FY2022 Annual Report

Immune Signal Unit
Associate Professor Hiroki Ishikawa

Abstract

Research in our unit has focused on molecular mechanisms of differentiation of immune cells. We are particularly interested in transcriptional regulatory mechanisms of T cell differentiation. Our recent data demonstrate important roles for the AP-1 transcription factor JunB in the functional diversification of T cell subsets. In this fiscal year, we reported that a glycolytic intermediate metabolite, phosphoenolpyruvate, inhibits the JunB-dependent Th17 transcriptional program and autoimmune diseases. Additionally, in collaboration with the Naha Medical Association, our study revealed that COVID-19 vaccine response is inversely correlated with baseline expression of AP-1 transcription factors in peripheral blood mononuclear cells. Furthermore, in another collaboration study with Shintake Unit, we demonstrated that ethanol vapor inhalation inhibits respiratory infection of influenza viruses.

1. Staff

Dr. Hiroki Ishikawa, Associate Professor
Dr. Rajukmar Singh, Staff Scientist
Dr. Miho Tamai, Staff Scientist
Ms. Mio Miyagi, Research Unit Technician
Dr. Daiki Sasaki, Research Unit Technician
Mr. Takeshi Toma Research Unit Technician
Mr. Tsung-Han Hsieh, Student
Ms. Shukla Sarker, Student
Mr. Ke Wang, Student
Mr. Tsung-Yen Huang, Student
Ms. Hsiao-Chiao Hsieh, Student
Mr. Charles Whitaker, Student
Ms. Mi Yang, Student
Mr. Masato Hirota, Student
Ms. Gulfiya Kadyrzhanova, Student
Ms. Sachie Yukawa, Student
Ms. Rika Yoshizawa, Research Unit Administrator

2. Collaborations

Identification of human and gut microbial parameters associated with COVID-19 vaccine responses

Description: Using blood and fecal samples collected from Okinawan subjects, we performed multiomics analyses and identified factors associated with COVID-19 vaccine responses.
Type of collaboration: Joint research
Researchers:
- Hiroaki Kitano (Integrated Open Systems Unit)
- Matthias Wolf (Molecular Electron Microscopy Unit)
- Dr. Mitsuo Kina (Kina Clinic)
- Dr. Osamu Tamai (Akebono Clinic)
- Dr. Kentaro Tamaki (Naha-nishi Clinic)
- Dr. Hiroaki Tomori (Yaesu Clinic)
- Dr. Masatake Miyagi (Arakawa Clinic)
- Dr. Chiaki Yamashiro (Yamashiro Orthopedic Surgery Ophthalmology Clinic)
- Dr. Eishin Sakihara (Lifestyle Related Disease Medical Center, Naha Medical Association)

3. Activities and Findings

3.1 Phosphoenolpyruvates regulates the Th17 transcriptional progr

We identified a glycolytic intermediate metabolite, phosphoenolpyruvate (PEP), as a negative regulator of Th17 differentiation (Huang et al., Cell Reports, 2023). Aerobic glycolysis, a metabolic pathway essential for effector T cell survival and proliferation, negatively regulates differentiation of autoimmune Th17 cells, but the mechanism underlying this regulation is largely unknown. We found that addition of PEP to cell culture media or inhibition of downstream glycolytic enzymes increases intracellular PEP and inhibits IL-17A expression in differentiating and mature Th17 cells. PEP supplementation does not significantly affect metabolic reprogramming, cell proliferation, or survival of T helper cells. Mechanistically, PEP binds to JunB and inhibits DNA binding of the JunB/BATF/IRF4 complex in a locus-specific manner, thereby regulating the Th17 transcriptional program. Furthermore, daily administration of PEP to mice inhibits generation of Th17 cells and ameliorates Th17-dependent autoimmune encephalomyelitis. These data demonstrate that PEP links aerobic glycolysis to the Th17 transcriptional program, suggesting the therapeutic potential of PEP for autoimmune diseases.

