Biology subjects

Inukai, T.

Publications and source records attributed to Inukai, T..

4 recordsLinked to original sources

bioRxiv2023

Covalent Targeting of Glutamate Cysteine Ligase to Inhibit Glutathione Synthesis

Dysregulated oxidative stress plays a major role in cancer pathogenesis and some types of cancer cells are particularly vulnerable to inhibiting cellular antioxidant capacity. Glutamate-cysteine ligase (GCL) is the first and rate-limiting step in the synthesis of the major cellular antioxidant glutathione (GSH). Developing a GCL inhibitor may be an attractive therapeutic strategy for certain cancer types that are particularly sensitive to oxidative stress. In this study, we reveal a cysteine-reactive covalent ligand EN25 that covalently targets an allosteric cysteine C114 on GCLM, the modifier subunit of GCL, leading to inhibition of GCL activity, lowering of cellular GSH levels, and impaired cell viability in ARID1A-deficient cancer cells that are particularly vulnerable to glutathione depletion, but not in ARID1A-positive cancer cells. Our studies uncover a novel potential ligandable site within GCLM that can be targeted to inhibit the GSH synthesis in cancer cells to target vulnerable cancer cell types.

biochemistry↗

bioRxiv2022

An excitatory-inhibitory fronto-cerebellar loop resolves the Stroop effect

The Stroop effect is a well-known behavioral phenomenon in humans that refers to robust interference between language and color information. Although this effect has long been studied, it remains unclear when the interference occurs and how it is resolved in the brain. By manipulating the verbality of stimulus perception and response generation, here we show that the Stroop effect occurs during perception of color-word stimuli and is resolved by a cross-hemispheric, excitatory-inhibitory functional loop involving the lateral prefrontal cortex and cerebellum. Humans performed a Stroop task and a control task in which the stimulus did not contain verbal information, and made a response either vocally or manually. The resolution of Stroop interference involved the lateral prefrontal cortex in the left hemisphere and the cerebellum in the right hemisphere, independently of whether the response was made vocally or manually. In contrast, such cross-hemispheric lateralization was absent during the non-verbal control task. Moreover, the prefrontal cortex amplified cerebellar activity, whereas the cerebellum suppressed prefrontal activity, and these effects were enhanced during interference resolution. These results suggest that this fronto-cerebellar loop involving language and cognitive systems regulates goal-relevant information to resolve the interference occurring during simultaneous perception of a word and color.

neuroscience↗

bioRxiv2021

Detection and phylogeny of Wolbachia in field-collected Aedes albopictus and Aedes aegypti from Manila City, Philippines

BackgroundWolbachia is the most common bacterial endosymbiont of arthropods, such as the medically important Aedes albopictus and recent reports also detected in Aedes aegypti. Our study showed additional support for the presence of wolbachia in Ae. albopictus and Ae. aegypti using Wolbachia specific markers, wsp, and 16S. Main textThis study collected 12 adult Ae. albopictus and 359 Ae. aegypti from 183 households in a dengue-prone area, Manila, Philippines, between June and September 2017. Aedes larvae (n = 509) were also collected from 17 water containers from 11 households. The DNA of the Aedes larvae and adults were screened for the presence of Wolbachia using the wsp and 16S markers, following optimized polymerase chain reaction (PCR) conditions, and sequenced. Our results showed that 3 out of 359 (0.84%) adult Ae. aegypti and 12 out of 12 (100%) adult Ae. albopictus were Wolbachia positive, whereas all larvae tested negative for Wolbachia (0/509; 0%). The wsp marker revealed six Wolbachia-positive Ae. albopictus, whereas the 16S marker showed Wolbachia in three Ae. aegypti and ten Ae. albopictus ConclusionOur results suggest that the utilization of two Wolbachia specific markers, wsp and 16S demonstrated Wolbachia presence in individual Ae. albopictus and Ae. aegypti. The results of the Wolbachia infection in Ae. albopictus showed that the detected strains were from either supergroups A and B. Despite the low infection rate of Wolbachia in Ae. aegypti, our results demonstrated the presence of Wolbachia in field-collected Ae. aegypti supporting previous studies.

microbiology↗

bioRxiv2018

Cas9/CRISPR genome editing to demonstrate the contribution of Cyp51A Gly138Ser to azole resistance in Aspergillus fumigatus

Azole resistance in Aspergillus fumigatus is predominantly associated with increased expression of Cyp51A (lanosterol 14-demethylase), the target enzyme of azole antifungal agents, or with single-nucleotide polymorphisms (SNPs) in cyp51A. Although several SNPs that may be linked to low susceptibility in azole-resistant isolates have previously been reported, few studies have been conducted to conclusively demonstrate the contribution of SNPs to decreased azole susceptibility. An A. fumigatus strain was isolated from the sputum of a 74-year-old male receiving long-term voriconazole treatment for chronic progressive pulmonary aspergillosis. Etest antifungal susceptibility testing showed low susceptibility to voriconazole, itraconazole, and posaconazole. Nucleotide sequencing of cyp51A from this isolate revealed the mutations Gly138Ser (GGC[->]AGC) and Asn248Lys (AAT[->]AAA) compared with the cyp51A of azole-susceptible isolates. PCR-amplified DNA fragments containing cyp51A with or without the mutations of interest and a hygromycin marker were simultaneously introduced along with the Cas9 protein and in vitro-synthesized single-guide RNA into protoplasts of the azole-resistant/susceptible strains. Etest azole susceptibility testing of recombinant strains showed an increased susceptibility via the replacement of Ser138 by glycine. In contrast, azole susceptibility was slightly decreased when a Ser138 mutation was introduced into the azole-susceptible strain AfS35, indicating that the serine at position 138 of Cyp51A contributes to low susceptibility in the azole-resistant isolate. Genetic recombination, which has been hampered thus far in clinical isolates, can now be achieved using Cas9/CRISPR genome editing. This technique could be useful to investigate the contribution of other SNPs of cyp51A to azole resistance.

microbiology↗