MYC phase separation selectively modulates the transcriptome.

Dysregulation and enhanced expression of MYC transcription factors (TFs) including MYC and MYCN contribute to the majority of human cancers. For example, MYCN is amplified up to several hundredfold in high-risk neuroblastoma. The resulting overexpression of N-myc aberrantly activates genes that are not activated at low N-myc levels and drives cell proliferation. Whether increasing N-myc levels simply mediates binding to lower-affinity binding sites in the genome or fundamentally changes the activation process remains unclear. One such activation mechanism that could become important above threshold levels of N-myc is the formation of aberrant transcriptional condensates through phase separation. Phase separation has recently been linked to transcriptional regulation, but the extent to which it contributes to gene activation remains an open question. Here we characterized the phase behavior of N-myc and showed that it can form dynamic condensates that have transcriptional hallmarks. We tested the role of phase separation in N-myc-regulated transcription by using a chemogenetic tool that allowed us to compare non-phase-separated and phase-separated conditions at equivalent N-myc levels, both of which showed a strong impact on gene expression compared to no N-myc expression. Interestingly, we discovered that only a small percentage (<3%) of N-myc-regulated genes is further modulated by phase separation but that these events include the activation of key oncogenes and the repression of tumor suppressors. Indeed, phase separation increases cell proliferation, corroborating the biological effects of the transcriptional changes. However, our results also show that >97% of N-myc-regulated genes are not affected by N-myc phase separation, demonstrating that soluble complexes of TFs with the transcriptional machinery are sufficient to activate transcription.

Poly(ethylene succinate-co-lactic acid) as a Multifunctional Additive for Modulating the Miscibility, Crystallization, and Mechanical Properties of Poly(lactic acid).

Polymer blending offers an effective and economical approach to overcome the performance limitations of poly(lactic acid) (PLA). In this study, a series of copolymers poly(ethylene succinate-co-lactic acid) (PESL) were synthesized, featuring lactic acid (LA) contents that ranged from 20 to 86 wt %. This synthesis involved a one-pot industrial melt polycondensation process using succinic acid (SA), ethylene glycol (EG), and LA, catalyzed by titanium tetraisopropoxide (TTP). The goal was to produce a fully biobased copolymer expected to exhibit partial miscibility with pure poly(lactic acid) (PLA). To assess the capability of PESL copolymers in toughening PLA, we conducted tensile testing on PLA/PESL blends containing 15 wt % PESL. As a result, an elongation at break for the blends with 15 wt % loading of the copolymer PESL72 was directly enhanced to 250% with an ultimate strength of 35 MPa, compared to brittle PLA with less 10% tensile length. The morphological features of interfacial adhesion before and after tensile failure were measured by scanning electron microscopy (SEM). A significant enhancement in the chain mobility of the PLA/PESL blends was further evidenced by differential scanning calorimetry (DSC) and dynamic mechanical analysis (DMA). These findings hold promise for the development of functional packaging materials based on PLA. The proposed copolymer design, which boasts strong industrial feasibility, can serve as a valuable guide for enhancing the toughness of PLA.

Phase separation of YAP-MAML2 differentially regulates the transcriptome.

Phase separation (PS) drives the formation of biomolecular condensates that are emerging biological structures involved in diverse cellular processes. Recent studies have unveiled PS-induced formation of several transcriptional factor (TF) condensates that are transcriptionally active, but how strongly PS promotes gene activation remains unclear. Here, we show that the oncogenic TF fusion Yes-associated protein 1-Mastermind like transcriptional coactivator 2 (YAP-MAML2) undergoes PS and forms liquid-like condensates that bear the hallmarks of transcriptional activity. Furthermore, we examined the contribution of PS to YAP-MAML2-mediated gene expression by developing a chemogenetic tool that dissolves TF condensates, allowing us to compare phase-separated and non-phase-separated conditions at identical YAP-MAML2 protein levels. We found that a small fraction of YAP-MAML2-regulated genes is further affected by PS, which include the canonical YAP target genes CTGF and CYR61, and other oncogenes. On the other hand, majority of YAP-MAML2-regulated genes are not affected by PS, highlighting that transcription can be activated effectively by diffuse complexes of TFs with the transcriptional machinery. Our work opens new directions in understanding the role of PS in selective modulation of gene expression, suggesting differential roles of PS in biological processes.