3.2 Identification of factors associated with COVID-19 vaccine responses

Using a systems biology approach based on multi-omics analyses of human blood and stool samples, we identified human and gut microbial parameters that are associated with COVID-19 vacine responses (Hirota et al., Commun. Biol., 2023). COVID-19 mRNA vaccines induce protective adaptive immunity against SARS-CoV-2 in most individuals, but there is wide variation in levels of vaccine-induced antibody and T-cell responses. However, factors associated with this inter-individual variation remain unclear. We found that baseline expression of AP-1 transcription factors, FOS and ATF3, is inversely correlated with BNT162b2 mRNA vaccine-induced T-cell responses. FOS expression is associated with transcription modules related to baseline immunity, but it is negatively associated with those related to T-cell activation upon BNT162b2 mRNA stimulation. Interestingly, the gut microbial fucose/rhamnose degradation pathway is positively correlated with FOS and ATF3 expression and inversely correlated with BNT162b2-induced T-cell responses. Taken together, these results suggest that gut microbial fucose/rhamnose degradation may promotes AP-1 expression in PBMCs, thereby inhibiting mRNA vaccine-induced T cell responses.

3.3 Ethanol vapor inhalation treatment for respiratory infectious diseases

In collaboration with Prof. Shintake at OIST, we have developed ethanol vapor inhalation treatment for respiratory infectious diseases (Tamai et al., J. Infect. Dis., 2023). Ethanol has been used as a disinfectant against enveloped viruses, including influenza and SARS-CoV-2. If ethanol can also inactivate these viruses in the respiratory tract without cytotoxicity, ethanol vapor inhalation could be a new therapy for various respiratory infections associated with enveloped viruses; however, there was no experimental evidence to support this treatment. We found that approximately 20% (v/v) ethanol solution, much lower than the minimum concentration required for conventional environmental surface disinfection, is sufficient to inactivate influenza A virus (IAV) at mammalian body temperatures. The minimum EtOH concentration required for IAV inactivation is inversely proportional to the reaction temperature, suggesting that thermal and chemical energies contribute cooperatively to viral inactivation. Furthermore, no cytotoxicity was observed when lung epithelial cells were exposed to ethanol solutions up to 22.5% (v/v) from the apical side, mimicking inhalation treatment with ethanol vapor. Based on these observations, we developed an ethanol vapor exposure system for mice, which is expected to expose the respiratory epithelial layer to about 20% (v/v) ethanol solution. Using this, we showed that inhalation administration of EtOH vapor does not cause obvious side effects and inhibits IAV infection in the lung, thereby protecting mice from lethal IAV respiratory infection (Fig. 4). These results demonstrate the potential of ethanol vapor inhalation as a novel therapeutic strategy against respiratory virus infection and suggest that the generation of about 20% (v/v) of EtOH solution in the respiratory tract is essential to the success of this treatment.

4. Publications

4.1 Journals

1. M. Tamai, S. Taba, T. Mise, M. Yamashita, H. Ishikawa, and T. Shintake, Effect of ethanol vapor inhalation treatment on lethal respiratory viral infection with Influenza A. J. Infect. Dis. doi: 10.1093/infdis/jiad089. Online ahead of print., 2023.
2. M. Hirota, M. Tamai, S. Yukawa, N. Taira, M. M. Matthews, T. Toma, T. Seto, M. Yoshida, S. Toguchi, M. Miyagi, T. Mori, H. Tomori, O. Tamai, M. Kina, E. Sakihara, C. Yamashiro, M. Miyagi, K. Tamaki, M. Wolf, M. K. Collins, H. Kitano, and H. Ishikawa, Human immune and gut microbial parameters associated with inter-individual variations in COVID-19 mRNA vaccine-induced immunity, Commun. Biol. 6, 368, 2023.
3. T-Y. Huang, M. Hirota, D. Sasaki, R. S. Kalra, H-C. Chien, M. Tamai, S. Sarkar, Y. Mi, M. Miyagi, Y. Seto, and H. Ishikawa, "Phosphoenolpyruvate regulates the Th17 transcriptional program and inhibits autoimmunity", Cell Rep. 28;42(3):112205, 2023.

4.2 Books and other one-time publications

Nothing to report

4.3 Oral and Poster Presentations

T-Y. Huang, M. Hirota, D. Sasaki, R. S. Kalra, H-C. Chien, M. Tamai, S. Sarkar, Y. Mi, M. Miyagi, Y. Seto, and H. Ishikawa. Phosphoenolpyruvate regulates the Th17 transcriptional program and inhibits autoimmunity. The 8th annual meeting of the Japanese Society for Living Systems Design Research, Tokyo, Japan from March 9-10 (2022)
M. Tamai, S. Taba, T. Mise, M. Yamashita, H. Ishikawa, and T. Shintake, Effect of ethanol vapor inhalation treatment on lethal respiratory viral infection with Influenza A. The 8th annual meeting of the Japanese Society for Living Systems Design Research, Tokyo, Japan from March 9-10 (2022)