OsGLU3, a Putative Membrane-Bound Endo-1, 4-Beta-Glucanase, Is Required for Root Cell Elongation and Division in Rice （Oryza sativa L.）

Plant roots move through the soil by elongation. This is vital to their ability to anchor the plant and acquire water and minerals from the soil. In order to identify new genes involved in root elongation in rice, we screened an ethyl methane sulfonate （EMS）-mutagenized rice library, and isolated a short root mutant, Osglu3-1. The map-based cloning results showed that the mutant was due to a point mutation in OsGLU3, which encodes a putative membrane-bound endo- 1,4-13-glucanase. Osglu3-1 displayed less crystalline cellulose content in its root cell wall, shorter root cell length, and a slightly smaller root meristem as visualized by restricted expression of OsCYCBI, I：GUS. Exogenous application of glu- cose can suppress both the lower root cell wall cellulose content and short root phenotypes of Osglu3-1. Consistently, OsGLU3 is ubiquitously expressed in various tissues with strong expression in root tip, lateral root, and crown root pri- modia. The fully functional OsGLU3-GFP was detected in plasma membrane, and FM4-64-1abeled compartments in the root meristem and elongation zones. We also found that phosphate starvation, an environmental stress, altered cell wall cel- lulose content to modulate root elongation in a OsGLU3-dependant way.

Activation of anthrachamycin biosynthesis in Streptomyces chattanoogensis L10 by site-directed mutagenesis of rpoB

Genome sequencing projects revealed massive cryptic gene clusters encoding the undiscovered secondary metabolites in Streptomyces. To investigate the metabolic products of silent gene clusters in Streptomyces chattanoogensis L10(CGMCC 2644), we used site-directed mutagenesis to generate ten mutants with point mutations in the highly conserved region of rpsL(encoding the ribosomal protein S12) or rpoB(encoding the RNA polymerase β-subunit). Among them, L10/RpoB(H437 Y) accumulated a dark pigment on a yeast extract-malt extract-glucose(YMG) plate. This was absent in the wild type. After further investigation, a novel angucycline antibiotic named anthrachamycin was isolated and determined using nuclear magnetic resonance(NMR) spectroscopic techniques. Quantitative real-time polymerase chain reaction(qRT-PCR) analysis and electrophoretic mobility shift assay(EMSA) were performed to investigate the mechanism underlying the activation effect on the anthrachamycin biosynthetic gene cluster. This work indicated that the rpoB-specific missense H437 Y mutation had activated anthrachamycin biosynthesis in S. chattanoogensis L10. This may be helpful in the investigation of the pleiotropic regulation system in Streptomyces.

m4C DNA methylation regulates biosynthesis of daptomycin in Streptomyces roseosporus L30

Despite numerous studies on transcriptional level regulation by single genes in drug producing Actinomyces,the global regulation based on epigenetic modification is not well explored.N4-methylcytosine(m4C),an abundant epigenetic marker in Actinomycetes’genome,but its regulatory mechanism remains unclear.In this study,we identify a m4C methyltransferase(SroLm3)in Streptomyces roseosporus L30 and multi-omics studies were performed and revealed SroLm3 as a global regulator of secondary metabolism.Notably,three BGCs inΔsroLm3 strain exhibited decreased expression compared to wild type.In-frame deletion of sroLm3 in S.roseosporus L30 further revealed its role in enhancing daptomycin production.In summary,we characterized a m4C methyltransferase,revealed the function of m4C in secondary metabolism regulation and biosynthesis of red pigment,and mapped a series of novel regulators for daptomycin biosynthesis dominated by m4C methylation.Our research further indicated that m4C DNA methylation may contribute to a metabolic switch from primary to secondary metabolism in Actinomyces.