Differentiation of cardiac Purkinje fibers requires precise spatiotemporal regulation of Nkx2-5 expression.
Nkx2-5 gene mutations cause cardiac abnormalities, including deficits of function in the atrioventricular conduction system (AVCS). In the chick, Nkx2-5 is elevated in Purkinje fiber AVCS cells relative to working cardiomyocytes. Here, we show that Nkx2-5 expression rises to a peak as Purkinje fibers progressively differentiate. To disrupt this pattern, we overexpressed Nkx2-5 from embryonic day 10, as Purkinje fibers are recruited within developing chick hearts. Overexpression of Nkx2-5 caused inhibition of slow tonic myosin heavy chain protein (sMHC), a late Purkinje fiber marker but did not affect Cx40 levels. Working cardiomyocytes overexpressing Nkx2-5 in these hearts ectopically up-regulated Cx40 but not sMHC. Isolated embryonic cardiomyocytes overexpressing Nkx2-5 also displayed increased Cx40 and suppressed sMHC. By contrast, overexpression of a human NKX2-5 mutant did not effect these markers in vivo or in vitro, suggesting one possible mechanism for clinical phenotypes. We conclude that a prerequisite for normal Purkinje fiber maturation is precise regulation of Nkx2-5 levels.
Harris
Brett S
BS
Department of Cell Biology, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
Spruill
Laura
L
Edmonson
Angela M
AM
Rackley
Mary S
MS
Benson
D Woodrow
DW
O'Brien
Terrence X
TX
Gourdie
Robert G
RG
D000256:Adenoviridae; D000818:Animals; D030161:Avian Proteins / Q000096:biosynthesis* / Q000235:genetics; D015415:Biomarkers; D002454:Cell Differentiation / Q000502:physiology*; D002467:Cell Nucleus / Q000378:metabolism; D002642:Chick Embryo; D017630:Connexins / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D005822:Genetic Vectors; D018398:Homeodomain Proteins / Q000096:biosynthesis* / Q000235:genetics; D032383:Myocytes, Cardiac / Q000378:metabolism; D018995:Myosin Heavy Chains / Q000096:biosynthesis / Q000235:genetics; D011690:Purkinje Fibers / Q000166:cytology* / Q000378:metabolism; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics; D000097003:Gap Junction alpha-5 Protein
D030161:Avian Proteins; D015415:Biomarkers; D017630:Connexins; D018398:Homeodomain Proteins; D018995:Myosin Heavy Chains; D014157:Transcription Factors; D000097003:Gap Junction alpha-5 Protein