Verification of Reinforced Surface Loose Layer of Zinc-Aluminum-Magnesium Steel Plate.

The corrosion resistance of zinc-aluminum-magnesium steel plates (Zn-Al-Mg steel plates) is significantly higher than that of galvanized steel plates. However, the unsatisfactory bonding performance of Zn-Al-Mg steel plates significantly limits their widespread application. In this study, X-ray photoelectron spectroscopy is employed to detect changes in the surface oxygen content of Zn-Al-Mg steel plates after different temperature treatments to confirm the existence of surface loose layers. In particular, changes in the surface oxygen content of the Zn-Al-Mg steel plates after the oxide layer is removed are investigated under saturated H2O vapor and O2 environmental conditions, and the cause of the formation of loose surface layers is determined. The uneven distribution of elements on the surface of the Zn-Al-Mg steel plates is investigated with scanning electron microscopy and energy dispersive spectroscopy. Nuclear magnetic resonance is employed to determine the size of the network spatial structure formed by silane coupling agents under different hydrolysis conditions and to further investigate the bonding performance of hydrolysate-modified Zn-Al-Mg steel plates. Several typical automotive adhesives are utilized to compare and examine the changes in the tensile strength of the Zn-Al-Mg steel plate bonding before and after modification with the silane coupling agent and analyze the structural damage of the adhesive at the bonding interface. The results confirm that the silane coupling agent strengthens the loose layer on the surface of the Zn-Al-Mg steel plate.

ATM-SPARK: A GFP phase separation-based activity reporter of ATM.

The kinase ataxia telangiectasia mutated (ATM) plays a key role in the DNA damage response (DDR). It is thus essential to visualize spatiotemporal dynamics of ATM activity during DDR. Here, we designed a robust ATM activity reporter based on phosphorylation-inducible green fluorescent protein phase separation, dubbed ATM-SPARK (separation of phases-based activity reporter of kinase). Upon ATM activation, it undergoes phase separation via multivalent interactions, forming intensely bright droplets. The reporter visualizes spatiotemporal dynamics of endogenous ATM activity in living cells, and its signal is proportional to the amount of DNA damage. ATM-SPARK also enables high-throughput screening of biological and small-molecule regulators. We identified the protein phosphatase 4 that blocks ATM activity. We also identified BGT226 as a potent ATM inhibitor with a median inhibitory concentration of ~3.8 nanomolars. Furthermore, BGT226 sensitizes cancer cells to the radiomimetic drug neocarzinostatin, suggesting that BGT226 might be combined with radiotherapeutic treatment. ATM-SPARK achieves large dynamic range, bright fluorescence, and simple signal pattern.

Designed peptides that assemble into cross-α amyloid-like structures.

Amyloids adopt 'cross-ß' structures composed of long, twisted fibrils with ß-strands running perpendicular to the fibril axis. Recently, a toxic peptide was proposed to form amyloid-like cross-α structures in solution, with a planar bilayer-like assembly observed in the crystal structure. Here we crystallographically characterize designed peptides that assemble into spiraling cross-α amyloid-like structures, which resemble twisted ß-amyloid fibrils. The peptides form helical dimers, stabilized by packing of small and apolar residues, and the dimers further assemble into cross-α amyloid-like fibrils with superhelical pitches ranging from 170 Å to 200 Å. When a small residue that appeared critical for packing was converted to leucine, it resulted in structural rearrangement to a helical polymer. Fluorescently tagged versions of the designed peptides form puncta in mammalian cells, which recover from photobleaching with markedly different kinetics. These structural folds could be potentially useful for directing in vivo protein assemblies with predetermined spacing and stabilities.

