Inhibitor of differentiation 4 (ID4) acts as an inhibitor of ID-1, -2 and -3 and promotes basic helix loop helix (bHLH) E47 DNA binding and transcriptional activity

Abstract

The four known ID proteins (ID1-4, Inhibitor of Differentiation) share a homologous helix loop helix (HLH) domain and act as dominant negative regulators of basic-HLH transcription factors. ID proteins also interact with many non-bHLH proteins in complex networks. The expression of ID proteins is increasingly observed in many cancers. Whereas ID-1, ID-2 and ID-3, are generally considered as tumor promoters, ID4 on the contrary has emerged as a tumor suppressor. In this study we demonstrate that ID4 heterodimerizes with ID-1, -2 and -3 and promote bHLH DNA binding, essentially acting as an inhibitor of inhibitors of differentiation proteins. Interaction of ID4 was observed with ID1, ID2 and ID3 that was dependent on intact HLH domain of ID4. Interaction with bHLH protein E47 required almost 3 fold higher concentration of ID4 as compared to ID1. Furthermore, inhibition of E47 DNA binding by ID1 was restored by ID4 in an EMSA binding assay. ID4 and ID1 were also colocalized in prostate cancer cell line LNCaP. The alpha helix forming alanine stretch N-terminal, unique to HLH ID4 domain was required for optimum interaction. Ectopic expression of ID4 in DU145 prostate cancer line promoted E47 dependent expression of CDKNI p21. Thus counteracting the biological activities of ID-1, -2 and -3 by forming inactive heterodimers appears to be a novel mechanism of action of ID4. These results could have far reaching consequences in developing strategies to target ID proteins for cancer therapy and understanding biologically relevant ID- interactions.

Id4 eliminates the pro-activation factor Ascl1 to maintain quiescence of adult hippocampal stem cells

“…It is well established that Id proteins, including Id4, form non-functional heterodimers with E proteins, which are dimerization partners of tissue-specific bHLH transcription factors such as Ascl1 (Imayoshi and Kageyama, 2014; Ling et al, 2014; Patel et al, 2015; Sharma et al, 2015). High levels of Ids are therefore expected to result in the sequestration of E-proteins away from functional dimers with Ascl1.…”

“…Id4 has been proposed to act differently from other members of the Id family (Boareto et al, 2017; Patel et al, 2015; Sharma et al, 2015). Accordingly, the function that we have identified for Id4 in the hippocampus is also distinct from the role reported for other Id factors in the adult ventricular-subventricular zone (V-SVZ), where Id1 and Id3 promote stem cell self-renewal (Nam and Benezra, 2009) and Id1-3 maintain stem cell function by keeping stem cells adherent to their niche environment (Niola et al, 2012).…”

“…Therefore, Id proteins promote a switch of the expression pattern of Hes proteins from oscillatory, resulting in oscillatory expression of target genes such as Ascl1, to stably high, resulting in constant repression of these targets (Bai et al, 2007; Boareto et al, 2017). Id4 can inhibit the action of Id1-3 proteins by interacting with stronger affinity with them than with the E-proteins (Patel et al, 2015; Sharma et al, 2015). This non-canonical role of Id4 has been proposed to explain how Id4 promotes the proliferation of embryonic neural progenitors (Bedford et al, 2005; Yun et al, 2004) by blocking the interaction of Id1-3 with Hes proteins and thus promoting Hes protein oscillations and progenitor proliferation (Boareto et al, 2017).…”

ID4 controls luminal lineage commitment in normal mammary epithelium and inhibits BRCA1 function in basal-like breast cancer

“…The ID4-mediated inhibition of ELF5 occurs indirectly via Notch signalling (Junankar et al 2015), while ID4 has been shown to inhibit ERα, FOXA1 and BRCA1 expression by direct interaction with the promoters of these genes (Beger et al 2001, Best et al 2014. As ID4 does not have a basic DNA-binding domain (reviewed in Sharma et al 2015), it is likely that it regulates gene expression by forming a complex with other transcriptional regulators. The identity of these transcriptional regulators is currently unknown.…”

Induction of Lumen Formation in a Three-dimensional Model of Mammary Morphogenesis by Transcriptional Regulator ID4

“…Because ID2 plays an important role in mammary gland development (41,42), but may be oncogenic when not switched off (9), its regulation by ID4 is a real possibility. Along this line, Sharma et al (10) have recently shown that ID4 can switch off ID1-3 inhibition by forming heterodimers, thus switching on their targets. Thus, it may be necessary to identify their targets, rather than the targets of ID4.…”