrated that PEI conjugate transferred the payloads to the cells over-expressing LAT-1 transporter, while the biological effects on the cells lacking the transporter was negligible. Also, shRNA-mediated down-regulation of P-gp led to the increase of toxic effects on the cells over-expressing P-gp. This study suggests a promising approach for co-delivery of small molecules and nucleic acid materials in a targeted manner for cancer therapy.

A biophysical study of DNA condensation mediated by histones and protamines

The compaction of long DNA strands into confined spaces such as the nuclei of eukaryotic cells is an essential phenomenon towards the emergence of elaborated forms of life. Histones and protamines are the major nucleoproteins involved in this task participating in the formation of chromatin in somatic and germinative cells, respectively. In addition to a fundamental understanding of critical biological processes, DNA condensation also holds strong potential in biotechnology. Herein, we investigate the mesoscale structure of complexes formed between DNA and histones or protamines. A sophisticated set of biophysical methods encompassing steady-state fluorimetry, small-angle X-ray scattering and infrared nano spectroscopy was used to unveil both the self-assembly and molecular interactions of these complexes. We explored the fluorescence of a molecular rotor, thioflavin T, to investigate the accessibility of ligands in the inter-base environment of DNA strands. AFM-based infrared spectroscopy was used for the first time to probe the vibrational signature of individual DNA/nucleoprotein nano assemblies and disclose secondary-structure features. Our results show that protamines form highly compact structures in which DNA folding hinders access to the inter-base spacing. These assemblies exhibit diversified secondary-structure conformations, with the presence of β-sheets stabilizing the packing. In contrast, histone-based complexes are characterized by fibrillar nano assemblies exhibiting larger inter strands separations and access to guest molecules that intercalate between bases. The findings presented here may help the understanding of DNA condensation mediated by these two major nucleoproteins and may assist the optimization of gene vehicles based on these promising nano assemblies.

Steering DNA Condensation on Engineered Nanointerfaces.

DNA has received increasing attention in nanotechnology due to its ability to fold into prescribed structures. Different from the commonly adopted base-pairing strategy, an emerging class of amorphous DNA materials are formed by DNA's abiological interactions. Despite the great successes, a lack of nanoscale nucleation/growth control disables more advanced considerations. This work aims at harnessing the heterogeneous nucleation of metal-ion-glued DNA condensates on nanointerfaces. Upon unveiling key orthogonal factors including solution pH, ionic cross-linkers, and surface functionalities, chemically programmable DNA condensation on nanoparticle seeds is achieved, resembling a famous Stöber process for silica coating. The nucleation rules discovered on individual nanoseeds can be passed on to their dimeric assemblies, where broken spherical symmetry and the existence of interparticle gaps help a regiospecific DNA gelation. The steerable DNA condensation, and the multifunctions from DNA, metal ions, and nanocores, hold a great promise in noncanonical DNA nanotechnology toward novel applications.

Pillar[5]arene-induced DNA condensation: Liquid–liquid phase separation in pillar[5]arene-oligonucleotide system

The spatial separation of cell components and the simultaneous organization of many chemical reactions are important constituents of cell functioning. It is achieved through cellular compartments – membrane and membraneless organelles. Membraneless organelles have liquid-like properties and are formed during liquid–liquid phase separation. These compartments contain a complex of polyvalent molecules such as proteins, DNA and RNA. Here, we present a sufficiently simple two-component system for studying liquid–liquid phase separation, which includes a positively charged macrocyclic pillar[5]arene containing amide-ammonium fragments and a negatively charged DNA decamer. The investigation of proposed model systems with modifiable components can help understand the processes of self-assembly and phase separation.

Synthesis of Double Hydrophilic Block Copolymers Poly(2-isopropyl-2-oxazoline-b-ethylenimine) and their DNA Transfection Efficiency.

