Mad1 function is regulated through elements within the carboxy terminus.
Myc and Mad are basic helix-loop-helix leucine zipper (bHLH-LZ) proteins that heterodimerize with Max to bind DNA and thereby influence the transcription of Myc-responsive genes. Myc-Max dimers transactivate whereas Mad-Max-mSin3 complexes repress Myc-mediated transcriptional activation. We have previously shown that the N-terminal mSin3 binding domain and the centrally located bHLH-LZ are required for Mad1 to function during a molecular switch from proliferation to differentiation. Here we demonstrate that the carboxy terminus (CT) of Mad1 contains previously unidentified motifs necessary for the regulation of Mad1 function. We show that removal of the last 18 amino acids of Mad1 (region V) abolishes the growth-inhibitory function of the protein and the ability to reverse a Myc-imposed differentiation block. Moreover, deletion of region V results in a protein that binds DNA weakly and no longer represses Myc-dependent transcriptional activation. In contrast, deletion of the preceding 24 amino acids (region IV) together with region V restores DNA binding and transcriptional repression, suggesting a functional interplay between these two regions. Furthermore, phosphorylation within region IV appears to mediate this interplay. These findings indicate that novel regulatory elements are present in the Mad1 CT.
Barrera-Hernandez
G
G
NCI-Navy Medicine Branch, Genetics Department, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20889-5105, USA.
Cultraro
C M
CM
Pianetti
S
S
Segal
S
S
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D004268:DNA-Binding Proteins / Q000235:genetics*; D005786:Gene Expression Regulation; D018257:Helix-Loop-Helix Motifs; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D014157:Transcription Factors