Identification Of NADPH Oxidases And Analysis Of The Response Mechanism Under Abiotic Stress In Flammulina Velutipes

It is reported that NADPH oxidases which can produce ROS play an important role in fungal mycelial growth and fruiting body development,while the ROS as signal molecules regulate a variety of signal transduction,modify the structure of protein,transcription factors and genes and then regulate their function directly.At present there are lots of papers about NADPH oxidases under abiotic stress and ROS in plants and animals,while less in fungi.Based on the completion of the genome sequencing in Flammulina velutipes,according to literature reports,this study preliminarily analyzes NADPH oxidases by bioinformatics analysis and build the mechanisms of the subunit Cdc24 response to abiotic stress,hoping to provide certain theoretical basis for the research on the mechanism of fungal mycelial growth and fruiting in Flammulina velutipes.The analysis results show that the NADPH oxidases contain six subunits,including cell division regulatory proteins(Cdc24),NADPH oxidases homologous isomer 1(Noxl),NADPH oxidases homologous isomer 2(Nox2),NADPH oxidases regulator(NoxR),protein kinase activator(Beml),small GTP enzyme(Rac1).This study identifies the genetic structure of the above six genes based on the transcriptome sequencing platform.We confirmed the reliability of NADPH oxidases by the analysis of phylogenetic analysis,the structure of the conserved domains,transmembrane structure and the signal peptide prediction.In this study,we find the optimum pH of Flammulina velutipes is 6,the optimum pH of acid stress is 4,alkali stress is 11,the optimal concentration of NaCl stress is 0.8mol/L,the optimal concentration of H2O2 stress is 10mmol/L by the biological characteristics experiments of Flammulina velutipes.NADPH oxidases are more sensitive to the factors like solid or liquid culture medium,culture time,incubation temperature,incubation time,and cellophane and so on.By abiotic stress treatments and inhibitor treatments,we found that when the Flammulina velutipes respond to abiotic stress,NADPH oxidases could produce ROS,while H2O2 and O2-can act as signaling molecules acting on the subunits Cdc24,which indicates that H2O2 and O2-not only play a regulatory role on downstream genes,but also on the NADPH oxidases.These results completed the initial exploration about the identification NADPH oxidases and its subunit Cdc24 response to abiotic stress mechanisms.