著者

Karla Cristine C. DOYSABAS Mami OBA Tomoki ISHIBASHI Hideki SHIBATA Hitoshi TAKEMAE Hiroshi SHIMODA Tetsuya MIZUTANI Atsuo IIDA Eiichi HONDO

出版者

JAPANESE SOCIETY OF VETERINARY SCIENCE

雑誌

Journal of Veterinary Medical Science (ISSN:09167250)

巻号頁・発行日

pp.20-0021, (Released:2020-02-12)

被引用文献数

2

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Adenosine 5’-triphosphate (ATP), the major energy currency of the cell, is involved in many cellular processes, including the viral life cycle, and can be used as an indicator of early signs of cytopathic effect (CPE). In this study, we demonstrated that CPE can be analyzed using an FRET-based ATP probe named ATP indicator based on Epsilon subunit for Analytical Measurements (ATeam). The results revealed that as early as 3 hr, the virus infected cells showed a significantly different Venus/cyan fluorescent protein (CFP) ratio compared to the mock-infected cells. The ATeam technology is therefore useful to determine the early signs of ATP-based CPE as early as 3 hr without morphology-based CPE by light microscopy, and enables high throughput determination of the presence of microorganisms in neglected samples stored in laboratories.

著者

Atsuo Iida Zi Wang Atsuko Sehara-Fujisawa

出版者

The Genetics Society of Japan

雑誌

Genes & Genetic Systems (ISSN:13417568)

巻号頁・発行日

pp.19-00033, (Released:2019-10-01)

参考文献数

15

被引用文献数

3

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Integrins, transmembrane molecules that facilitate cell-to-cell and cell-to-extracellular matrix interactions, are heterodimers that consist of an α- and β-subunit. The integrin α4 gene (itgα4) is expressed in various type of cells and tissues. Its biochemical functions and physiological roles have been revealed using cultured cell assays. In contrast, the primary effect caused by itgα4 deletion on vertebrate development is poorly understood, because knockout mice exhibit multiple defects that can lead to embryonic lethality in the uterus. Zebrafish are a convenient vertebrate model to investigate morphogenesis during embryogenesis, because of their external fertilization and subsequent development outside the female's body. Here, we generated a zebrafish mutant line named itgα4 ko108 using the CRISPR/Cas9 genome editing system; the mutant genome harbored an approximately 2.0-kb deletion in the itgα4 locus. A truncated transcript was detected in itgα4 (+/-) or (-/-) fish but not in (+/+) fish. The mutant transcript was hypothesized to encode a truncated Itgα4 protein due to a premature stop codon. itgα4 (-/-) embryos obtained from the mating of heterozygous parents exhibited no apparent phenotype during development at 24 hours post-fertilization (hpf). However, approximately half of them exhibited cephalic hemorrhage at 48 hpf. The incidence ratio was significantly higher than that in (+/+) or (+/-) embryos. Embryonic hemorrhage has also been reported previously in Itgα4 knockout mice. In contrast, embryonic lethality with the other defects reported in the knockout mice was not observed in our zebrafish model. Therefore, the mutant line itgα4 ko108 should be a useful model to investigate a physiological function for Itgα4 in the blood circulation system.