Ⅲ-4. Radiation Chemistry & Biology

RIKEN Accel. Prog. Rep. 53

Mutants of the ectomycorrhizal mushroom Tricholoma matsutake thatexert detrimental eﬀects on its symbiotic partner Pinus densiflora †H. Murata,∗1 S. Nakano,∗1 T. Yamanaka,∗1 T. Abe,∗2 H. Ichida,∗2 Y. Hayashi,∗2 T. Shimokawa,∗1 K. Tahara,∗1and A. Ohta∗3
Tricholoma matsutake is an ectomycorrhizal fungus
that produces the prized mushrooms “matsutake” insymbiotic association with conifers.1) In our attempts
to induce mutants of T. matsutake that could be useful in artiﬁcial cultivation for fruiting by γ-ray irradiation at 500 Gy, we isolated a mutant, designated asG1, that grew better in substrates than the wild-type
NBRC 33136.1) The phenotype of G1 on agar plates
somewhat resembled that of Ar 59, which had been isolated by argon-ion-beam irradiation at 500 Gy; bothmutants overproduced amylase and cellulase activities
and exhibited a mycelial morphology diﬀerent from thewild-type, forming a fuzzy cotton-ball-like colony, rather
than a mycelial mat.1,2) In the present study, we examined how G1 and Ar 59 inﬂuence the plant growth ofPinus densiflora seedlings in vitro.
The inoculation was performed with two diﬀerent
methods. The ﬁrst involved sterile 1-week-old P. densiflora seedlings, in which the seedlings were planted
in a pumiceous soil substrate ﬁlled with T. matsutake
mycelia and axenically co-cultured at 23◦ C for 120 days.
The second involved sterile 1-month-old P. densiflora
seedlings already grown in the pumiceous soil, in which
T. matsutake mycelia were placed on the surface of thesubstrate where the shoot base emerged; they were axenically co-cultured at 23◦ C for 150 days.
In the ﬁrst experiment, G1 exhibited a signiﬁcantdetrimental eﬀect on both above- and below-ground organs; root systems were severely underdeveloped or collapsed, leading in turn to aboveground wilting (Fig. 1).

Fig. 1. Biomass measurements of 1-week-old P. densiflora
seedlings co-cultured with T. matsutake strains for 120
days.
†∗1
∗2
∗3

Condensed from the article in Botany 97, 463–474 (2019)
Forest Research and Management Organization
RIKEN Nishina Center
Lake Biwa Environmental Research InstituteFig. 2. Biomass measurements of 1-month-old P. densiflora
seedlings co-cultured with T. matsutake strains for 150
days.

This result contrasts with the plant-growth-promoting
eﬀect of Ar 59, as well as NBRC 33136; the former allowed the seedlings to gain signiﬁcantly higher aboveground and total biomasses than the control, while thelatter allowed them to gain signiﬁcantly higher aboveground, belowground, and total biomasses (Fig. 1).
In the second experiment, seedlings associated with
G1 exhibited signiﬁcant decreases in their belowgroundbiomasses, and many lateral roots degenerated significantly, reducing their rhizosphere biomasses (Fig. 2).
Seedlings associated with Ar 59, in contrast, exhibited neither plant-growth-promoting nor plant-growthinhibiting eﬀects (Fig. 2). Unlike these mutants, NBRC
33136 promoted plant growth; the aboveground andtotal biomasses signiﬁcantly increased, but the belowground biomass did not (Fig. 2).
Data suggest that there could be a regulatory region in
the genome of T. matsutake that determines plant associating behaviors, depending upon where the mutations
occur and what environment the fungus inhabits basedon its endophytic association with non-host plants.3)
The overproduction of degrading enzymatic activities is
not the cause of G1’s trait of harming its symbiotic partner, because Ar 59 overproduced the same enzymatic
activities without harming P. densiflora seedlings. Genomic and transcriptomic analyses of these mutants willallow us to unearth the unknown regulatory mechanisms
involved in the ectomycorrhizal symbiosis of T. matsutake.
References

1) H. Murata et al., Botany 97, 463 (2019).
2) H. Murata et al., Mycorrhiza 28, 171 (2018).
3) H. Murata et al., Mycorrhiza 23, 235 (2013).
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