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      Advances in Genetic Engineering Technology and Its Application in the Industrial Fungus Aspergillus oryzae

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          Abstract

          The filamentous fungus Aspergillus oryzae is an important strain in the traditional fermentation and food processing industries and is often used in the production of soy sauce, soybean paste, and liquor-making. In addition, A. oryzae has a strong capacity to secrete large amounts of hydrolytic enzymes; therefore, it has also been used in the enzyme industry as a cell factory for the production of numerous native and heterologous enzymes. However, the production and secretion of foreign proteins by A. oryzae are often limited by numerous bottlenecks that occur during transcription, translation, protein folding, translocation, degradation, transport, secretion, etc. The existence of these problems makes it difficult to achieve the desired target in the production of foreign proteins by A. oryzae. In recent years, with the decipherment of the whole genome sequence, basic research and genetic engineering technologies related to the production and utilization of A. oryzae have been well developed, such as the improvement of homologous recombination efficiency, application of selectable marker genes, development of large chromosome deletion technology, utilization of hyphal fusion techniques, and application of CRISPR/Cas9 genome editing systems. The development and establishment of these genetic engineering technologies provided a great deal of technical support for the industrial production and application of A. oryzae. This paper reviews the advances in basic research and genetic engineering technologies of the fermentation strain A. oryzae mentioned above to open up more effective ways and research space for the breeding of A. oryzae production strains in the future.

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          Most cited references163

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          A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity.

          Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems provide bacteria and archaea with adaptive immunity against viruses and plasmids by using CRISPR RNAs (crRNAs) to guide the silencing of invading nucleic acids. We show here that in a subset of these systems, the mature crRNA that is base-paired to trans-activating crRNA (tracrRNA) forms a two-RNA structure that directs the CRISPR-associated protein Cas9 to introduce double-stranded (ds) breaks in target DNA. At sites complementary to the crRNA-guide sequence, the Cas9 HNH nuclease domain cleaves the complementary strand, whereas the Cas9 RuvC-like domain cleaves the noncomplementary strand. The dual-tracrRNA:crRNA, when engineered as a single RNA chimera, also directs sequence-specific Cas9 dsDNA cleavage. Our study reveals a family of endonucleases that use dual-RNAs for site-specific DNA cleavage and highlights the potential to exploit the system for RNA-programmable genome editing.
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            Multiplex genome engineering using CRISPR/Cas systems.

            Functional elucidation of causal genetic variants and elements requires precise genome editing technologies. The type II prokaryotic CRISPR (clustered regularly interspaced short palindromic repeats)/Cas adaptive immune system has been shown to facilitate RNA-guided site-specific DNA cleavage. We engineered two different type II CRISPR/Cas systems and demonstrate that Cas9 nucleases can be directed by short RNAs to induce precise cleavage at endogenous genomic loci in human and mouse cells. Cas9 can also be converted into a nicking enzyme to facilitate homology-directed repair with minimal mutagenic activity. Lastly, multiple guide sequences can be encoded into a single CRISPR array to enable simultaneous editing of several sites within the mammalian genome, demonstrating easy programmability and wide applicability of the RNA-guided nuclease technology.
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              Signal integration in the endoplasmic reticulum unfolded protein response.

              The endoplasmic reticulum (ER) responds to the accumulation of unfolded proteins in its lumen (ER stress) by activating intracellular signal transduction pathways - cumulatively called the unfolded protein response (UPR). Together, at least three mechanistically distinct arms of the UPR regulate the expression of numerous genes that function within the secretory pathway but also affect broad aspects of cell fate and the metabolism of proteins, amino acids and lipids. The arms of the UPR are integrated to provide a response that remodels the secretory apparatus and aligns cellular physiology to the demands imposed by ER stress.
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                Author and article information

                Contributors
                Journal
                Front Microbiol
                Front Microbiol
                Front. Microbiol.
                Frontiers in Microbiology
                Frontiers Media S.A.
                1664-302X
                23 February 2021
                2021
                : 12
                : 644404
                Affiliations
                Co-Innovation Center for Sustainable Forestry in Southern China, College of Biology and the Environment, Nanjing Forestry University , Nanjing, China
                Author notes

                Edited by: Wanping Chen, Georg August University of Göttingen, Germany

                Reviewed by: Bin He, Jiangxi Science and Technology Normal University, China; Moriyuki Kawauchi, Kyoto University, Japan; Katsuya Gomi, Tohoku University, Japan

                *Correspondence: Feng-Jie Jin, jinfj@ 123456njfu.edu.cn

                This article was submitted to Food Microbiology, a section of the journal Frontiers in Microbiology

                Article
                10.3389/fmicb.2021.644404
                7940364
                33708187
                643847aa-a8a4-4bea-a304-7ac1405b031e
                Copyright © 2021 Jin, Hu, Wang and Jin.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                History
                : 21 December 2020
                : 05 February 2021
                Page count
                Figures: 0, Tables: 2, Equations: 0, References: 163, Pages: 14, Words: 0
                Funding
                Funded by: National Natural Science Foundation of China 10.13039/501100001809
                Categories
                Microbiology
                Review

                Microbiology & Virology
                aspergillus oryzae,fermentation industry,genetic engineering technology,cell factory,heterologous protein production

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