High-level expression of Mastermind-like 2 contributes to aberrant activation of the NOTCH signaling pathway in human lymphomas

Abstract

Inappropriate activation of the NOTCH signaling pathway, for example, by activating mutations, contributes to the pathogenesis of various human malignancies. Here, we demonstrate that aberrant expression of an essential NOTCH coactivator of the Mastermind-like (MAML) family provides an alternative mechanism to activate NOTCH signaling in human lymphoma cells. We detected high-level MAML2 expression in several B cell-derived lymphoma types, including classical Hodgkin lymphoma (cHL) cells, relative to normal B cells. Inhibition of MAML-protein activity by a dominant negative form of MAML or by small hairpin RNAs targeting MAML2 in cHL cells resulted in downregulation of the NOTCH target genes HES7 and HEY1, which we identified as overexpressed in cHL cells, and in reduced proliferation. Furthermore, a NOTCH gene-expression signature in cHL cells confirmed their cell-autonomous NOTCH activity. Finally, in line with the essential role of MAML proteins for assembly and activity of the NOTCH transcriptional complex (NTC), we show that MAML-derived small-peptide constructs block NOTCH activity and disrupt NTC formation in vitro. These data strongly suggest direct targeting of the NTC as treatment strategy for NOTCH-dependent malignancies.

Notch is an essential upstream regulator of NF-κB and is relevant for survival of Hodgkin and Reed–Sternberg cells

“…3 Jagged1 is expressed in HRS cells and bystander cells in vivo, 3 and the Notch ligands of the Delta-like family (DLL1, DLL3, DLL4) and Jagged2 are expressed in cultured HRS cells to activate Notch signaling. 4 We further demonstrated that absence of the main Notch1 inhibitor Deltex1 and high-level expression of the Notch co-activator of the Mastermind-like family contribute to deregulated Notch signaling in HRS cells. 4,5 Notch1 activity reduces the expression of the B cell transcription factors E12 and E47, and EBF1 and induces the transcription of activated B cell factor 1 (ABF-1).…”

“…4 We further demonstrated that absence of the main Notch1 inhibitor Deltex1 and high-level expression of the Notch co-activator of the Mastermind-like family contribute to deregulated Notch signaling in HRS cells. 4,5 Notch1 activity reduces the expression of the B cell transcription factors E12 and E47, and EBF1 and induces the transcription of activated B cell factor 1 (ABF-1). 5 In addition, Notch1 binds to PAX5 in HRS cells, which might impair PAX5 function.…”

“…In addition, we recently demonstrated that all cultured HRS cells and the Burkitt lymphoma cell line BJAB express variable levels of Notch1 IC . 4 However, neither Notch1 nor Notch1 IC levels alone predict the functional activity of the Notch transcriptional complex. 4 Therefore, we determined the activity of the Notch transcriptional complex by analyzing Notch target gene expression (ABF-1 (refs 5,23) and HEY1 (ref.…”

“…4 However, neither Notch1 nor Notch1 IC levels alone predict the functional activity of the Notch transcriptional complex. 4 Therefore, we determined the activity of the Notch transcriptional complex by analyzing Notch target gene expression (ABF-1 (refs 5,23) and HEY1 (ref. 4) ), using semi-quantitative reverse transcription-PCR analysis (Figure 1a, lower panel).…”

“…4 Therefore, we determined the activity of the Notch transcriptional complex by analyzing Notch target gene expression (ABF-1 (refs 5,23) and HEY1 (ref. 4) ), using semi-quantitative reverse transcription-PCR analysis (Figure 1a, lower panel). Our previous data demonstrated that the E2A antagonist ABF-1 is overexpressed in HRS cells, and we identified ABF-1 as a Notch1-specific target gene using siRNA against Notch1.…”