MyoD uses overlapping but distinct elements to bind E-box and tetraplex structures of regulatory sequences of muscle-specific genes.
Muscle differentiation and expression of muscle-specific proteins are initiated by the binding of heterodimers of the transcription factor MyoD with E2A proteins to E-box motif d(CANNTG) in promoters or enhancers of muscle-specific genes. MyoD homodimers, however, form tighter complexes with tetraplex structures of guanine-rich regulatory sequences of some muscle genes. In this work, we identified elements in MyoD that bind E-box or tetraplex structures of promoter sequences of the muscle-specific genes alpha7 integrin and sarcomeric Mitochondrial Creatine Kinase (sMtCK). Deletions of large domains of the 315 amino acids long recombinant MyoD indicated that the binding site for both E-box and tetraplex DNA is its basic region KRKTTNADRRKAATMRERRR that encompasses the three underlined clusters of basic residues designated R(1), R(2) and R(3). Deletion of a single or pairs of R triads or R111C substitution completely abolished the E-box-binding capacity of MyoD. By contrast, the MyoD deletion mutants Delta102-114, DeltaR(3), DeltaR(1)R(3) or DeltaR(2)R(3) maintained comparable tetraplex DNA-binding capacity as reflected by the similar dissociation constants of their protein-DNA complexes. Only deletion of all three basic clusters abolished the binding of tetraplex DNA. Implications of the binding of E-box and tetraplex DNA by non-identical MyoD elements are considered.
Shklover
Jeny
J
Department of Biochemistry, Rappaport Faculty of Medicine, Technion - Israel Institute of Technology, POB 9649 Bat Galim, Haifa 31096, Israel.
Etzioni
Shulamit
S
Weisman-Shomer
Pnina
P
Yafe
Anat
A
Bengal
Eyal
E
Fry
Michael
M
D024361:Amino Acids, Basic / Q000737:chemistry; D000818:Animals; D015703:Antigens, CD / Q000235:genetics; D001665:Binding Sites; D052281:Creatine Kinase, Mitochondrial Form / Q000235:genetics; D024721:E-Box Elements; D054856:G-Quadruplexes; D039001:Integrin alpha Chains / Q000235:genetics; D051379:Mice; D009154:Mutation; D017570:MyoD Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic
D024361:Amino Acids, Basic; D015703:Antigens, CD; D039001:Integrin alpha Chains; D017570:MyoD Protein; C057363:MyoD1 myogenic differentiation protein; C085725:integrin alpha7; D052281:Creatine Kinase, Mitochondrial Form