The c‐Myc protein is involved in cell proliferation, differentiation and apoptosis though heterodimerization with Max to form a transcriptionally active sequence‐specific DNA binding complex. By means of sequential immunoprecipitation of chromatin using anti‐Max and anti‐Myc antibodies, we have identified a Myc‐regulated gene and genomic sites occupied by Myc‐Max in vivo. Four of 27 sites recovered by this procedure corresponded to the highest affinity ‘canonical’ CACGTG sequence. However, the most common in vivo binding sites belonged to the group of ‘non‐canonical’ E box‐related binding sites previously identified by in vitro selection. Several of the genomic fragments isolated contained transcribed sequences, including one, MrDb, encoding an evolutionarily conserved RNA helicase of the DEAD box family. The corresponding mRNA was induced following activation of a Myc‐estrogen receptor fusion protein (Myc‐ER) in the presence of a protein synthesis inhibitor, consistent with this helicase gene being a direct target of Myc‐Max. In addition, as for c‐Myc, the expression of MrDb is induced upon proliferative stimulation of primary human fibroblasts as well as B cells and down‐regulated during terminal differentiation of HL60 leukemia cells. Our results indicate that Myc‐Max heterodimers interact in vivo with a specific set of E box‐related DNA sequences and that Myc is likely to activate multiple target genes including a highly conserved DEAD box protein. Therefore, Myc may exert its effects on cell behavior through proteins that affect RNA structure and metabolism.

The B cell‐associated surface molecule CD40 plays a key role in T cell‐dependent B cell maturation, as individuals with defects in either CD40 or its ligand are impaired in immunoglobulin isotype class switching and germinal center formation. CD40 signaling activates downstream effectors, including the tyrosine protein kinase, Lyn, the phosphatidylinositol‐3‐kinase (PI‐3 kinase), and the transcription factor, NF‐kappa B. In this study, we demonstrate that stress‐activated protein kinases (SAPK) are activated after CD40 cross‐linking on various B cell lines or human tonsillar B cells. The activation is rapid and transient and is mediated through a cyclosporin A‐insensitive pathway. Furthermore, this signaling pathway appears not to rely on protein kinase C. While CD40 ligation strongly activates the SAPKs (up to 25‐fold), it does not affect members of the mitogen‐activated protein kinase family (MAPK; ERK1 and ERK2). Consistent with these data, CD40 signals up‐regulate c‐jun but not c‐fos mRNA and alter the transcription factor ATF2 but not the Raf‐1 protein. In summary, CD40 signaling preferentially induces SAPK but not MAPK.

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