The expression of the human myeloid cell nuclear differentiation antigen (MNDA) was observed specifically in cells of the granulocyte-macrophage lineage in our earlier reports. The specificity of MNDA expression for cells in the granulocyte-macrophage lineage was reexamined in cell lines established from patients with Philadelphia chromosome-positive chronic myeloid leukemia. Cell lines that expressed MNDA exhibited myeloid cell features and granulocyte or monocyte differentiation could be induced in vitro, while cell lines exhibiting properties of very early stage cells or multipotential cells did not express MNDA. Cells originating from cases of Burkitt's lymphoma were negative. By contrast, three lymphoblastoid cell lines (immortalized in vitro with Epstein-Barr virus) were weakly positive and MNDA was up-regulated by interferon-alpha (IFN-alpha) treatment. As we reported previously, MNDA mRNA level in adherent monocytes is elevated by IFN-alpha; in this study, we further assessed MNDA expression in in vitro monocyte-derived macrophages. Three additional agents (endotoxin, phytohemagglutinin, and phorbol ester) and other conditions that affect function, cytokine production, differentiation, and/or growth of monocytes were examined for their ability to alter MNDA expression. The results varied with the agent, cell type, and stage of differentiation. Changes in MNDA expression occurred slowly (hours to days), suggesting that MNDA could mediate changes realized over a long period. The results also reveal a discordance in certain MNDA positive cells between steady-state levels or changes in levels of protein and mRNA indicating that the regulation of MNDA expression occurs at more than one point. Changes in MNDA expression are consistent with a role in opposing macrophage differentiation and activation of monocytes/macrophages.

The human myeloid cell nuclear differentiation antigen (MNDA) is expressed constitutively in cells of the myeloid lineage, appearing in myeloblast cells in some cases of acute myeloid leukemia and consistently being detected in promyelocyte stage cells as well as in all later stage cells including peripheral blood monocytes and granulocytes. The human myeloid leukemia cell lines, HL-60, U937, and THP-1, express similar levels of immunochemically detectable MNDA. Although, the level of MNDA mRNA in primary monocytes is very low it was up-regulated at 6 h following the addition of interferon alpha. The effect of interferon alpha on the MNDA mRNA is also observed in the cell lines HL-60, U937, and THP-1. The MNDA mRNA level in primary granulocytes was unaffected by addition of interferon alpha and other agents including interferon gamma, endotoxin, poly (I).poly (C), and FMLP. The MNDA mRNA level in the myeloid cell lines was also unaffected by the latter four agents. Induction of differentiation in the myeloid cell lines with phorbol ester induces monocyte differentiation which was accompanied by a decrease in MNDA mRNA level. This reduced level of mRNA could then be elevated with subsequent interferon alpha treatment. The effects of phorbol ester on MNDA mRNA appeared to be associated with induced differentiation since inhibiting cell proliferation did not alter the level of MNDA mRNA and cell cycle variation in MNDA mRNA levels were not observed. The ability of interferon alpha to up-regulate MNDA mRNA in phorbol ester treated myeloid cell lines is consistent with the observations made in primary monocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

We have previously shown that the human myeloid cell nuclear differentiation antigen (MNDA) is expressed at both the antigen and mRNA levels specifically in human monocytes and granulocytes and earlier stage cells in the myeloid lineage. A 200 amino acid region of the MNDA is strikingly similar t o a region in the proteins encoded by a family of interferon-inducible mouse genes, designated Mi-201, Mi-202, K-203, etc, that are not regulated in a cell-or tissue-specific fashion. However, a new member of the lfi-ZOO gene family, 03, is induced in mouse mononuclear phagocytes but not in fibroblasts by interferon. The same 200 amino acid region, duplicated in the mouse Mi-ZOO gene family, is also repeated in the recently characterized human IF1 16 gene that is constitutively expressed specifically in lymphoid cells and is induced in myeloid cells by interferon y. The 1.8-kb MNDA mRNA, which contains an interferon-stimulated response HE HUMAN MYELOID cell nuclear differentiation antigen (MNDA) is detected only in nuclei of cells of the granulocyte-monocyte lineage.' We also evaluated MNDA expression in 21 cases of human acute leukemia classified by French-American-British (FAB) Group criteria.2 Cases in the FAB M2, M3, M4, and M5 categories, in which myeloid cell maturation is evident, were all positive with variability in percentage of positive cells and in the intensity of the staining reactions. In five cases of acute myeloblastic leukemia without maturation (FAB MI), three were negative and two were positive. The four cases of acute lymphocytic leukemia were all negative. Two cases of biphenotypic acute leukemia and one case of lymphoid blast crisis of chronic granulocytic leukemia were also negative.2 These results indicated that MNDA expression is correlated with granulocyte and monocyte differentiation in cases of acute leukemia. Our MNDA amino acid and cDNA sequence a n a l y~i s~.~ provided insight into its functional significance. The strict nuclear localization of the MNDA in interphase cells is consistent with its containing sequence motifs commonly found in gene regulatory protein^.^ In addition, DNA binding activity of the MNDA was previously demonstrated using DNAprotein cross-linking.5 MNDA contains extended regions of sequence at both the DNA and protein levels similar to a class of mouse genes cluster) that are interferon-ind~cible.~.~ The mouse @-202 and @-204 genes each encode contiguous 200 amino acid (aa) regions that are highly conserved within each protein and between proteins in the family. The recently characterized mouse D3 gene, a new member of the @-ZOO gene family, contains only a single copy of the 200-aa conserved sequence and exhibits a sequence similar to the I$-204 gene outside of the 200-aa conserved sequence that is unique from other members of the @-200 gene family.' The MNDA also contains only a single copy of the 200-aa conserved sequence and more than half of the 67 residues of its NH2 terminal sequence are identical to the NH2 terminal domain of the mouse 204 protein.' In the ...

We have previously shown that the human myeloid cell nuclear differentiation antigen (MNDA) is expressed at both the antigen and mRNA levels specifically in human monocytes and granulocytes and earlier stage cells in the myeloid lineage. A 200 amino acid region of the MNDA is strikingly similar to a region in the proteins encoded by a family of interferon-inducible mouse genes, designated Ifi-201, Ifi- 202, Ifi-203, etc, that are not regulated in a cell- or tissue-specific fashion. However, a new member of the Ifi-200 gene family, D3, is induced in mouse mononuclear phagocytes but not in fibroblasts by interferon. The same 200 amino acid region, duplicated in the mouse Ifi- 200 gene family, is also repeated in the recently characterized human IFI 16 gene that is constitutively expressed specifically in lymphoid cells and is induced in myeloid cells by interferon gamma. The 1.8-kb MNDA mRNA, which contains an interferon-stimulated response element in the 5′ untranslated region, was significantly upregulated in human monocytes exposed to interferon alpha. Characterization of the MNDA gene showed that it is a single-copy gene and localized to human chromosome 1q 21–22 within the large linkage group conserved between mouse and human that contains the Ifi-200 gene family. The IFI 16 gene is also located on human chromosome 1q. Our observations are consistent with the proposal that the MNDA is a member of a cluster of related human interferon-regulated genes, similar to the mouse Ifi-200 gene family. In addition, one mouse gene in the Ifi-200 gene family and the human MNDA and IFI 16 genes show expression and/or regulation restricted to cells of the hematopoietic system, suggesting that these genes participate in blood cell-specific responses to interferons.

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