Comparative Transcriptome Analysis Of Rhizopus Chinensis With Different Mycelia Morphology In Submerged Fermentation For Lipase Production

The metabolism and fermentation of filamentous fungus are influenced significantly by mycelia morphology.Rhizopus chinensis CCTCC M201021 can form different mycelia morphology in submerge fermentation of lipase with different fermentation performance.We investigated transcriptome profiles of two typical mycelia morphology of R.chinensis,the aggregated and dispersed,in order to seek the possible regulation mechanisms on mycelia morphology.Basing on high throughput RNA sequencing,comparative analysis of differential genes transcription in R.chinensis with different morphology were carried out.The results provided some clues about regulation mechanisms on mycelia morphology and the influence of mycelia morphology.The main results were described as follows:(1)To investigate the intrinsic differences between the two mycelia of R.chinensis,two sets of mycelia samples were harvest from the aggregated and dispersed cultures before and after morphology change respectively.After RNA isolution and RNA sequencing,transcriptome data were obtained.The percentage of reads of these transcripts mapped to genes surpassed 90%,while the percentages of reads mapped to exons were all higher than 99%,which meet the requirements of transcriptome analysis.(2)Based on the RNA sequencing data,the overall transcriptome profiles of R.chinensis in the aggregated and dispersed culture at different culture times were analysed.These results indicated that transcription patterns between the aggregated and the dispersed cultures were quite different,even at the early stage of cultures.The earlier stage of fermentation,there were more down-regulated genes in the aggregated mycelia,and later period there were more upregulated genes in the aggregated mycelia.(3)The RPKM values of gene expression of R.chinensis in the aggregated and dispersed culture at different culture times were analysed.Through analysis of the 20 genes featuring the highest RPKM values in the transcriptome of R.chinensis,we found that most of the highest RPKM value genes were ribosomal proteins,especially in the aggregated mycelia.In addition,chitinase genes and signal transduction genes in dispersed of R.chinensis were highly expressed.The analysis of 20 most up-regulated genes and 20 most down-regulated genes indicated that the cellular process and signal genes were up-regulated in dispersed.of R.chinensis,besides different metabolism genes for two morphology of R.chinensis.The value of RPKM suggested that most of unique transcribed geneswere transcribed at low level.The different morphology influenced the variety and expression level of genes for R.chinensis,and some genes were expressed in a specific culture,even the same function protein were expressed by different genes.(4)All the DEGs of the mycelia with two typical morphology were annotated and functionally classified based on the Gene Ontology(GO).GO functional enrichment analysis indicated the importance of biological processes' differences in the mycelia with different morphology during the cultures.The DEGs enriched significantly in GO functional catalogies were annotated and classified according to the KEGG and KOG.The change of mycelia morphology could be interpreted as a very complicated biological process.Further functional analysis and comparison of these DEGs would provide us clues to understand the underlying difference of mycelia morphology.The functional analysis of these DEGs suggested that the dispersed mycelia were regulated by complicated signal transduction mechanisms in cellular responses to stress or stimuli at the early stage of the culture,even during the whole culture.In other words,the aggregation of mycelia might be a simple biological process naturally,while the dispersed cells tended to remain their dispersed state for growth under complicated regulation mechanisms.In addition,the aggregation of mycelia would result in the mass transfer limitation,these differential expressions of genes might be caused by the change of microenvironment around the cells.Moreover,The lipase activity of R.chinensis agreed with the transcription level of the lipase genes.In conclusion,the change of mycelial morphology should be considered as cell differentiation with different biological function,instead of simply mechanical conglomerates of cells.