Gene delivery is now a part of the therapeutic arsenal for vaccination and treatments of inherited or acquired diseases. Polymers represent an opportunity to develop new synthetic vectors for gene transfer, with a prerequisite of improved delivery and reduced toxicity compared to existing polymers. Here, the synthesis in a two-step's procedure of linear poly(ethylenimine-b-2-isopropyl-2-oxazoline) block copolymers with the linear polyethylenimine (lPEI) block of various molar masses is reported; the molar mass of the poly(2-isopropyl-2-oxazoline) (PiPrOx) block has been set to 7kg mol-1 . Plasmid DNA condensation is successfully achieved, and in vitro transfection efficiency of the copolymers is at least comparable to that obtained with the lPEI of same molar mass. lPEI-b-PiPrOx block copolymers are however less cytotoxic than their linear counterparts. PiPrOx can be a good alternative to PEG which is often used in drug delivery systems. The grafting of histidine moieties on the lPEI block of lPEI-b-PiPrOx does not provide any real improvement of the transfection efficiency. A weak DNA condensation is observed, due to increased steric hindrance along the lPEI backbone. The low cytotoxicity of lPEI-b-PiPrOx makes this family a good candidate for future gene delivery developments.

RNA Captures More Cations than DNA: Insights from Molecular Dynamics Simulations.

The distribution of cations around nucleic acids is essential for a broad variety of processes ranging from DNA condensation and RNA folding to the detection of biomolecules in biosensors. Predicting the exact distribution of ions remains challenging since the distribution and, hence, a broad variety of nucleic acid properties depend on the salt concentration, the valency of the ions, and the ion type. Despite the importance, a general theory to quantify ion-specific effects for highly charged biomolecules is still lacking. Moreover, recent experiments reveal that despite their similar building blocks, DNA and RNA duplexes can react differently to the same ionic conditions. The aim of our current work is to provide a comprehensive set of molecular dynamics simulations using more than 180 μs of simulation time. For the mono- and divalent cations Li+, Na+, K+, Cs+, Ca2+, Sr2+, and Ba2+, the simulations allow us to reveal the ion-specific distributions and binding patterns for DNA and RNA duplexes. The microscopic insights from the simulations display the origin of ion-specificity and shed light on the question of why DNA and RNA show opposing behavior in the same ionic conditions. Finally, the detailed binding patterns from the simulations reveal why RNA can capture more cations than DNA.

Effect of phosphorylation of protamine-like cationic peptide on the binding affinity to DNA

Protamines are more arginine-rich and more basic than histones and are responsible for providing a highly compacted shape to the sperm heads in the testis. Phosphorylation and dephosphorylation are two events that occur in the late phase of spermatogenesis before the maturation of sperms. In this work, we have studied the effect of phosphorylation of protamine-like cationic peptides using all-atom molecular dynamics simulations. Through thermodynamic analyses, we found that phosphorylation reduces the binding efficiency of such cationic peptides on DNA duplexes. Peptide phosphorylation leads to a less efficient DNA condensation, due to a competition between DNA-peptide and peptide-peptide interactions. We hypothesize that the decrease of peptide bonds between DNA together with peptide self-assembly might allow an optimal re-organization of chromatin and an efficient condensation through subsequent peptide dephosphorylation. Based on the globular and compact conformations of phosphorylated peptides mediated by arginine-phosphoserine H-bonding, we furthermore postulate that phosphorylated protamines could more easily intrude into chromatin and participate to histone release through disruption of histone-histone and histone-DNA binding during spermatogenesis.

The Hydrophobicity of AIE Dye Facilitates DNA Condensation for Carrier‐Free Gene Therapy

AbstractA nanomaterial with a high payload of nucleic acid drugs and low cytotoxicity is desirable for gene therapy. Herein, a new DNA assembly method triggered by hydrophobic interaction and strengthened by Mg2+ electrostatic interaction for carrier‐free gene transfection is reported. It is discovered that a hydrophobic aggregation‐induced emission dye with a low molecular weight that is modified to an oligonucleotide will greatly change the condensation properties of DNA. The conditions and the possible mechanism of such an aggregation process are carefully investigated. Further, it is revealed that the resultant aggregate is suitable for gene delivery, as in which the oligonucleotides are protected from degradation, and the intracellular permeability is enhanced. For the delivery of Bcl‐2 antisense oligonucleotide, such a system exhibits a high gene transfection efficiency both in vitro and in vivo. Accordingly, an interesting observation is reported in this study that an aggregation‐induced emission dye with a molecular weight much lower than the nucleic acid drugs can cause oligonucleotide aggregation and facilitate gene transfection. Such a delivery system provides a practical and inspiring example for the development of gene therapy in the future.

Ku proteins promote DNA binding and condensation of cyclic GMP-AMP synthase.

