Basic helix-loop-helix factors recruit nuclear factor I to enhance expression of the NaV 1.4 Na+ channel gene.
We have previously shown that the basic helix-loop-helix (bHLH) transcription factors coordinate Na(V) 1.4 Na(+) channel gene expression in skeletal muscle, but the identity of the co-factors they direct is unknown. Using C2C12 muscle cells as a model system, we test the hypothesis that the bHLH factors counteract negative regulation exerted through a repressor E box (-90/-85) by recruiting positive-acting transcription factors to the nucleotides (-135/-57) surrounding the repressor E box. We used electrophoretic mobility shift assays to identify candidate factors that bound the repressor E box or these adjacent regions. Repressor E box-binding factors included the known transcription factor, ZEB/AREB6, and a novel repressor E box-binding factor designated REB. Mutations of the repressor E box that interfere with the binding of these factors prevented repression. The transcription factor, nuclear factor I (NFI), bound immediately upstream and downstream of the repressor E box. Mutation of the NFI-binding sites diminished the ability of myogenin and MRF4 to counteract repression. Based on these observations we suggest that bHLH factors recruit NFI to enhance skeletal muscle Na(+) channel expression.
Hebert
Sadie L
SL
Department of Molecular and Biomedical Pharmacology, University of Kentucky Medical Center, Lexington, KY 40536, USA. slhebe2@email.uky.edu
Simmons
Christine
C
Thompson
Amy L
AL
Zorc
Catherine S
CS
Blalock
Eric M
EM
Kraner
Susan D
SD
D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000502:physiology*; D024721:E-Box Elements; D005786:Gene Expression Regulation; D018398:Homeodomain Proteins / Q000502:physiology; D006801:Humans; D009124:Muscle Proteins / Q000235:genetics*; D018482:Muscle, Skeletal / Q000378:metabolism*; D062553:NAV1.4 Voltage-Gated Sodium Channel; D051839:NFI Transcription Factors / Q000502:physiology*; D010766:Phosphorylation; D015222:Sodium Channels / Q000235:genetics*; D014157:Transcription Factors / Q000502:physiology; D000071799:Zinc Finger E-box-Binding Homeobox 1
D051792:Basic Helix-Loop-Helix Proteins; D018398:Homeodomain Proteins; D009124:Muscle Proteins; D062553:NAV1.4 Voltage-Gated Sodium Channel; D051839:NFI Transcription Factors; D015222:Sodium Channels; D014157:Transcription Factors; D000071799:Zinc Finger E-box-Binding Homeobox 1; C106251:SCN4A protein, human; C486307:ZEB1 protein, human