National Institute for Basic Biology

Evolutionary Biology and Biodiversity

LABORATORY OF EVOLUTIONARY GENOMICS
Professor:

SHIGENOBU, Shuji

Specially Appointed Assistant Professor:
YODA, Shinichi
Postdoctoral Fellow:
KOBAYASHI, Yuuki
JSPS Postdoctoral Fellow: NOZAKI, Tomonari
MATSUDA, Naoki
SOKENDAI Graduate Student: YORIMOTO, Shunta
TAN, Kathrine
Visiting Scientist :HUANGH, Jia-Hsin
TONG, Xin
CHUNG, Chen-yo
Technical Assistant:
SUZUKI, Miyuzu
WEN, Hsin-I
IKEDA, Mika
Admin Support Staff:
ICHIKAWA, Mariko

Overview“Nothing, it seems, exists except as part of a network ofinteractions.” (Gilbert & Epel, 2008)
Every creature on Earth exists among a network of various
biological interactions. For example, many multicellularorganisms, including humans, harbor symbiotic bacteria intheir bodies. Some of them provide their hosts with essential
nutrients deficient in the host’s diet and others digest foods
that are indigestible by the host alone. Despite numerous
examples of symbioses and its intriguing outcomes, thegenetic and molecular basis underlying these interactions
remains elusive. The goal of our group is to establish a newinterdisciplinary science known as “Symbiogenomics”, wherewe aim to understand the network of biological interactions at
the molecular and genetic level. To this end, we take advantage of state-of-the-art genomics, such as next-generation
sequencing technologies and CRISPR-Cas9 genome editing.

Genomic revelations of a mutualism: the pea
aphid and its obligate bacterial symbiont

Aphid species bear intracellular symbiotic bacteria in thecytoplasm of bacteriocytes, which are specialized cells forharboring said bacteria. This mutualism is so obligate thatneither can reproduce independently. The genome sequenceof the pea aphid, Acyrthosiphon pisum, in consort withthat of bacterial symbiont Buchnera aphidicola illustrates
the remarkable interdependency between these two organisms (IAGC, 2010; Shigenobu et al., 2000; Shigenobu &Yorimoto, 2022). The genetic capacities of the pea aphid
and the symbiont for amino acid biosynthesis are complementary. Genome analysis revealed that the pea aphid has
undergone characteristic gene losses and duplications. The
IMB antibacterial immune pathway is missing several criticalgenes, which might account for the evolutionary success ofaphids in obtaining beneficial symbionts. Lineage-specificgene duplications have occurred in genes over a broad rangeof functional categories, which include signaling pathways,
miRNA machinery, chromatin modification and mitosis. The
importance of these duplications for symbiosis remains to
be determined. We found several instances of lateral genetransfer from bacteria to the pea aphid genome. Some ofthem are highly expressed in bacteriocytes.
We recently discovered a novel class of genes in the pea
aphid genome that encode small cysteine-rich proteins withsecretion signals that are expressed exclusively in the bacteriocytes of the pea aphid, and named these bacteriocytespecific cysteine-rich proteins (BCR) (Shigenobu & Stern,2013). The BCR mRNAs are first expressed at a developmental time point coinciding with the incorporation ofsymbionts strictly in the cells that contribute to the bacteriocyte, and this bacteriocyte-specific expression is maintained throughout the aphid’s life. Furthermore, some BCRs
showed antibiotic activity (Uchi et al., 2019). These results
suggest that BCRs act within bacteriocytes to mediate thesymbiosis with bacterial symbionts, which is reminiscent of
the cysteine-rich secreted proteins of leguminous plants thatalso regulate endosymbionts. Employment of small cysteinerich peptides may be a common tactic of host eukaryotes tomanipulate bacterial symbionts.

Figure 1. Pea aphids and the bacterial symbiont, Buchnera. Adult aphids(Left). A developing viviparous embryo which symbionts are infecting(Right). Scale bar = 20 um.

Visual overview of this lab’s work.

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Genomics of emerging model insects

Recent remarkable technological advancements allow us
to study any organisms at molecular and genetic levels,
which has been difficult for non-model organisms. Insectscience is also benefited by the technological innovation.
New entomology is starting to study the amazing variety ofmorphology, phisiology and ethology of insects by using
these state-of-the-art technologies. Our group is developing
a platform for emerging model insects to read the genomes/transcriptomes and then manipulate their genomes.
Termites are emerging model social organisms characterized by a sophisticated caste system. We recently sequenced
the genome of the subterranean termite Reticulitermes
sepratus (Shigenobu et al., 2022). The analyses of ~1.0 Gb
genome and 15,591 genes revealed the significance of gene
duplication in social evolution. Gene duplication associated with caste-biased gene expression was prevalent in thetermite genome. The duplicate genes comprised diverse categories related to social functions, such as chemical communication, social immunity and defense, and they were often
expressed in a caste-specific organ. Gene duplication may
facilitates social evolution through regulatory diversification
leading to caste-biased expression and functional specialization.
Beyond insect genomics, we are involved in several
genome projects of emerging model organisms. We recently
published the genome information of a photosynthetic sea
slug. Some sea slugs take up chloroplasts from the algaethat they consume into their cells. These chloroplasts retain
their ability to perform photosynthetic activity within the
animal cells for several months, and thus provide them withphotosynthesis-derived nutrition. This process is called kleptoplasty. Our genome analysis of Plakobranchus ocellatus
revealed the chloroplast acquisition without gene transfers in
the photosynthetic sea slug (Maeda et al., 2021).