Cyclic GMP-AMP synthase (cGAS) is a cytosolic DNA sensor that plays a critical role in regulating antiviral signaling. cGAS binds to DNA and catalyzes the synthesis of cyclic GMP-AMP (cGAMP), which is essential for downstream signal transduction. The antiviral response is a rapid biological process; however, cGAS itself has relatively low DNA binding affinity, implying that formation of the cGAS-DNA complex requires an additional factor(s) that promotes cGAS-DNA binding, allowing efficient antiviral signal transduction. Here, we report that the Ku proteins (Ku80 and Ku70) directly interact with cGAS and positively regulate cGAS-mediated antiviral signaling. Mechanistically, we find that the interaction of the Ku proteins with cGAS significantly increases the DNA-binding affinity of cGAS and promotes cGAS condensation in the cytosol, thereby enhancing cGAS catalytic activity. Our results show that the Ku proteins are critical partners of cGAS in sensing DNA virus infection and ensuring efficient innate immune signal transduction.

Structural, enzymatic and spatiotemporal regulation of PP2A-B55 phosphatase in the control of mitosis.

Cells require major physical changes to induce a proper repartition of the DNA. Nuclear envelope breakdown, DNA condensation and spindle formation are promoted at mitotic entry by massive protein phosphorylation and reversed at mitotic exit by the timely and ordered dephosphorylation of mitotic substrates. This phosphorylation results from the balance between the activity of kinases and phosphatases. The role of kinases in the control of mitosis has been largely studied, however, the impact of phosphatases has long been underestimated. Recent data have now established that the regulation of phosphatases is crucial to confer timely and ordered cellular events required for cell division. One major phosphatase involved in this process is the phosphatase holoenzyme PP2A-B55. This review will be focused in the latest structural, biochemical and enzymatic insights provided for PP2A-B55 phosphatase as well as its regulators and mechanisms of action.

The Growth Rate of DNA Condensate Droplets Increases with the Size of Participating Subunits.

Liquid-liquid phase separation (LLPS) is a common phenomenon underlying the formation of dynamic membraneless organelles in biological cells, which are emerging as major players in controlling cellular functions and health. The bottom-up synthesis of biomolecular liquid systems with simple constituents, like nucleic acids and peptides, is useful to understand LLPS in nature as well as to develop programmable means to build new amorphous materials with properties matching or surpassing those observed in natural condensates. In particular, understanding which parameters determine condensate growth kinetics is essential for the synthesis of condensates with the capacity for active, dynamic behaviors. Here we use DNA nanotechnology to study artificial liquid condensates through programmable star-shaped subunits, focusing on the effects of changing subunit size. First, we show that LLPS is achieved in a 6-fold range of subunit size. Second, we demonstrate that the rate of growth of condensate droplets scales with subunit size. Our investigation is supported by a general model that describes how coarsening and coalescence are expected to scale with subunit size under ideal assumptions. Beyond suggesting a route toward achieving control of LLPS kinetics via design of subunit size in synthetic liquids, our work suggests that particle size may be a key parameter in biological condensation processes.

Condensation and Protection of DNA by the Myxococcus xanthus Encapsulin: A Novel Function.

Encapsulins are protein nanocages capable of harboring smaller proteins (cargo proteins) within their cavity. The function of the encapsulin systems is related to the encapsulated cargo proteins. The Myxococcus xanthus encapsulin (EncA) naturally encapsulates ferritin-like proteins EncB and EncC as cargo, resulting in a large iron storage nanocompartment, able to accommodate up to 30,000 iron atoms per shell. In the present manuscript we describe the binding and protection of circular double stranded DNA (pUC19) by EncA using electrophoretic mobility shift assays (EMSA), atomic force microscopy (AFM), and DNase protection assays. EncA binds pUC19 with an apparent dissociation constant of 0.3 ± 0.1 µM and a Hill coefficient of 1.4 ± 0.1, while EncC alone showed no interaction with DNA. Accordingly, the EncAC complex displayed a similar DNA binding capacity as the EncA protein. The data suggest that initially, EncA converts the plasmid DNA from a supercoiled to a more relaxed form with a beads-on-a-string morphology. At higher concentrations, EncA self-aggregates, condensing the DNA. This process physically protects DNA from enzymatic digestion by DNase I. The secondary structure and thermal stability of EncA and the EncA−pUC19 complex were evaluated using synchrotron radiation circular dichroism (SRCD) spectroscopy. The overall secondary structure of EncA is maintained upon interaction with pUC19 while the melting temperature of the protein (Tm) slightly increased from 76 ± 1 °C to 79 ± 1 °C. Our work reports, for the first time, the in vitro capacity of an encapsulin shell to interact and protect plasmid DNA similarly to other protein nanocages that may be relevant in vivo.

