Presentation Information
[P02-165]Potential role of agmatine production in growth and asexual development in Aspergillus oryzae
○Jenna Hamaring Pinkan Kairupan1, Adina Amy Reikanisuji1, Akasaka Naoki1, Jun-ichi Maruyama2, Daisuke Watanabe1 (1. NAIST (Japan), 2. The Univ. Tokyo (Japan))
Keywords:
Agmatine,Arginine decarboxylase,Aspergillus oryzae,Solid-state fermentation
Agmatine is a naturally occurring polyamine synthesized from arginine via decarboxylation catalyzed by arginine decarboxylase (ADC). Agmatine has gained increasing attention because of its pleiotropic physiological benefits such as improvement of cognitive function and alleviation of neuropathic pain (1). Our previous studies demonstrated that Aspergillus oryzae, a filamentous fungus used for traditional Asian and Japanese fermented foods, produces agmatine specifically under solid-state cultivation conditions (2). We subsequently identified a novel arginine decarboxylase (Ao-ADC1), which catalyzes the conversion of arginine to agmatine in vitro (3). These findings suggest that agmatine biosynthesis may play a key role in the adaptation of A. oryzae to solid-state environments. However, its physiological relevance has yet to be elucidated.
To investigate the significance of Ao-adc1 (agmatine production) in various cellular processes, including mycelial growth and conidiation, under solid-state cultivation conditions, the Ao-adc1 disruptant (Δadc1) was constructed from A. oryzae RIB40 using the CRISPR-Cas9 genome editing system (4). Sequencing analysis identified a single base pair deletion in the coding region of Ao-adc1, leading to a frameshift mutation causing premature translation termination. The parental RIB40 and Δadc1 strains were cultivated on steamed rice, and the ADC activity was evaluated by in vitro assay. The cells of Δadc1 showed no detectable ADC activity, whereas those of RIB40 exhibited 137.16 pmol/min/µg GlcNAc of ADC activity. Consistent with this, Δadc1 accumulated no agmatine during solid-state fermentation of steamed rice, while RIB40 produced 4.4 mM agmatine. These results indicate that Ao-ADC1 is responsible for agmatine production in A. oryzae in vivo.
Morphological analysis found that the aerial mycelia were less fluffy in Δadc1 than in RIB40 when the strains were cultivated on a nutrient-rich agar plate and steamed rice. To examine this, the two strains were cultivated on a nutrient-rich agar plate, and the diameter of the colonies was measured over time. The colony diameter was significantly larger in RIB40 than in Δadc1 throughout cultivation, and the diameters after cultivation for 3 days were 43.7 ± 0.5 and 39.0 ± 0.8 mm in RIB40 and Δadc1, respectively. It was also observed that the onset of conidiation appeared earlier in Δadc1 than in the parental strain. We then cultivated the two strains on a nutrient-rich agar plate and counted the numbers of conidia generated after cultivation for 2 days. The conidial numbers of Δadc1 ((4.6 ± 0.1) × 107 conidia/plate) was markedly higher than those of the parental strain ((8.9 ± 2.8) × 106 conidia/plate). In contrast, the viability of the conidia was lower in Δadc1 (85.6 ± 5.8%) than in RIB40 (98.4 ± 2.3%). These findings suggest that agmatine may have important roles in mycelial elongation, asexual development, and conidial viability during solid-state cultivation in A. oryzae.
(1) Akasaka et al. 2026. Amino Acids 58:13.
(2) Akasaka et al. 2018. Appl Environ Microbiol 84:e00722-18.
(3) Murakami et al. 2024. Appl Environ Microbiol 90:e0029424.
(4) Katayama et al. 2019. Appl Environ Microbiol 85:e01896-18.
To investigate the significance of Ao-adc1 (agmatine production) in various cellular processes, including mycelial growth and conidiation, under solid-state cultivation conditions, the Ao-adc1 disruptant (Δadc1) was constructed from A. oryzae RIB40 using the CRISPR-Cas9 genome editing system (4). Sequencing analysis identified a single base pair deletion in the coding region of Ao-adc1, leading to a frameshift mutation causing premature translation termination. The parental RIB40 and Δadc1 strains were cultivated on steamed rice, and the ADC activity was evaluated by in vitro assay. The cells of Δadc1 showed no detectable ADC activity, whereas those of RIB40 exhibited 137.16 pmol/min/µg GlcNAc of ADC activity. Consistent with this, Δadc1 accumulated no agmatine during solid-state fermentation of steamed rice, while RIB40 produced 4.4 mM agmatine. These results indicate that Ao-ADC1 is responsible for agmatine production in A. oryzae in vivo.
Morphological analysis found that the aerial mycelia were less fluffy in Δadc1 than in RIB40 when the strains were cultivated on a nutrient-rich agar plate and steamed rice. To examine this, the two strains were cultivated on a nutrient-rich agar plate, and the diameter of the colonies was measured over time. The colony diameter was significantly larger in RIB40 than in Δadc1 throughout cultivation, and the diameters after cultivation for 3 days were 43.7 ± 0.5 and 39.0 ± 0.8 mm in RIB40 and Δadc1, respectively. It was also observed that the onset of conidiation appeared earlier in Δadc1 than in the parental strain. We then cultivated the two strains on a nutrient-rich agar plate and counted the numbers of conidia generated after cultivation for 2 days. The conidial numbers of Δadc1 ((4.6 ± 0.1) × 107 conidia/plate) was markedly higher than those of the parental strain ((8.9 ± 2.8) × 106 conidia/plate). In contrast, the viability of the conidia was lower in Δadc1 (85.6 ± 5.8%) than in RIB40 (98.4 ± 2.3%). These findings suggest that agmatine may have important roles in mycelial elongation, asexual development, and conidial viability during solid-state cultivation in A. oryzae.
(1) Akasaka et al. 2026. Amino Acids 58:13.
(2) Akasaka et al. 2018. Appl Environ Microbiol 84:e00722-18.
(3) Murakami et al. 2024. Appl Environ Microbiol 90:e0029424.
(4) Katayama et al. 2019. Appl Environ Microbiol 85:e01896-18.
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