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首页> 外文期刊>Annals of Human Genetics >Molecular and cytological investigations of phosphoglucomutase (PGM1) in the K562 cell line.
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Molecular and cytological investigations of phosphoglucomutase (PGM1) in the K562 cell line.

机译：K562细胞系中磷酸葡萄糖突变酶（PGM1）的分子和细胞学研究。

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Phosphoglucomutase 1 (PGM1) deficiency is a stable characteristic of the erythroleukaemic cell line, K562, whereas the activity of the isozymes of the other two PGM loci (PGM2 and PGM3) is slightly elevated. In this study the molecular basis of PGM1 deficiency was investigated by a combined approach utilising protein electrophoresis, immunodetection, cytogenetic techniques, and DNA and RNA analysis. Isoelectric focusing and activity staining confirmed that K562 has no detectable PGM1 activity. Immunoblot analysis of extracts, separated by isoelectric focusing, starch gel and SDS gel electrophoresis, using monospecific anti-PGM1 antibodies showed that K562 contained no detectable immunoreactive material. Karyotype analysis revealed the presence of two intact chromosomes 1 and a derivative chromosome 1, der(1)t(1;11), each of which carried a copy of the PGM1 gene as demonstrated by fluorescence in situ hybridization using a PGM1 cosmid as probe. Southern blot analysis using a PGM1 cDNA clone as probe suggested that the PGM1 genes had not been subject to any gross structural rearrangements. We were also able to determine that K562 is type PGM1 2+1+ by restriction endonuclease analysis of genomic DNA. Very low levels of PGM1 mRNA which appeared to be full length transcripts were detected in K562 using a reverse transcriptase PCR technique. We conclude that the most likely cause of PGM1 enzyme deficiency in K562 is abnormal regulation of transcription.
机译：磷酸葡萄糖突变酶1（PGM1）缺乏是红白血病细胞系K562的稳定特征，而其他两个PGM位点（PGM2和PGM3）的同工酶的活性略有提高。在这项研究中，通过使用蛋白质电泳，免疫检测，细胞遗传学技术以及DNA和RNA分析的组合方法研究了PGM1缺乏症的分子基础。等电聚焦和活性染色证实K562没有可检测的PGM1活性。使用单特异性抗PGM1抗体通过等电聚焦，淀粉凝胶和SDS凝胶电泳对提取物进行免疫印迹分析，结果表明K562不包含可检测的免疫反应物质。染色体核型分析显示存在两个完整的染色体1和一个衍生染色体1，der（1）t（1; 11），每个染色体都带有一个PGM1基因的拷贝，如使用PGM1粘粒作为探针的荧光原位杂交所证明。使用PGM1 cDNA克隆作为探针的Southern印迹分析表明，PGM1基因未经历任何总体结构重排。通过基因组DNA的限制性核酸内切酶分析，我们还能够确定K562是PGM1 2 + 1 +型。使用逆转录酶PCR技术在K562中检测到极低水平的PGM1 mRNA，似乎是全长转录本。我们得出结论，K562中PGM1酶缺乏的最可能原因是转录的异常调节。

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