Manipulation of the Magnetic State of a Porphyrin-Based Molecule on Gold: From Kondo to Quantum Nanomagnet via the Charge Fluctuation Regime.

By moving individual Fe-porphyrin-based molecules with the tip of a scanning tunneling microscope in the vicinity of the elbow of the herringbone-reconstructed Au(111) containing a Br atom, we reversibly and continuously control their magnetic state. Several regimes are obtained experimentally and explored theoretically: from the integer spin limit, through intermediate magnetic states with renormalized magnetic anisotropy, until the Kondo-screened regime, corresponding to a progressive increase of charge fluctuations and mixed valency due to an increase in the interaction of the molecular Fe states with the substrate Fermi sea. Our study demonstrates the potential of utilizing charge fluctuations to generate and tune quantum magnetic states in molecule-surface hybrids.

RNF126-Mediated MRE11 Ubiquitination Activates the DNA Damage Response and Confers Resistance of Triple-Negative Breast Cancer to Radiotherapy.

Triple-negative breast cancer (TNBC) has higher molecular heterogeneity and metastatic potential and the poorest prognosis. Because of limited therapeutics against TNBC, irradiation (IR) therapy is still a common treatment option for patients with lymph nodes or brain metastasis. Thus, it is urgent to develop strategies to enhance the sensitivity of TNBC tumors to low-dose IR. Here, the authors report that E3 ubiquitin ligase Ring finger protein 126 (RNF126) is important for IR-induced ATR-CHK1 pathway activation to enhance DNA damage repair (DDR). Mechanistically, RNF126 physically associates with the MRE11-RAD50-NBS1 (MRN) complex and ubiquitinates MRE11 at K339 and K480 to increase its DNA exonuclease activity, subsequent RPA binding, and ATR phosphorylation, promoting sustained DDR in a homologous recombination repair-prone manner. Accordingly, depletion of RNF126 leads to increased genomic instability and radiation sensitivity in both TNBC cells and mice. Furthermore, it is found that RNF126 expression is induced by IR activating the HER2-AKT-NF-κB pathway and targeting RNF126 expression with dihydroartemisinin significantly improves the sensitivity of TNBC tumors in the brain to IR treatment in vivo. Together, these results reveal that RNF126-mediated MRE11 ubiquitination is a critical regulator of the DDR, which provides a promising target for improving the sensitivity of TNBC to radiotherapy.

Construction and Optimization of Herpes Simplex Virus Vectors for Central Nervous System Gene Delivery based on CRISPR/Cas9-mediated Genome Editing.

AIMS: We aim to define parameters that affect the safety and long-term transgene expression of attenuated HSV-1 vectors and optimize the expression cassettes to achieve robust and sustained expression in CNS. BACKGROUND: Engineered, attenuated Herpes Simplex Virus (HSV) vectors are promising vehicles for gene delivery to the peripheral and central nervous systems. The virus latent promoter (LAP) is commonly used to drive exogenous gene expression; however, parameters affecting the safety and longterm transgene expression of attenuated HSV-1 vectors have not been fully understood. OBJECTIVE: The study aimed at using CRISPR-Cas9 system to construct attenuated HSV-1 vectors and examine the influence of transgene cassette construction and insertion site on transgene expression and vector safety. METHODS: In this study, we used a CRISPR-Cas9 system to accurately and efficiently edit attenuated HSV-1 strain 1716, and construct two series of recombinant virus LMR and LMRx with different sets of gene cassettes insertion in Exon1(LAP2) and 2.0 kb intron downstream of LAP, respectively. The transgene expression and viral gene transcriptional kinetics were compared in in vitro cell lines. The reporter gene expression and safety profiles of each vector were further evaluated in mouse hippocampus gene transduction model. RESULTS: The in vitro cell line analysis indicated that the insertion of a gene expression cassette would disrupt virus gene transcription. Mouse hippocampus transducing analysis suggested that complete expression cassette insertion at 2.0 kb intron could achieve robust and longtime gene expression than the other constructs. Recombinants with gene expression cassettes lacking Poly (A) induced significant neuronal inflammation due to persistent viral antigen expression and microglia activation. CONCLUSION: Our results indicated that the integrity of LAT transcripts was not necessary for establishment of long-term latent expression. Exogenous strong promoters (like cBh promoter) could remain active during latency when placed in Exon1 or 2.0 Kb Intron of LAT locus, although their transcriptional activity declined with time. Consistent with previous research, the foreign gene expression would last much longer when the gene cassette was located downstream of Exon1, which suggested a role of LAP2 in maintaining promoter activity during latency. Besides, over-transcription of the downstream part of LAT may induce continuous activation of the attenuated vectors, which suggests an important role of LAT in maintaining viral reactivation potential.