Whole-Genome Sequence Analysis of <i>Flammulina filiformis</i> and Functional Validation of <i>Gad</i>, a Key Gene for γ-Aminobutyric Acid Synthesis
<i>Flammulina filiformis</i> is one of the widely produced edible fungi worldwide. It is rich in γ-aminobutyric acid (GABA), a non-protein amino acid with important physiological functions in humans. To investigate the functions of key genes in the GABA metabolic pathway of <i>F. f...
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2024-12-01
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| author | Wenyun Li Junjun Shang Dapeng Bao Jianing Wan Chenli Zhou Zhan Feng Hewen Li Youran Shao Yingying Wu |
| author_facet | Wenyun Li Junjun Shang Dapeng Bao Jianing Wan Chenli Zhou Zhan Feng Hewen Li Youran Shao Yingying Wu |
| author_sort | Wenyun Li |
| collection | DOAJ |
| description | <i>Flammulina filiformis</i> is one of the widely produced edible fungi worldwide. It is rich in γ-aminobutyric acid (GABA), a non-protein amino acid with important physiological functions in humans. To investigate the functions of key genes in the GABA metabolic pathway of <i>F. filiformis</i>, we isolated the monokaryon <i>Fv-HL23-1</i> from the factory-cultivated <i>F. filiformis</i> strain <i>Fv-HL23</i> and then sequenced and assembled the genome using the PacBio Sequel and Illumina NovaSeq sequencing platforms. The results showed that the genome comprised 140 scaffolds with a total length of 40.96 Mb, a GC content of 49.62%, an N50 of 917,125 bp, and 14,256 protein-coding genes. Phylogenetic analysis based on the whole genome revealed a close evolutionary relationship of <i>Fv-HL23-1</i> with <i>Armillaria mellea</i>, <i>Lentinula edodes</i>, and <i>Schizophyllum commune</i>. A total of 589 carbohydrate-active enzymes were identified in the genome of <i>Fv-HL23-1</i>, suggesting its strong lignocellulose degradation ability, and 108 CYP450 gene family members were identified, suggesting important functions such as resistance to stress, secondary metabolite synthesis, and growth and development. The <i>F. filiformis</i> proteins glutamate decarboxylase 1 (Ff-GAD1) and glutamate decarboxylase 2 (Ff-GAD2), which may be responsible for GABA synthesis, were identified by protein alignment. Molecular docking analysis showed that Ff-GAD2 may have better catalytic activity than Ff-GAD1. To verify the function of <i>Ff-gad2</i>, its heterologous expression in the mycelia of the mononuclear <i>Hypsizigus marmoreus</i> was analyzed. Compared with wild type, the GABA content of mycelia was increased by 85.40–283.90%, the growth rate was increased by 9.39 ± 2.35%, and the fresh weight was increased by 18.44 ± 7.57%. Ff-GAD2 may play a catalytic role in GABA synthesis. In addition, the expression of the full-length <i>Ff-gad2</i> gene was increased by 7.96 ± 1.39 times compared with the exon expression level in <i>H. marmoreus</i> mycelia, suggesting that the intron may contribute to the heterologous expression of Ff-GAD2. Based on whole-genome sequencing, we analyzed the enzyme system related to the important life activities of <i>F</i>. <i>filiformis</i>, focusing on the function of Ff-GAD, a key enzyme in the GABA synthesis pathway. The results lay a foundation for elucidating the GABA metabolism pathway of edible fungi and developing targeted breeding strategies for GABA-producing edible fungi. |
| format | Article |
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| institution | Kabale University |
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| language | English |
| publishDate | 2024-12-01 |
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| spelling | doaj-art-442fae09253b4c01a527f7eaa4a1a4e82024-12-27T14:33:58ZengMDPI AGJournal of Fungi2309-608X2024-12-01101286210.3390/jof10120862Whole-Genome Sequence Analysis of <i>Flammulina filiformis</i> and Functional Validation of <i>Gad</i>, a Key Gene for γ-Aminobutyric Acid SynthesisWenyun Li0Junjun Shang1Dapeng Bao2Jianing Wan3Chenli Zhou4Zhan Feng5Hewen Li6Youran Shao7Yingying Wu8College of Food Sciences & Technology, Shanghai Ocean University, Shanghai 201306, ChinaCollege of Food Sciences & Technology, Shanghai Ocean University, Shanghai 201306, ChinaCollege of Food Sciences & Technology, Shanghai Ocean University, Shanghai 201306, ChinaNational Engineering Research Center of Edible Fungi, Key Laboratory of Applied Mycological Resources and Utilization of Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, ChinaNational Engineering Research Center of Edible Fungi, Key Laboratory of Applied Mycological Resources and Utilization of Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, ChinaJiangsu Chinagreen Biological Technology