Peroxisomal ß-oxidation regulates histone acetylation and DNA methylation in Arabidopsis.

Epigenetic markers, such as histone acetylation and DNA methylation, determine chromatin organization. In eukaryotic cells, metabolites from organelles or the cytosol affect epigenetic modifications. However, the relationships between metabolites and epigenetic modifications are not well understood in plants. We found that peroxisomal acyl-CoA oxidase 4 (ACX4), an enzyme in the fatty acid ß-oxidation pathway, is required for suppressing the silencing of some endogenous loci, as well as Pro35S:NPTII in the ProRD29A:LUC/C24 transgenic line. The acx4 mutation reduces nuclear histone acetylation and increases DNA methylation at the NOS terminator of Pro35S:NPTII and at some endogenous genomic loci, which are also targeted by the demethylation enzyme REPRESSOR OF SILENCING 1 (ROS1). Furthermore, mutations in multifunctional protein 2 (MFP2) and 3-ketoacyl-CoA thiolase-2 (KAT2/PED1/PKT3), two enzymes in the last two steps of the ß-oxidation pathway, lead to similar patterns of DNA hypermethylation as in acx4 Thus, metabolites from fatty acid ß-oxidation in peroxisomes are closely linked to nuclear epigenetic modifications, which may affect diverse cellular processes in plants.

METHIONINE ADENOSYLTRANSFERASE4 Mediates DNA and Histone Methylation.

DNA and histone methylation coregulate heterochromatin formation and gene silencing in animals and plants. To identify factors involved in maintaining gene silencing, we conducted a forward genetic screen for mutants that release the silenced transgene Pro35S::NEOMYCIN PHOSPHOTRANSFERASE II in the transgenic Arabidopsis (Arabidopsis thaliana) line L119 We identified MAT4/SAMS3/MTO3/AT3G17390, which encodes methionine (Met) adenosyltransferase 4 (MAT4)/S-adenosyl-Met synthetase 3 that catalyzes the synthesis of S-adenosyl-Met (SAM) in the one-carbon metabolism cycle. mat4 mostly decreases CHG and CHH DNA methylation and histone H3K9me2 and reactivates certain silenced transposons. The exogenous addition of SAM partially rescues the epigenetic defects of mat4 SAM content and DNA methylation were reduced more in mat4 than in three other mat mutants. MAT4 knockout mutations generated by CRISPR/Cas9 were lethal, indicating that MAT4 is an essential gene in Arabidopsis. MAT1, 2, and 4 proteins exhibited nearly equal activity in an in vitro assay, whereas MAT3 exhibited higher activity. The native MAT4 promoter driving MAT1, 2, and 3 cDNA complemented the mat4 mutant. However, most mat4 transgenic lines carrying native MAT1, 2, and 3 promoters driving MAT4 cDNA did not complement the mat4 mutant because of their lower expression in seedlings. Genetic analyses indicated that the mat1mat4 double mutant is dwarfed and the mat2mat4 double mutant was nonviable, while mat1mat2 showed normal growth and fertility. These results indicate that MAT4 plays a predominant role in SAM production, plant growth, and development. Our findings provide direct evidence of the cooperative actions between metabolism and epigenetic regulation.

Optical and interfacial layer properties of SiO2 films deposited on different substrates.

SiO2 films were deposited on fused silica, silicon, glass, germanium, and sapphire substrates by an ion beam sputtering technique. The optical properties of SiO2 films on different substrates and interfacial layer properties between SiO2 films and different substrates were researched by the spectroscopic ellipsometry technique. The refractive indices of SiO2 films deposited on different substrates are about 1.477 at the wavelength of 632.8 nm. The optical anisotropy property of SiO2 films on fused silica substrate is the best. The impact of thermal treatment on surface roughness and interfacial layer properties between SiO2 films and Si substrates were also investigated. When the annealing temperature is 550°C, the least surface thickness and thinnest interface layer thickness between v films and silicon substrate can be achieved. The results indicate that the surface and interface layer properties between SiO2 films and silicon substrate can be greatly improved when the optimum annealing temperature is selected.

Correlation between properties of HfO2 films and preparing parameters by ion beam sputtering deposition.

Ion beam sputtering is one of the most important technologies for preparing hafnium dioxide thin films. In this paper, the correlation between properties of hafnium dioxide thin films and preparing parameters was systematically researched by using the orthogonal experiment design method. The properties of hafnium oxide films (refractive index, extinction coefficient, deposition rate, stress, and inhomogeneity of refractive index) were studied. The refractive index, extinction coefficient, physical thickness, and inhomogeneity of refractive index were obtained by the multiple wavelength curve-fitting method from the reflectance and transmittance of single layers. The stress of thin film was measured by elastic deformation of the thin film-substrate system. An orthogonal experimental strategy was designed using substrate temperature, ion beam voltage, ion beam current, and oxygen flow rate as the variables. The experimental results indicated that the temperature of the substrate is the key influencing parameter on the properties of hafnium oxide films, while other preparing parameters are also correlated with specific properties. The experimental results are significant for selecting proper parameters for preparing hafnium oxide films with different applications.

[Applied research on the physical model for calculating optical constant of metal oxide films].

In the present paper, two metal oxide films of HfO2 and Ta2 O5 were prepared by ion beam sputtering technology. Through measuring ellipsometric parameters of HfO2 and Ta2O5 films, their optical constants can be inversion-calculated by nonlinear least squares techniques. In the fitting process, eight experiment groups were arranged by the orthogonal table L8 (2(7)). After selecting Cauchy model, the largest influencing factor for fitting optical constant of HfO2 is surface layer model and the largest influencing factor for fitting optical constant of Ta2 O5 is refractive index gradient model. The impact of different physical model on MSE and the order for selecting model in the fitting process are determined. According to the selecting model and the determined fitting order, optical constants of the two metal oxide films were inversion-calculated with adding weak absorption model, and the obtained MSE can descend 79% and 39% according to the initial value. The results indicated that the selecting model possesses definite physical significance in the fitting process. The obtained method can be applied in inversion-calculating many metal oxide films with weak absorption. It has wide application value. At 500 nm, the refractive index gradient of Ta2O5 films is greater than HfO2 films, while the extinction coefficient of HfO2 films is greater than Ta2O5. It was shown that Hf metal is easier to form stable oxide than Ta metal. And the absorption of HfO2 films is larger than Ta2O5 films.

ALBA protein complex reads genic R-loops to maintain genome stability in Arabidopsis.

The R-loop, composed of a DNA-RNA hybrid and the displaced single-stranded DNA, regulates diverse cellular processes. However, how cellular R-loops are recognized remains poorly understood. Here, we report the discovery of the evolutionally conserved ALBA proteins (AtALBA1 and AtALBA2) functioning as the genic R-loop readers in Arabidopsis. While AtALBA1 binds to the DNA-RNA hybrid, AtALBA2 associates with single-stranded DNA in the R-loops in vitro. In vivo, these two proteins interact and colocalize in the nucleus, where they preferentially bind to genic regions with active epigenetic marks in an R-loop-dependent manner. Depletion of AtALBA1 or AtALBA2 results in hypersensitivity of plants to DNA damaging agents. The formation of DNA breaks in alba mutants originates from unprotected R-loops. Our results reveal that the AtALBA1 and AtALBA2 protein complex is the genic R-loop reader crucial for genome stability in Arabidopsis.