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A PU.1 suppressive target gene, metallothionein 1G, inhibits retinoic acid-induced NB4 cell differentiation.

Hirako N1, Nakano H2, Takahashi S3.

PLoS One. 2014 ;9(7):e103282.

1Division of Molecular Hematology, Kitasato University Graduate School of Medical Sciences, Minami-ku, Sagamihara, Japan.and

2Division of Hematology, Kitasato University School of Allied Health Sciences, Minami-ku, Sagamihara, Japan.and

3Division of Molecular Hematology, Kitasato University Graduate School of Medical Sciences, Minami-ku, Sagamihara, Japan; Division of Hematology, Kitasato University School of Allied Health Sciences, Minami-ku, Sagamihara, Japan.

 

Abstract

We recently revealed that myeloid master regulator SPI1/PU.1 directly represses metallothionein (MT) 1G through its epigenetic activity of PU.1, but the functions of MT1G in myeloid differentiation remain unknown. To clarify this, we established MT1G-overexpressing acute promyelocytic leukemiaNB4 (NB4MTOE) cells, and investigated whether MT1G functionally contributes to all-trans retinoic acid (ATRA)-induced NB4 cell differentiation. Real-time PCR analyses demonstrated that the inductions of CD11b and CD11c and reductions in myeloperoxidase and c-myc by ATRA were significantly attenuated in NB4MTOE cells. Morphological examination revealed that the percentages of differentiated cells induced by ATRA were reduced in NB4MTOE cells. Since G1 arrest is a hallmark of ATRA-induced NB4 cell differentiation, we observed a decrease in G1 accumulation, as well as decreases in p21WAF1/CIP1 and cyclin D1 inductions, by ATRA in NB4MTOE cells. Nitroblue tetrazolium (NBT) reduction assays revealed that the proportions of NBT-positive cells were decreased in NB4MTOE cells in the presence of ATRA. Microarray analyses showed that the changes in expression of several myeloid differentiation-related genes (GATA2, azurocidin 1, pyrroline-5-carboxylate reductase 1, matrix metallopeptidase -8, S100 calcium-binding protein A12, neutrophil cytosolic factor 2 and oncostatin M) induced by ATRA were disturbed in NB4MTOE cells. Collectively, overexpression of MT1G inhibits the proper differentiation of myeloid cells.

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Significance Statement

As we recently revealed that MT1G is a direct suppressive target gene of PU.1, the series of findings led to the discovery that PU.1 not only controls myeloid differentiation through positive regulations of myeloid genes (i.e. G-CSFR, GM-CSFR etc), but also through negative regulations of MT1G.

Furthermore, current study indicates that expression analysis of MT1G in acute promyelocytic leukemia patients may be a good prediction marker to estimate the efficacy of ATRA.

 

A PU.1 Suppressive Target Gene, Metallothionein 1G, Inhibits Retinoic Acid-Induced NB4 Cell Differentiation