G protein α 12/13 is involved in the heart tube formation via S1P signaling
○Ryu-ichi Fukuda1, Tomoya Kotani2, Atsuo Kawahara3, and Koichi Kawakami1,4
1)Division of Molecular and Developmental Biology, National Institute of Genetics.
In the early heart development, the myocardial precursors migrate between the pharyngeal
endoderm and the extra-embryonic yolk syncytial layer (YSL) toward the midline and then fuse
to form the functional heart tube. Sphingosine-1-phosphate (S1P) receptor 2 (S1PR2), which is
encoded by the miles-apart (mil) gene, is a component of the S1P signaling pathway and the
homozygous mil mutant shows abnormal heart tube formation, a defect called cardia bifida.
Previously, it was reported that S1PR2 is coupled with G protein α 12 and 13 (Gα12/13) and
involved in migration of vascular smooth muscle cells in vitro. However, the role of Gα12/13 in
the S1P signaling pathway during cardiac tube formation is largely unclear.
To determine if Gα12/13 is required for heart tube formation in vivo, we injected MOs
against Gα12, Gα13a and Gα13b and analyzed the injected embryos. The embryos injected
with all of the three MOs displayed cardia bifida. Consistent with this, injection of mRNA
encoding dominant negative forms of Gα12 (Gα12CT) or Gα13 (Gα13CT) mRNA showed
cardia bifida. These data indicate that Gα12/Gα13a/Gα13b are essential for heart tube
We have been performing a gene trap screen in zebrafish by using the Tol2-based gene trap
vector containing a splice acceptor and the GFP gene. We found a gene trap line that expressed
GFP in the heart and carried a transposon insertion in the first exon of the gna12 gene, which
encodes Gα12. We hypothesized that the insertion might reduce the activity of Gα12 and hence
reduce the activity of the S1P signaling. To test this, we injected mil MO into the embryo
homozygous for the insertion, expecting that the mil knockdown phenotype might be enhanced.
However, unexpectedly, the homozygous embryo injected with mil MO did not show cardia
bifida but showed normal heart tube formation.
From these data, we think that Gα12/Gα13a/Gα13b play an essential role for the heart tube
formation downstream of the S1P signaling in vivo, and hypothesize that the transposon
insertion may constitutively activate Gα12. Studies to test this hypothesis are currently ongoing.
2)Laboratoryof Molecular and Cellular Interactions, Graduate School of Science, Hokkaido University.
3)Department of Structural Analysis, National Cardiovascular Center Research Institute.
4)Department of Genetics, Graduate University of Advanced Studies (SOKENDAI).