摘要
Background : Iron-sulfur cluster assembly 1( ISCA 1) is an iron-sulfur(Fe/S) carrier protein that accepts Fe/S from a scaffold protein and transfers it to target proteins including the mitochondrial Fe/S containing proteins. ISCA 1 is also the newly identified causal gene for multiple mitochondrial dysfunctions syndrome( MMDS). However, our knowledge about the physiological function of ISCA 1 in vivo is currently limited. In this study, we generated an ISCA 1 knockout rat line and analyzed the embryo development. Methods : ISCA 1 knockout rats were generated by replacing the exon1 of ISCA 1 gene with the mC herry-Cre fusion gene using CRISPR-Cas9 technology. The ISCA 1 expression pattern was analyzed by fluorescence imaging using ISCA 1 promotor driven Cre and mC herry expression. The embryonic morphology was examinated by microscope and mitochondrial proteins were tested by Western blot. Results : An ISCA 1 knockout rat line was obtained, which expressed mC herry-Cre fusion protein. Both of the fluorescence images from mC herry and Cre induced mC herry in a reporter rat strain, showing that ISCA 1 expressed in most of the tissues in rats. The ISCA 1 knockout resulted in abnormal development at 8.5 days, with a significant decrease of NDUFA 9 protein and an increase of aconitase 2( ACO 2) in rat embryos. Conclusion : Deletion of ISCA 1 induced early death in rats. ISCA 1 affected the expression of key proteins in the mitochondrial respiratory chain complex, suggesting that ISCA 1 has an important influence on the respiratory complex and energy metabolism.
Background : Iron-sulfur cluster assembly 1( ISCA 1) is an iron-sulfur(Fe/S) carrier protein that accepts Fe/S from a scaffold protein and transfers it to target proteins including the mitochondrial Fe/S containing proteins. ISCA 1 is also the newly identified causal gene for multiple mitochondrial dysfunctions syndrome( MMDS). However, our knowledge about the physiological function of ISCA 1 in vivo is currently limited. In this study, we generated an ISCA 1 knockout rat line and analyzed the embryo development. Methods : ISCA 1 knockout rats were generated by replacing the exon1 of ISCA 1 gene with the mC herry-Cre fusion gene using CRISPR-Cas9 technology. The ISCA 1 expression pattern was analyzed by fluorescence imaging using ISCA 1 promotor driven Cre and mC herry expression. The embryonic morphology was examinated by microscope and mitochondrial proteins were tested by Western blot. Results : An ISCA 1 knockout rat line was obtained, which expressed mC herry-Cre fusion protein. Both of the fluorescence images from mC herry and Cre induced mC herry in a reporter rat strain, showing that ISCA 1 expressed in most of the tissues in rats. The ISCA 1 knockout resulted in abnormal development at 8.5 days, with a significant decrease of NDUFA 9 protein and an increase of aconitase 2( ACO 2) in rat embryos. Conclusion : Deletion of ISCA 1 induced early death in rats. ISCA 1 affected the expression of key proteins in the mitochondrial respiratory chain complex, suggesting that ISCA 1 has an important influence on the respiratory complex and energy metabolism.
基金
CAMS Innovation Fund for Medical Sciences,Grant/Award Number:2017-I2M-3-015 and 2016-I2M-1-004