Chromatin sequesters pioneer transcription factor Sox2 from exerting force on DNA

Biomolecular condensation constitutes an emerging mechanism for transcriptional regulation. Recent studies suggest that the co-condensation between transcription factors (TFs) and DNA can generate mechanical forces driving genome rearrangements. However, the reported forces generated by protein-DNA co-condensation are typically below one piconewton (pN), questioning its physiological significance. Moreover, the force-generating capacity of these condensates in the chromatin context remains unknown. Here, we show that Sox2, a nucleosome-binding pioneer TF, forms co-condensates with DNA and generates forces up to 7 pN, exerting considerable mechanical tension on DNA strands. We find that the disordered domains of Sox2 are required for maximum force generation but not for condensate formation. Furthermore, we show that nucleosomes dramatically attenuate the mechanical stress exerted by Sox2 by sequestering it from coalescing on bare DNA. Our findings reveal that TF-mediated DNA condensation can exert significant mechanical stress on the genome which can nonetheless be attenuated by the chromatin architecture.

The Activation of Protamine 1 Using Epigenome Editing Decreases the Proliferation of Tumorigenic Cells.

DNA methyltransferases (DNMT) and histone deacetylases (HDAC) inhibitors are used as cancer epigenome drugs. However, these epigenetic drugs lack targeting specificity and could risk inducing genome instability and the expression of oncogenes. Therefore, there is a need to develop new therapeutic strategies where specific cancer genes can be targeted for silencing or activation. The CRISPR/dCas9 system represents a promising, powerful therapeutic tool because of its simplicity and specificity. Protamine 1 (PRM1) is exclusively expressed in sperm and has a vital role in the tight packaging of DNA, thus inducing transcriptional silencing in sperm cells. We hypothesized that the activation of the PRM1 gene in tumorigenic cells would lead to DNA condensation and reduce the proliferation of these cells. To test our hypothesis, we transfected human embryonic kidney cells 293T with a dCas9-P300 plasmid that adds acetyl groups to the promoter region of PRM1 via specific gRNAs plasmids. RNA-Seq analysis of transfected cells revealed high specificity of targeted gene activation. PRM1 expression resulted in a significant decrease in cell proliferation as measured by the BrdU ELISA assay. To confirm that the activation of PRM1 was due to acetyl groups deposited to H3K27, a ChIP-qPCR was performed. The acetylation of the PRM1 promoter region targeted by dCas9-p300 in transfected cells was higher than that of the control cells. Interestingly, the targeted promoter region for acetylation showed reduced DNA methylation. These findings demonstrate the efficacy of epigenome editing in activating PRM1 in non-expressing tumorigenic cells, which could be used as a promising therapeutic strategy in cancer treatment.

The role of packaging signals in virus assembly and the interplay between the nucleation and elongation rates

Virion assembly and genome packaging are among the most interesting biophysical phenomena in biology. However, these processes are not universal and can be very different, even for closely related viruses. The details for these phenomena depend on the type of genome, the assembly site in the host cell, the replication cycle, and the mechanism for selective packaging of the viral genome (1). The study of virion assembly and genome packaging has led to the understanding of exciting phenomena, such as DNA condensation and its role in pressurized viruses (2,3), the role of electrostatics in the RNA packaging for single-stranded RNA viruses (4), the dependence of the assembly pathway to the relative strength of RNA-protein and protein-protein interactions (5), assembly kinetics (6), and the requirement of packaging signals (7).

Essential oil of Foeniculumvulgare subsp. piperitum fruits exerts an anti‑tumor effect in triple‑negative breast cancer cells.

