Expression of the Musashi1 gene encoding the RNA-binding protein in human hepatoma cell lines

doi:10.1016/S0006-291X(02)00175-4

Biochemical and Biophysical Research Communications

Volume 293, Issue 1, 26 April 2002, Pages 150-154

https://doi.org/10.1016/S0006-291X(02)00175-4 Get rights and content

Abstract

Musashi1, a neural RNA-binding protein, plays an important role in regulating cell differentiation in precursor cells. Recently, expression of Musashi1 has been detected in human tumor tissues such as gliomas and melanomas, suggesting its involvement in oncogenic development. To determine any association between Musashi1 and the development of liver cancer, we investigated its gene expression in seven human hepatoma cell lines: HuH6, HuH7, Hep3B, SK-Hep1, HepG2, HLE, and HLF. Musashi1 mRNA expression was analyzed using the reverse-transcription polymerase chain reaction (PCR), and the PCR products were sequenced using a subcloning procedure. Musashi1 protein expression was analyzed in HuH7 and HepG2 cells by Western blot and immunofluorescence staining. Musashi1 mRNA was detected in the HuH6, HuH7, and Hep3B hepatoma cell lines, but not in the others. Sequencing of the PCR-amplified Musashi1 cDNA in these three cell lines showed the expected sequence of the human Musashi1 gene. Musashi1 protein expression was confirmed in HuH7 cells, which were positive for Musashi1 mRNA expression, but not in HepG2 cells. These results suggest that Musashi1 expression may be an important factor in the development of several types of carcinoma such as human hepatoma, and may be a useful molecular marker for tumor detection.

Section snippets

Materials and methods

Cell lines and culture. Seven human hepatoma cell lines, HuH6, HuH7, Hep3B, SK-Hep1, HLE, HLF, and HepG2, were used for the mRNA gene-expression analysis of Musashi1. The cells were seeded into 250 ml flasks and maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 2 mM l-glutamine, 100 units/ml penicillin, 100 units/ml streptomycin, and 10% fetal bovine serum at 37 °C in a humidified atmosphere containing 5% CO₂. All media and chemicals used in the cell cultures were

Musashi1 protein expression in fetal mouse liver

In mouse fetal liver at 14.5 gestational days, the Musashi1 protein was detected in the cytoplasm of early hepatocytes (Fig. 1).

Musashi1 mRNA expression in human hepatoma cell lines

The PCR-amplified product obtained using the Musashi1-specific primers produced clear bands of the predicted 542-bp size. Musashi1 gene expression was observed in three hepatoma cell lines (HuH6, HuH7, and Hep3B) but was not detected in the other four hepatoma cell lines (SK-Hep1, HepG2, HLE, and HLF) (Fig. 2). Sequencing of the PCR-amplified Musashi1 cDNA in the three

Discussion

The Musashi1 gene, which encodes a neural RNA-binding protein, is closely associated with cell differentiation in CNS stem cells [1], [2], [3], [4], [5]. In the endodermal cells, Musashi1 is known to be expressed in intestinal epithelial stem cells [16]. We demonstrated in this study that, in the mouse, Musashi1 is expressed in early hepatocytes during the embryonic stage of liver development. Abnormalities of cell-regulatory gene expression may be involved in carcinogenesis; however, little is

Acknowledgements

We thank Dr. Hiroshi Kawai (Kanazawa University School of Medicine, Japan), and Dr. Tadahisa Fukui (Yamagata University School of Medicine, Japan) for the kind support of this work.

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^☆: This study was supported by a grant-in-aid from the Japan Society for the Promotion of Science (grant number: 13470112). The nucleotide sequence data reported in this paper appear in the DDBJ/EMBL/GenBank nucleotide sequence database with the accession numbers AB072590–AB072592.

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Expression of the Musashi1 gene encoding the RNA-binding protein in human hepatoma cell lines☆

Abstract

Section snippets

Materials and methods

Musashi1 protein expression in fetal mouse liver

Musashi1 mRNA expression in human hepatoma cell lines

Discussion

Acknowledgements

Dev. Biol.

Genomics

Differentiation

Curr. Opin. Cell. Biol.

J. Hepatol.

Hepatology

Semin. Cancer Biol.

Neuron

J. Neurosci.

Dev. Neurosci.