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Abstract
Lack of the conserved NK2-domain of the cardiac transcription factor Nkx2.5 causes multiple heart
defects
Martina Schinke, Beth Israel Deaconess Med Ctr, Boston, MA; Silvio Litovsky, Texas Heart Inst,
Houston, TX; Anny Usheva, Beth Israel Deaconess Med Ctr, Boston, MA; Makoto Tanaka, Kyoto Univ, Kyoto
Japan; Colin Maguire, Charles Berul, Children's Hosp, Boston, MA; Seigo Izumo, Beth Israel Deaconess
Med Ctr, Harvard Med Sch, Boston, MA
The NK2 family of homeobox genes constitutes a family of transcription factors that play an
important role in different developmental processes. Members of this group are characterized by two
highly conserved protein domains: the homeodomain, conferring DNA binding activity, and the
NK2-specific domain (NK2-SD) of yet unknown function. One of the best characterized members of this
group is the early cardiogenic marker Nkx2.5. Loss of function of Nkx2.5 leads to embryonic lethality
around E10.5 due to an arrest of heart development at the looping stage. We have further dissected the
function of Nkx2.5 in vivo by creating a knockout mouse line harboring an in frame deletion of the
NK2-SD by Cre/loxP mediated excision. Homozygous mutant mice die at E14.5 due to severe cardiac
malformations, e.g. common AV canal, DORV, and VSD. Lack of the NK2-SD leads to downregulation of the
ventricular markers MLC-2v and Irx4 specifically in the right ventricle, and is accompanied with
reduced right ventricular function. This function of Nkx2.5 seems to be independent of its ability to
bind target DNA, since lack of the NK2-SD does not alter the DNA binding activity of Csx/Nkx2.5 in
vitro. Heterozygous mutant mice show a spectrum of cardiac defects related to cardiac septation and
valve morphogenesis, but lack conduction system defects as reported for heterozygous Nkx2.5 mice. The
phenotype observed in NK2-SD mutant mice shows that Nkx2.5 is not only crucial during early steps of
cardiogenesis but also plays an important role at later developmental stages.
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Experimental Design
Embryos were isolated at embryonic day 12.5. The entire embryo heart was taken and isolated in
ice-cold PBS and immediately frozen on dry-ice. Total RNA was extracted from pooled samples of wildtype
(3 samples), heterozygous (3 samples) and mutant embryos (3 samples) as described in Methodology.
Page last modified: 21-Jan-2003
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