Publication List

〔Original papers〕

K., Ikuta, T., Tame, A., Yamaguchi, K., Shigenobu, S.,• Igawa-Ueda,
Hongo, Y., Takaki, Y., Fujikura, K., Maruyama, T., and Yoshida, T.

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•
•
•
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Caste development and symbiosis in the social
aphid Ceratovacuna japonica

Eusociality has evolved repeatedly among insect lineages
and represents one of the major transitions in evolution.
Molecular mechanisms underlying sociality have been extensively studied with hymenopteran lineages, such as ants,bees, and wasps, and termites. Some aphid species show
eusociality and important groups for better understandingof the eusocial evolution, since they are one of the primitive eusocial species that belong to Hemiptera. We focus
on the social aphid Ceratovacuna japonica, because werecently established the laboratory culture system. First,
we investigated the sterility regulation in the soldier casteof the aphids (Chung and Shigenobu, 2022). We foundthat soldiers, that are completely sterile, possess a pair ofovarioles, but soldiers’ ovarioles were small and lacking gastrulating embryos. We also found that soldier reproduction
is constrained by apoptosis in maternal nutritive cells andnecrosis in oocytes and embryos. C. japonica also contains
endosymbionts. The eusocial aphid C. japonica provides us
with a unique opportunity to study how symbiosis operates ina social setting. We are currently investigating the interfaceof symbiosis and sociality.

(2021). Symbiont Transmission onto the Cell Surface of Early Oocytes
in the Deep-sea Clam Phreagena okutanii. Zoolog. Sci. 38, 140–147.
DOI: 10.2108/zs200129
Kobayashi, Y., Katsuren, M., Hojo, M., Wada, S., Terashima, Y.,
Kawaguchi, M., Tokuda, G., Kinjo, K., and Shigenobu, S. (2022).
Taxonomic revision of Termitomyces species found in Ryukyu
Archipelago, Japan, based on phylogenetic analyses with three loci.
Mycoscience 63, 33–38. DOI: 10.47371/mycosci.2021.11.001
Korgaonkar, A., Han, C., Lemire, A.L., Siwanowicz, I., Bennouna, D.,
Kopec, R.E., Andolfatto, P., Shigenobu, S., and Stern, D.L. (2021). A
novel family of secreted insect proteins linked to plant gall development.
Curr. Biol. 31, 1836-1849.e12. DOI: 10.1016/j.cub.2021.01.104
Maeda, T., Takahashi, S., Yoshida, T., Shimamura, S., Takaki, Y.,Nagai, Y., Toyoda, A., Suzuki, Y., Arimoto, A., Ishii, H., Satoh, N.,
Nishiyama, T., Hasebe, M., Maruyama, T., Minagawa, J., Obokata, J.,
and Shigenobu, S. (2021). Chloroplast acquisition without the genetransfer in kleptoplastic sea slugs, Plakobranchus ocellatus. eLife 10,e60176. DOI: 10.7554/eLife.60176
Miyazaki, S., Fujiwara, K., Kai, K., Masuoka, Y., Gotoh, H., Niimi,T., Hayashi, Y., Shigenobu, S., and Maekawa, K. (2021). Evolutionarytransition of doublesex regulation from sex-specific splicing to malespecific transcription in termites. Sci. Rep. 11, 15992. DOI: 10.1038/
s41598-021-95423-7
Nozaki, T., Suetsugu, K., Sato, K., Sato, R., Takagi, T., Funaki, S.,
Ito, K., Kurita, K., Isagi, Y., and Kaneko, S. (2021). Development ofmicrosatellite markers for the geographically parthenogenetic stickinsect Phraortes elongatus (Insecta: Phasmatodea). Genes Genet. Syst.
96, 199–203. DOI: 10.1266/ggs.21-00022
Nozaki, T., and Matsuura, K. (2021). Oocyte resorption in termite
queens: Seasonal dynamics and controlling factors. J. Insect Physiol.
131, 104242. DOI: 10.1016/j.jinsphys.2021.104242
Nozaki, T., Hojo, M., Yashiro, T., and Matsuura, K. (2021). First
Record of the Soil-feeding Higher Termites (Isoptera: Termitidae) from
Okinawa Island, the Central Ryukyus, Japan. SOCIOBIOLOGY 68,e6159. DOI: 10.13102/sociobiology.v68i2.6159
Shigenobu, S., Hayashi, Y., Watanabe, D., Tokuda, G., Hojo, M.Y.,
Toga, K., Saiki, R., Yaguchi, H., Masuoka, Y., Suzuki, R., Suzuki, S.,
Kimura, M., Matsunami, M., Sugime, Y., Oguchi, K., Niimi, T., Gotoh,H., Hojo, M.K., Miyazaki, S., Toyoda, A., Miura, T., and Maekawa,
K. (2022). Genomic and transcriptomic analyses of the subterranean
termite Reticulitermes speratus: Gene duplication facilitates socialevolution. Proc. Natl. Acad. Sci. U.S.A. 119, e2110361119. DOI:10.1073/pnas.2110361119

