Gene-regulatory properties of Myc helix-loop-helix/leucine zipper mutants: Max-dependent DNA binding and transcriptional activation in yeast correlates with transforming capacity

Abstract

Max is a basic helix-loop-helix/leucine zipper (bHLH/LZ) protein that forms sequence-specific DNA-binding complexes with the c-Myc oncoprotein (Myc). Using Saccharomyces cerevisiae, we have shown that the Max bHLH/LZ domain enables Myc to activate transcription through CACGTG and CACATG sequences in vivo, and that the number and context of such sites determines the level of activation. In addition, we have used yeast to investigate the role of the Myc helix-loop-helix (HLH) and leucine zipper (LZ) motifs in mediating Max-dependent DNA-binding and transcriptional activation in vivo using HLH/LZ mutants generated by site-directed mutagenesis. The results show that, while both motifs are essential for Myc to activate transcription, helix 2 of the HLH together with the contiguous LZ suffice to mediate complex formation with Max, whilst helix 1 is essential for sequence-specific DNA binding of Myc-Max complexes. Furthermore, the ability of Myc HLH/LZ mutants to bind DNA and activate transcription in collaboration with Max correlates closely with their neoplastic transforming activity in higher eukaryotic cells.