Targeting of the transcription factor Max during apoptosis: phosphorylation-regulated cleavage by caspase-5 at an unusual glutamic acid residue in position P1.
Max is the central component of the Myc/Max/Mad network of transcription factors that regulate growth, differentiation and apoptosis. Whereas the Myc and Mad genes and proteins are highly regulated, Max expression is constitutive and no post-translational regulation is known. We have found that Max is targeted during Fas-induced apoptosis. Max is first dephosphorylated and subsequently cleaved by caspases. Two specific cleavage sites for caspases in Max were identified, one at IEVE(10) decreasing S and one at SAFD(135) decreasing G near the C-terminus, which are cleaved in vitro by caspase-5 and caspase-7 respectively. Mutational analysis indicates that both sites are also used in vivo. Thus Max represents the first caspase-5 substrate. The unusual cleavage after a glutamic acid residue is observed only with full-length, DNA-binding competent Max protein but not with corresponding peptides, suggesting that structural determinants might be important for this activity. Furthermore, cleavage by caspase-5 is inhibited by the protein kinase CK2-mediated phosphorylation of Max at Ser-11, a previously mapped phosphorylation site in vivo. These findings suggest that Fas-mediated dephosphorylation of Max is required for cleavage by caspase-5. The modifications that occur on Max in response to Fas signalling affect the DNA-binding activity of Max/Max homodimers. Taken together, our findings uncover three distinct processes, namely dephosphorylation and cleavage by caspase-5 and caspase-7, that target Max during Fas-mediated apoptosis, suggesting the regulation of the Myc/Max/Mad network through its central component.
Krippner-Heidenreich
A
A
Institut für Molekularbiologie, Medizinische Hochschule Hannover, Carl-Neuberg Strasse 1, 30623 Hannover, Germany.
Talanian
R V
RV
Sekul
R
R
Kraft
R
R
Thole
H
H
Ottleben
H
H
Lüscher
B
B
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D050976:Basic-Leucine Zipper Transcription Factors; D019556:COS Cells; D053179:Caspase 7; D020169:Caspases / Q000378:metabolism*; D002522:Chlorocebus aethiops; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D019281:Dimerization; D018698:Glutamic Acid; D006801:Humans; D007075:Immunoglobulin M / Q000494:pharmacology; D019169:Jurkat Cells; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000737:chemistry; D010449:Peptide Mapping; D010766:Phosphorylation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism; D014162:Transfection; D019014:fas Receptor / Q000502:physiology
D000590:Amino Acid Chloromethyl Ketones; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D050976:Basic-Leucine Zipper Transcription Factors; D015853:Cysteine Proteinase Inhibitors; D004268:DNA-Binding Proteins; D007075:Immunoglobulin M; C499330:MAX protein, human; C067850:Myc associated factor X; D010446:Peptide Fragments; D011994:Recombinant Proteins; D014157:Transcription Factors; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D019014:fas Receptor; D018698:Glutamic Acid; C099217:CASP5 protein, human; C505520:CASP7 protein, human; D053179:Caspase 7; D020169:Caspases