At present, the growing spread of tumor cases worldwide renders the research of new promising and selective anticancer drugs urgent. The biological action of extracts of medicinal plants or their essential oils (EOs) is an emerging field of interest, since they could comprise a rich source of phytochemicals that can prove promising. In the present study, the biological activity and mechanism of action of the EO of Foeniculumvulgare subsp. piperitum fruits (FVPEO) were investigated using MTT assays, morphological analyses and western blotting in MDA‑MB231 cells, a triple‑negative breast cancer cell line. The findings revealed that FVPEO could exert strong anticancer effects, causing a dose‑dependent inhibition of breast cancer MDA‑MB231 cell growth, accompanied with DNA condensation and fragmentation. The cytotoxic effect of FVPEO was counteracted by the addition of the antioxidant N‑acetylcysteine and was associated with a marked increase in reactive oxygen species and stress‑related proteins; such as manganese superoxide dismutase, c‑Jun, phospho‑c‑Jun N‑terminal kinase and nuclear factor E2‑related factor 2, and the latter's transcriptional targets, Heme oxygenase‑1 and NAD(P)H quinone oxidoreductase 1 (NQO1). As evidenced by the activation of caspase‑3 and fragmentation of poly(ADP‑ribose) polymerase‑1, which are typical apoptosis markers, FVPEO promoted apoptotic cell death accompanied with an increase in phosphorylated H2A histone family member X and the activation of the NQO1/p53 axis. In combination, the present experiments provided evidence that FVPEO could represent a reservoir of biologically active compounds suitable for both cancer prevention and treatment.

An insight into the morphology of DNA compaction induced by homobinuclear Ru(II) polypyridyl complexes

Binuclear Ru(II) polypyridyl complexes [Ru2(NN)4(BIPMB)]4+ (1–4), where N-N = 2,2′-bipyridine (bpy), 1,10-phenanthroline (phen), dipyrido [3,2-d:2′,3′-f] quinoxaline (dpq), and dipyrido[3,2-a:2′,3′-c]phenazine (dppz), have been synthesized using suitable precursors and bridging ligand (BIPMB), where BIPMB = 3,3′-bis-{(imidazol-1-yl)-[4,5-f]-1,10-phenanthroline) methyl}-1,1′-biphenyl. The binding mode and affinity of complexes 1–4 with Calf Thymus DNA (CT-DNA) were determined by absorption and steady-state fluorescence spectroscopy. The decrease in viscosity of CT-DNA on sequential addition of these complexes indicated DNA condensation and the result was corroborated by circular dichroism (CD). The nanosized globular aggregates of CT-DNA induced by complexes 1–4 were observed by dynamic light scattering (DLS) and transmission electron microscopy (TEM) measurements. The gel electrophoretic mobility studies revealed the small orderly particles of supercoiled plasmid pBR322 DNA induced by these complexes. Additionally, the morphology and size of compact plasmid DNA condensates were studied by DLS and atomic force microscopy (AFM). The complexes were moderately cytotoxic against MCF-7 cells.

Photoprotective effects of Sargassum thunbergii on ultraviolet B-induced mouse L929 fibroblasts and zebrafish

BackgroundChronic exposure to ultraviolet B (UVB) causes a series of adverse skin reactions, such as erythema, sunburn, photoaging, and cancer, by altering signaling pathways related to inflammation, oxidative stress, and DNA damage. Marine algae have abundant amounts and varieties of bioactive compounds that possess antioxidant and anti-inflammatory properties. Thus, the objective of this study was to investigate the photoprotective effects of an ethanol extract of Sargassum thunbergii.MethodsSargassum thunbergii phenolic-rich extract (STPE) was prepared, and its activity against UVB damage was evaluated using L929 fibroblast cells and zebrafish. STPE was extracted and purified by 40% ethanol and macroporous resin XDA-7. Reactive oxygen species (ROS) and antioxidant markers, such as superoxide dismutase (SOD), catalase (CAT) activities, and malondialdehyde (MDA) content were analyzed. The effect of STPE on UVB-induced inflammation was determined by inflammatory cytokine gene and protein expression. The expression of signaling molecules in the Nuclear Factor KappaB (NF-κB) pathway was determined by western blotting. DNA condensation was analyzed and visualized by Hoechst 33342 staining. In vivo evaluation was performed by tail fin area and ROS measurement using the zebrafish model.ResultsThe total polyphenol content of STPE was 72%. STPE reduced ROS content in L929 cells, improved SOD and CAT activities, and significantly reduced MDA content, thereby effectively alleviating UVB radiation-induced oxidative damage. STPE inhibited the mRNA and protein expression of TNF-α, IL-6, and IL-1α. STPE reversed DNA condensation at concentrations of 20 and 40 μg/mL compared with the UVB control. Moreover, STPE inhibited NF-κB signaling pathway activation and alleviated DNA agglutination in L929 cells after UVB irradiation. Additionally, 1.67 μg/mL STPE significantly increased the tail fin area in zebrafish, and 0.8–1.6 μg/mL STPE effectively eliminated excessive ROS after UVB radiation.ConclusionsSTPE inhibited UVB-induced oxidative stress, inflammatory cytokine expression, and DNA condensation via the downregulation of the NF-κB signaling pathway, suggesting that it prevents UVB-induced photodamage, and has potential for clinical development for skin disease treatment.