Co., Ltd., Siyang 223700, ChinaJiangsu Chinagreen Biological Technology Co., Ltd., Siyang 223700, ChinaNational Engineering Research Center of Edible Fungi, Key Laboratory of Applied Mycological Resources and Utilization of Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, ChinaCollege of Food Sciences & Technology, Shanghai Ocean University, Shanghai 201306, China<i>Flammulina filiformis</i> is one of the widely produced edible fungi worldwide. It is rich in γ-aminobutyric acid (GABA), a non-protein amino acid with important physiological functions in humans. To investigate the functions of key genes in the GABA metabolic pathway of <i>F. filiformis</i>, we isolated the monokaryon <i>Fv-HL23-1</i> from the factory-cultivated <i>F. filiformis</i> strain <i>Fv-HL23</i> and then sequenced and assembled the genome using the PacBio Sequel and Illumina NovaSeq sequencing platforms. The results showed that the genome comprised 140 scaffolds with a total length of 40.96 Mb, a GC content of 49.62%, an N50 of 917,125 bp, and 14,256 protein-coding genes. Phylogenetic analysis based on the whole genome revealed a close evolutionary relationship of <i>Fv-HL23-1</i> with <i>Armillaria mellea</i>, <i>Lentinula edodes</i>, and <i>Schizophyllum commune</i>. A total of 589 carbohydrate-active enzymes were identified in the genome of <i>Fv-HL23-1</i>, suggesting its strong lignocellulose degradation ability, and 108 CYP450 gene family members were identified, suggesting important functions such as resistance to stress, secondary metabolite synthesis, and growth and development. The <i>F. filiformis</i> proteins glutamate decarboxylase 1 (Ff-GAD1) and glutamate decarboxylase 2 (Ff-GAD2), which may be responsible for GABA synthesis, were identified by protein alignment. Molecular docking analysis showed that Ff-GAD2 may have better catalytic activity than Ff-GAD1. To verify the function of <i>Ff-gad2</i>, its heterologous expression in the mycelia of the mononuclear <i>Hypsizigus marmoreus</i> was analyzed. Compared with wild type, the GABA content of mycelia was increased by 85.40–283.90%, the growth rate was increased by 9.39 ± 2.35%, and the fresh weight was increased by 18.44 ± 7.57%. Ff-GAD2 may play a catalytic role in GABA synthesis. In addition, the expression of the full-length <i>Ff-gad2</i> gene was increased by 7.96 ± 1.39 times compared with the exon expression level in <i>H. marmoreus</i> mycelia, suggesting that the intron may contribute to the heterologous expression of Ff-GAD2. Based on whole-genome sequencing, we analyzed the enzyme system related to the important life activities of <i>F</i>. <i>filiformis</i>, focusing on the function of Ff-GAD, a key enzyme in the GABA synthesis pathway. The results lay a foundation for elucidating the GABA metabolism pathway of edible fungi and developing targeted breeding strategies for GABA-producing edible fungi.https://www.mdpi.com/2309-608X/10/12/862monokaryon<i>Hypsizigus marmoreus</i>glutamate decarboxylasemolecular dockingheterologous expressionmetabolic pathways |
| spellingShingle | Wenyun Li Junjun Shang Dapeng Bao Jianing Wan Chenli Zhou Zhan Feng Hewen Li Youran Shao Yingying Wu Whole-Genome Sequence Analysis of <i>Flammulina filiformis</i> and Functional Validation of <i>Gad</i>, a Key Gene for γ-Aminobutyric Acid Synthesis Journal of Fungi monokaryon <i>Hypsizigus marmoreus</i> glutamate decarboxylase molecular docking heterologous expression metabolic pathways |
| title | Whole-Genome Sequence Analysis of <i>Flammulina filiformis</i> and Functional Validation of <i>Gad</i>, a Key Gene for γ-Aminobutyric Acid Synthesis |
| title_full | Whole-Genome Sequence Analysis of <i>Flammulina filiformis</i> and Functional Validation of <i>Gad</i>, a Key Gene for γ-Aminobutyric Acid Synthesis |
| title_fullStr | Whole-Genome Sequence Analysis of <i>Flammulina filiformis</i> and Functional Validation of <i>Gad</i>, a Key Gene for γ-Aminobutyric Acid Synthesis |
| title_full_unstemmed | Whole-Genome Sequence Analysis of <i>Flammulina filiformis</i> and Functional Validation of <i>Gad</i>, a Key Gene for γ-Aminobutyric Acid Synthesis |
| title_short | Whole-Genome Sequence Analysis of <i>Flammulina filiformis</i> and Functional Validation of <i>Gad</i>, a Key Gene for γ-Aminobutyric Acid Synthesis |
| title_sort | whole genome sequence analysis of i flammulina filiformis i and functional validation of i gad i a key gene for γ aminobutyric acid synthesis |
| topic | monokaryon <i>Hypsizigus marmoreus</i> glutamate decarboxylase molecular docking heterologous expression metabolic pathways |
| url | https://www.mdpi.com/2309-608X/10/12/862 |
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