Generating the recombinant mouse Aldehyde dehydrogenase 1A1 (ALDH1A1) with an N-terminal 6xHis-tag involves cloning the target gene, fused with the 6xHis-tag sequence, into a plasmid vector and transforming the yeast cells. The target gene encodes the 2-501aa of the mouse Aldh1a1. The cells are cultured and induced for protein expression, followed by cell lysis. Nickel affinity chromatography is used for protein purification, capitalizing on the His-tag's affinity for nickel. The protein's purity is evaluated using SDS-PAGE, exceeding 90%.
ALDH1A1 is a vital enzyme responsible for oxidizing aldehydes to their corresponding acids [1]. This property can deactivate essential chemotherapeutic agents [2]. ALDH1A1 is involved in various biological processes such as UV-radiation resistance, melanogenesis, and stem cell maintenance [3]. ALDH1A1 has been associated with tumor growth and drug resistance in lung adenocarcinoma and triple-negative breast carcinoma [2][4]. It is linked to poor differentiation, 'right-sidedness,' and reduced survival in colorectal cancer [5]. ALDH1A1 is a marker for malignant prostate stem cells and has predictive value for prostate cancer patient outcomes [9].
Furthermore, ALDH1A1 is implicated in retinoic acid synthesis, particularly in the developing retina, where it regulates gene expression involved in tumor-initiating stem-like cells [6][4]. In the nigrostriatal dopamine system, ALDH1A1 is selectively expressed by dopaminergic neurons and mitigates dyskinesia through transsynaptic retinoic acid signaling [7][8].
References:
[1] B. Wang, X. Chen, Z. Wang, W. Xiong, T. Xu, X. Zhaoet al., Aldehyde dehydrogenase 1a1 increases nadh levels and promotes tumor growth via glutathione/dihydrolipoic acid-dependent nad+ reduction, Oncotarget, vol. 8, no. 40, p. 67043-67055, 2017. https://doi.org/10.18632/oncotarget.17688
[2] Y. Wei, S. Wu, W. Xu, Y. Liang, Y. Li, W. Zhaoet al., Depleted aldehyde dehydrogenase 1a1 (aldh1a1) reverses cisplatin resistance of human lung adenocarcinoma cell a549/ddp, Thoracic Cancer, vol. 8, no. 1, p. 26-32, 2016. https://doi.org/10.1111/1759-7714.12400
[3] E. Paterson, H. Ho, R. Kapadia, & A. Ganesan, 9‐cis retinoic acid is the aldh1a1 product that stimulates melanogenesis, Experimental Dermatology, vol. 22, no. 3, p. 202-209, 2013. https://doi.org/10.1111/exd.12099
[4] M. Mohamed, B. Ibrahim, S. Sheikh, & M. Magid, Stem cell marker aldehyde dehydrogenase 1a1 expression in triple-negative breast carcinoma, Open Access Macedonian Journal of Medical Sciences, vol. 10, no. A, p. 287-294, 2022. https://doi.org/10.3889/oamjms.2022.8439
[5] L. Waals, I. Rinkes, & O. Kranenburg, Aldh1a1 expression is associated with poor differentiation, ‘right-sidedness’ and poor survival in human colorectal cancer, Plos One, vol. 13, no. 10, p. e0205536, 2018. https://doi.org/10.1371/journal.pone.0205536
[6] X. Fan, A. Molotkov, S. Manabe, C. Donmoyer, L. Deltour, M. Foglioet al., Targeted disruption of aldh1a1 (raldh1) provides evidence for a complex mechanism of retinoic acid synthesis in the developing retina, Molecular and Cellular Biology, vol. 23, no. 13, p. 4637-4648, 2003. https://doi.org/10.1128/mcb.23.13.4637-4648.2003
[7] J. Pan, J. Yu, L. Sun, C. Xie, L. Chang, J. Wuet al., Aldh1a1 regulates postsynaptic μ–opioid receptor expression in dorsal striatal projection neurons and mitigates dyskinesia through transsynaptic retinoic acid signaling, Scientific Reports, vol. 9, no. 1, 2019. https://doi.org/10.1038/s41598-019-40326-x
[8] K. Carmichael, R. Evans, E. Lopez, L. Sun, M. Kumar, J. Dinget al., Function and regulation of aldh1a1-positive nigrostriatal dopaminergic neurons in motor control and parkinson’s disease, Frontiers in Neural Circuits, vol. 15, 2021. https://doi.org/10.3389/fncir.2021.644776
[9] T. Li, Y. Su, Y. Mei, Q. Leng, B. Leng, Z. Liuet al., Aldh1a1 is a marker for malignant prostate stem cells and predictor of prostate cancer patients' outcome, Laboratory Investigation, vol. 90, no. 2, p. 234-244, 2010. https://doi.org/10.1038/labinvest.2009.127
