LC3B drives transcription-associated homologous recombination via direct interaction with R-loops.

Exploring the connection between ubiquitin-like modifiers (ULMs) and the DNA damage response (DDR), we employed several advanced DNA damage and repair assay techniques and identified a crucial role for LC3B. Notably, its RNA recognition motif (RRM) plays a pivotal role in the context of transcription-associated homologous recombination (HR) repair (TA-HRR), a particular subset of HRR pathways. Surprisingly, independent of autophagy flux, LC3B interacts directly with R-loops at DNA lesions within transcriptionally active sites via its RRM, promoting TA-HRR. Using native RNA immunoprecipitation (nRIP) coupled with high-throughput sequencing (nRIP-seq), we discovered that LC3B also directly interacts with the 3'UTR AU-rich elements (AREs) of BRCA1 via its RRM, influencing its stability. This suggests that LC3B regulates TA-HRR both proximal to and distal from DNA lesions. Data from our LC3B depletion experiments showed that LC3B knockdown disrupts end-resection for TA-HRR, redirecting it towards the non-homologous end joining (NHEJ) pathway and leading to chromosomal instability, as evidenced by alterations in sister chromatid exchange (SCE) and interchromosomal fusion (ICF). Thus, our findings unveil autophagy-independent functions of LC3B in DNA damage and repair pathways, highlighting its importance. This could reshape our understanding of TA-HRR and the interaction between autophagy and DDR.

USP39 promotes non-homologous end-joining repair by poly(ADP-ribose)-induced liquid demixing.

Mutual crosstalk among poly(ADP-ribose) (PAR), activated PAR polymerase 1 (PARP1) metabolites, and DNA repair machinery has emerged as a key regulatory mechanism of the DNA damage response (DDR). However, there is no conclusive evidence of how PAR precisely controls DDR. Herein, six deubiquitinating enzymes (DUBs) associated with PAR-coupled DDR were identified, and the role of USP39, an inactive DUB involved in spliceosome assembly, was characterized. USP39 rapidly localizes to DNA lesions in a PAR-dependent manner, where it regulates non-homologous end-joining (NHEJ) via a tripartite RG motif located in the N-terminus comprising 46 amino acids (N46). Furthermore, USP39 acts as a molecular trigger for liquid demixing in a PAR-coupled N46-dependent manner, thereby directly interacting with the XRCC4/LIG4 complex during NHEJ. In parallel, the USP39-associated spliceosome complex controls homologous recombination repair in a PAR-independent manner. These findings provide mechanistic insights into how PAR chains precisely control DNA repair processes in the DDR.

Crystallization Kinetics in BaTiO3 Synthesis from Hydrate Precursors via Microwave-Assisted Heat Treatment.

The crystallization kinetics in BaTiO3 synthesis from hydrate precursors via microwave-assisted heating (MWH) were investigated. The structural and chemical features of powders synthesized via MWH and conventional heating (CH) were compared. The charged radicals generated under microwave irradiation were identified by chemical analysis and real-time charge flux measurements. Using Ba(OH)2∙H2O (BH1), Ba(OH)2 (BH0), and BaCO3 (BC) as the precursors for a Ba source, and TiO2∙4H2O (TH) for a Ti source, three different mixture samples, BH1TH (BH1 + TH), BH0TH (BH0 + TH), and BCTH (BC + TH), were heat-treated in the temperature range of 100-900 °C. BaTiO3 powders were synthesized at temperatures as low as 100 °C when sample BH1TH was subjected to MWH. Based on the growth exponent (n), the synthesis reactions were inferred to be diffusion-controlled processes (3 ≤ n ≤ 4) for MWH and interface-controlled processes (2 ≤ n ≤ 3) for CH. Current densities of approximately 0.073 and 0.022 mA/m2 were measured for samples BH1TH and BH0TH, respectively, indicating the generation of charged radicals by the interaction between the precursors and injected microwaves. The radicals were determined as OH- groups by X-ray photoelectron spectroscopy and Fourier transform infrared spectroscopy.

Effects of Microwave-Assisted Heat Treatments on the Nanostructural Evolution of Amorphous Silicon Oxycarbide Thin Films.

This study investigated the effects of heat treatment on changes in the nanostructure of amorphous silicon oxycarbide thin films. Hydrogenated amorphous silicon oxycarbide (a-Si0.6C0.3O0.1:H) thin films were prepared via plasma-enhanced chemical vapor deposition. The films were subjected to post-deposition heat treatments via microwave-assisted heating, which resulted in the formation of nanocrystals of SiC and Si in the a-Si0.6C0.3O0.1:H matrix at temperatures as low as ~800 °C. The crystallization activation energies of SiC and Si were determined to be 1.32 and 1.04 eV, respectively lower than those obtained when the sample was heat-treated via conventional heating (CH). Microwaves can be used to fabricate nanocrystals at a temperature approximately ~300 °C lower than that required for CH. The optical and nanostructural evolutions after post-deposition heat treatments were examined using photoluminescence (PL) and X-ray diffraction. The position of the PL peaks of the nanocrystals varied from ~425 to ~510 nm as the annealing temperature was increased from 800 to 1000 °C. In this study the optical band gap of SiC and Si varied from ~2.92 to ~2.40 eV and from ~2.00 to ~1.79 eV, as the size of the SiC and Si nanocrystals varied with respect to the heating temperature and isothermal holding time, respectively.

Post-Translational Regulation of the RSF1 Chromatin Remodeler under DNA Damage.

Chromatin remodeling factors are involved in many cellular processes such as transcription, replication, and DNA damage response by regulating chromatin structure. As one of chromatin remodeling factors, remodeling and spacing factor 1 (RSF1) is recruited at double strand break (DSB) sites and regulates ataxia telangiectasia mutated (ATM) -dependent checkpoint pathway upon DNA damage for the efficient repair. RSF1 is overexpressed in a variety of cancers, but regulation of RSF1 levels remains largely unknown. Here, we showed that protein levels of RSF1 chromatin remodeler are temporally upregulated in response to different DNA damage agents without changing the RSF1 mRNA level. In the absence of SNF2h, a binding partner of RSF1, the RSF1 protein level was significantly diminished. Intriguingly, the level of RSF1-3SA mutant lacking ATM-mediated phosphorylation sites significantly increased, and upregulation of RSF1 levels under DNA damage was not observed in cells overexpressing ATM kinase. Furthermore, failure in the regulation of RSF1 level caused a significant reduction in DNA repair, whereas reconstitution of RSF1, but not of RSF1-3SA mutants, restored DSB repair. Our findings reveal that temporal regulation of RSF1 levels at its post-translational modification by SNF2h and ATM is essential for efficient DNA repair.