The helix-loop-helix (HLH) protein Id lacks the basic DNA-binding domain common to this class of proteins. In vitro experiments suggested that Id could associate tightly with two other HLH proteins encoded by the E2A gene, El2 and E47 (referred to here collectively as E proteins) and prevent their binding to a sequence present in the muscle creatine kinase (MCK) enhancer either as homo-oligomers or hetero-oligomers with MyoD. In this report we present evidence for the in vivo roles of Id and E proteins: (1) Id and E proteins cofractionate and coimmunoprecipitate in whole-cell extracts prepared from myoblasts; (2) the loss of Id protein observed during the conversion of proliferating myoblasts into mature myotubes correlates with the formation of MyoD/E hetero-oligomeric complexes in whole-cell extracts (these complexes do not form when purified Id protein is added to the extracts); and (3) stable overexpression of Id mRNA and protein in the C2C12 muscle cell line inhibits differentiation in these cells 16 hr postinduction. The myotubes that do eventually form 48 hr postinduction have no detectable Id protein in the nucleus despite the persistence of exogenous Id mRNA. These data support a model in which Id can inhibit muscle cell differentiation by associating with E proteins and preventing them from forming active hetero-oligomeric complexes with the muscle determination gene products.

The murine dominant negative helix-loop-helix (dnHLH) proteins inhibit the activities of bHLH transcription factors in diverse cell lineages (Benezra et al. . Currently, there are four members in the dnHLH family, Idl, Id2, Id3, and Id4. In this report, we have performed a detailed comparative in situ hybridization analysis to examine their expression pattern during post-gastrulational mouse development. Idl, 2, and 3 are expressed in multiple tissues, whereas Id4 expression can only be detected in neuronal tissues and in the ventral portion of the epithelium of the developing stomach. The regions where I d 1 3 genes are expressed, such as gut, lung, kidney, tooth, whisker, and several glandular structures, are undergoing active morphogenetic activities. The expression patterns of Idl, 2, and 3 overlap in many organs, except in the tissues derived from primitive gut. In the latter, Id1 and Id3 signals are detected in the mesenchyme surrounding the epithelium, whereas Id2 is expressed within the epithelium. The difference in the patterns of expressions of I d 1 3 and Id4 suggest that the dominant negative transcriptional activity of these two subclasses of the Id family may have different physiological consequences. o 1996 Witey-Liss, Inc.

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The murine dominant negative helix‐loop‐helix (dnHLH) proteins inhibit the activities of bHLH transcription factors in diverse cell lineages (Benezra et al. [1990] Cell 61:49–59; Christy et al [1991] Proc. Natl. Acad. Sci. U.S.A. 88:1815–1819; Sun et al [1991] Mol. Cell Biol. 11: 5603–5611; Riechmann et al. [1994] Nucleic Acids Res. 22:749–755). Currently, there are four members in the dnHLH family, Id1, Id2, Id3, and Id4. In this report, we have performed a detailed comparative in situ hybridization analysis to examine their expression pattern during post‐gastrulational mouse development. Id1, 2, and 3 are expressed in multiple tissues, whereas Id4 expression can only be detected in neuronal tissues and in the ventral portion of the epithelium of the developing stomach. The regions where Id1–3 genes are expressed, such as gut, lung, kidney, tooth, whisker, and several glandular structures, are undergoing active morphogenetic activities. The expression patterns of Id1, 2, and 3 overlap in many organs, except in the tissues derived from primitive gut. In the latter, Id1 and Id3 signals are detected in the mesenchyme surrounding the epithelium, whereas Id2 is expressed within the epithelium. The difference in the patterns of expressions of Id1–3 and Id4 suggest that the dominant negative transcriptional activity of these two subclasses of the Id family may have different physiological consequences. © 1996 Wiley‐Liss, Inc.