T., Matsubara, H., Minamitani, F., Satoh, Y., Tozawa, S.,• Takeuchi,Moriyama, T., Maruyama, K., Suzuki, K.T., Shigenobu, S., Inoue, T.,••••

Tamura, K., Agata, K., and Hayashi, T. (2022). Newt Hoxa13 has an
essential and predominant role in digit formation during development
and regeneration. Development 149, dev200282. DOI: 10.1242/dev.200282
Tanaka, S., Hashimoto, K., Kobayashi, Y., Yano, K., Maeda, T.,Kameoka, H., Ezawa, T., Saito, K., Akiyama, K., and Kawaguchi, M.
(2022). Asymbiotic mass production of the arbuscular mycorrhizal
fungus Rhizophagus clarus. Commun. Biol. 5, 43. DOI: 10.1038/
s42003-021-02967-5
Tominaga, T., Miura, C., Sumigawa, Y., Hirose, Y., Yamaguchi, K.,
Shigenobu, S., Mine, A., and Kaminaka, H. (2021). Conservation andDiversity in Gibberellin-Mediated Transcriptional Responses Among
Host Plants Forming Distinct Arbuscular Mycorrhizal Morphotypes.
Front. Plant Sci. 12, 795695. DOI: 10.3389/fpls.2021.795695
Tong, X., Kanzaki, N., and Akimoto, S. (2021). First record of amermithid nematode (Nematoda: Mermithidae) parasitizing winged
females of gall-forming aphids (Hemiptera: Aphididae: Eriosomatinae).
Entomol. Sci. 24, 354–360. DOI: 10.1111/ens.12486
Yashiro, T., Tea, Y.-K., van der Wal, C., Nozaki, T., Mizumoto,N., Hellemans, S., Matsuura, K., and Lo, N. (2021). Enhanced57

National Institute for Basic Biology

Evolutionary Biology and Biodiversity

heterozygosity from male meiotic chromosome chains is superseded byhybrid female asexuality in termites. Proc. Natl. Acad. Sci. U.S.A. 118,•e2009533118. DOI: 10.1073/pnas.2009533118
Yoshida, Y., Shaikhutdinov, N., Kozlova, O., Itoh, M., Tagami, M.,Murata, M., Nishiyori-Sueki, H., Kojima-Ishiyama, M., Noma,S., Cherkasov, A., Gazizova, G., Nasibullina, A., Deviatiiarov, R.,
Shagimardanova, E., Ryabova, A., Yamaguchi, K., Bino, T., Shigenobu,S., Tokumoto, S., Miyata, Y., Cornette, R., Yamada, T.G., Funahashi,A., Tomita, M., Gusev, O., and Kikawada, T. (2022). High quality
genome assembly of the anhydrobiotic midge provides insights on asingle chromosome-based emergence of extreme desiccation tolerance.
NAR Genomics Bioinforma. 4, lqac029. DOI: 10.1093/nargab/lqac029〔Review articles〕

S. & Yorimoto, S. Aphid hologenomics: current status and• Shigenobu,
future challenges. Curr Opin Insect Sci4, 100882. DOI:10.1016/j.

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cois.2022.100882.
Shigenobu, S. (2022). Editorial overview: Insect genomics: Genomics
of emerging model insects. Curr. Opin. Insect Sci. 51, 100913. DOI:
10.1016/j.cois.2022.100913
Miura, T., Oguchi, K., Yamaguchi, H., Nakamura, M., Sato, D.,
Kobayashi, K., Kutsukake, N., Miura, K., Hayashi, Y., Hojo, M.,
Maekawa, K., Shigenobu, S., Kano, T., and Ishiguro, A. (2022).
Understanding of superorganisms: collective behavior, differentiationand social organization. Artif. Life Robot. 27, 204–212. DOI: 10.1007/
s10015-022-00754-x