Aquaporin 1 Modulates Apoptosis Susceptibility in Pulmonary Hypertension

Pulmonary arterial hypertension (PAH) remains a fatal diagnosis despite available therapies. PAH is characterized by extensive remodeling of the pulmonary vasculature involving the formation of vaso‐occlusive lesions and a thickened medial layer of the vascular wall, both of which contain pulmonary arterial smooth muscle cells (PASMCs). We have demonstrated that PASMCs isolated from a well‐established rat model of PAH are resistant to apoptosis under both basal and stimulated conditions. The cell membrane protein aquaporin 1 (AQP1) was initially described as a water transport channel, but more recently has been implicated in other cellular functions including migration and proliferation, and in several distinct cancer types, has been associated with apoptosis resistance. AQP1 is upregulated in PASMCs isolated from rat models of PAH, although its role in apoptosis resistance is unclear. We hypothesized that increased expression of AQP1 contributes to apoptosis resistance in PASMCs.MethodsOur animal model is the Sugen‐hypoxia (SuHx) rat model of PAH in which Sugen‐5416, a vascular endothelial growth factor receptor type 2 inhibitor is injected into rats before they are placed in chronic hypoxia (10% O2) for 3 weeks, then returned to normoxia (21% O2) for 2 weeks. Control animals receive vehicle injection and are kept in normoxic (21% O2) conditions. Cells from distal pulmonary arteries are obtained via removal of the endothelium and enzymatic digestion of the tissue to obtain a pure smooth muscle population. To deplete AQP1, we transfected control and SuHx PASMCs with small interfering RNA (siRNA) directed toward either the AQP1 protein (siAQP1) or a non‐targeted sequence (siNT). Apoptosis was measured under basal conditions by Hoechst staining and visualization of DNA condensation. In complementary experiments, control PASMCs were infected with adenoviral constructs to over‐express AQP1 (AdAQP1) or green fluorescent protein (AdGFP; infection control). Cells were treated with either PBS or the apoptotic stimulus hydrogen peroxide (250 µm for 24 h). Apoptosis was again measured by Hoechst staining and visualization of DNA condensation.ResultsIn both control and SuHx PASMCs, AQP1 knock down increased apoptosis under basal conditions. In control PASMCs with increased AQP1 expression, there was no difference in apoptosis under basal conditions. To further provoke a response in these infected cells, the apoptotic stimulus hydrogen peroxide was applied; however, again there was no difference in apoptosis levels between AdAQP1 and AdGFP infected cells.ConclusionsThese data suggest that increased AQP1 may be necessary but not sufficient for apoptosis resistance. Future studies further characterizing the role of AQP1 in apoptosis resistance are necessary to understand molecular mechanisms governing resistance to apoptosis in PASMCs and provide novel pathways for new therapeutic agents targeting vascular remodeling with the potential to reverse disease.

Spontaneous Condensation of RNA into Nanoring and Globular Structures.

The effect of polyvalent cations, like spermine, on the condensation of DNA into very well-defined toroidal shapes has been well studied and understood. A great effort has been made to obtain similar condensed structures from RNA molecules, but so far, it has been elusive. In this work, we show that single-stranded RNA (ssRNA) molecules can easily be condensed into nanoring and globular structures on a mica surface, where each nanoring structure is formed mostly by a single RNA molecule. The condensation occurs in a concentration range of different cations, from monovalent to trivalent, but at a higher concentration, globular structures appear. RNA nanoring structures were observed on mica surfaces by atomic force microscopy (AFM). The samples were observed in tapping mode and were prepared by drop evaporation of a solution of RNA in the presence of one type of the different cations used. As far as we know, this is the first time that nanorings or any other well-defined condensed RNA structures have been reported in the presence of simple salts. The RNA nanoring formation can be understood by an energy competition between the hydrogen bonding forming hairpin stems—weakened by the salts—and the hairpin loops. This result may have an important biological relevance since it has been proposed that RNA is the oldest genome-coding molecule, and the formation of these structures could have given it stability against degradation in primeval times. Even more, the nanoring structures could have the potential to be used as biosensors and functionalized nanodevices.