Impact of Secondary Structure of Polypeptides on Glucose Concentration Sensitivity of Nanocarriers for Insulin Delivery.

A reasonably intelligent response to glucose concentration fluctuations is crucial for developing a self-regulated insulin delivery system. Inspired by the relationship between the higher ordered structures of proteins and their versatile functions, the introduction of polypeptides capable of mimicking different secondary structures into the delivery system will be anticipated for adjusting glucose concentration sensitivity. Herein, this work presents the impact of different secondary structural architectures of polypeptide blocks on the stability of glucose-responsive complex nanoparticles (CNPs) in the normal physiological environment and their response to the stimuli of normoglycemic and hyperglycemic conditions in vitro. Results from the conformational investigations of the CNPs carried out using circular dichroism and insulin release under the different stimuli suggested that the stability and glucose sensitivity of the CNPs are closely related to the secondary structure composition of the polypeptide blocks. The CNPs with a dominant α-helix structure exhibit a promising potential to improve normal glycemic control and to reduce the incidences of hyperglycemia and hypoglycemia both in vitro and in vivo.

A poly(ascorbyl acrylate)-containing nanoplatform with anticancer activity and the sequential combination therapy with its loaded paclitaxel.

Despite progress, the combination therapy of a nanoscale delivery system and its loaded drug to increase the efficiency of anticancer treatment still remains a challenge. In this study, taking advantage of ascorbic acid with anticancer activity, complex nanovehicles were designed and constructed by co-assembly of the amphiphilic block polymers poly(ascorbyl acrylate)-block-poly(lactic acid) (PAA-b-PLA) and maleimide-decorating poly(ethylene glycol)-block-poly(lactic acid) (Mal-PEG-b-PLA) in aqueous solution. The combination of the nanoparticles' large surface and structural repeating characteristics of PAA led to an exponential increase in the ascorbyl content on the nanoparticle surface, which endowed the nanovehicles themselves with desired anticancer activity. In vitro cytotoxicity assays against normal cell line NIH3T3 and breast cancer cell line MCF-7 demonstrated that PAA-b-PLA/Mal-PEG-b-PLA complex nanoparticles exhibited benign biocompatibility against normal cells and prominent cancer inhibition ability. Paclitaxel (PTX)-loaded complex nanoparticles against MCF-7 were further investigated by MTS assay and flow cytometry. As a result, a synergistic effect of the complex nanoparticles and the loaded PTX in inducing cancer cell apoptosis was apparently noted. The newly developed PAA-b-PLA/Mal-PEG-b-PLA complex nanoparticles not only served as an effective and safe vector to deliver the therapeutic agents to the targeted site, but more importantly, they could also combine with the loaded therapeutic agents to achieve a synergistic effect for improving tumor inhibition efficiency.

Synthesis of Y-shaped poly(solketal acrylate)-containing block copolymers and study on the thermoresponsive behavior for micellar aggregates.

One novel type of Y-shaped amphiphilic copolymers with two hydrophobic poly(solketal acrylate) (PSA) branches and one hydrophilic monomethoxy poly(ethylene glycol) (MPEG) block was synthesized by atom transfer radical polymerization (ATRP). These Y-shaped polymers can disperse in aqueous media to self-assemble into micellar aggregates with a spherical core-shell structure. The aqueous copolymer solutions exhibited transmittancy transition in the temperature range of 30-60 °C via optical transmittance measurements. An interesting thermo-dependent size of the micellar aggregates was observed by dynamic light scattering techniques and transmission electron microscopy, which showed that the micelle diameters were decreased with temperature increasing. The nile red release from the micelles at 25 °C and 37 °C under various pHs showed that temperature has great influence on release behavior. With good biocompatibility, the micellar aggregates formed from MPEG-block-(PSA)(2) may serve as one promising thermosensitive nanovehicle for targeted drug delivery.