Congenital heart diseases (CHDs) represent the most common form of birth defect and present as structural malformations of the heart and/or great vessels. During normal cardiogenesis, the primitive heart tube undergoes looping and chamber formation to generate the mature heart structure. Proper looping depends on elongation of the heart tube through addition of new myocardium and endocardium to the outflow pole from a recently discovered cardiac progenitor cell (CPC) population termed the second heart field (SHF). In both chick and mouse, defects in SHF cause ventricular, outflow tract and great vessel malformations suggesting that SHF defects are major causes of embryonic lethality and CHD in humans. However, the genetic alterations underlying most CHDs remain unknown. Unpublished data from our lab demonstrates that the SHF is evolutionarily conserved in zebrafish and marked by the TGFb signaling component Latent Transforming Growth Factor Beta Binding Protein 3 (LTBP3). LTBP3 expression analyses, cre/loxP-based lineage tracing, and genetic loss-of-function studies have localized the SHF domain, determined the cell types and structures derived from the SHF, and characterized the phenotypes arising from SHF deficiencies. Most importantly, these findings demonstrate that ventricular and outflow tract developments are highly conserved with that of higher vertebrates prior to septation. Thus, the zebrafish is an ideal model organism to discover and analyze conserved genetic programs potentially underlying human CHDs with their genetic tractability, unmatched real-time imaging, and ease of embryological manipulations. As a "reverse" approach to forward genetic screening for identifying essential regulators of cardiogenesis, we generated a list of several-hundred transcripts restricted to the zebrafish embryonic heart by comparing the transcriptomes of purified hearts and whole embryos. Compelling preliminary data show that a transcriptional co-factor, Four and a Half LIM domains (FHL), represents the earliest known marker of the zebrafish SHF and is required for SHF-mediated cardiogenesis.
In Aim 1, we propose to characterize the full array of cardiac phenotypes present in embryos lacking FHL function and elucidate the cellular defects present in these SHF CPCs.
In Aim 2, we will investigate a potential genetic and/or physical interaction between FHL and Nkx2.5, a conserved transcription factor required for SHF development in higher vertebrates. Finally, in Aim 3 we will utilize a unique cre/loxP-based transgenic SHF reporter strain to perform the first unbiased small molecule screen to reveal genetic pathways essential for SHF development. Together, these studies will reveal new regulatory mechanisms of SHF development that may lead to improved diagnostics/therapeutics and potentially identify novel genes underlying CHD.

Public Health Relevance

Congenital heart disease (CHD) is the most common birth defect among infants and it has been demonstrated that these defects arise from alterations in the secondary heart field (SHF). The genetic causes of CHD are largely unknown, this project aims to dissect new regulatory mechanisms of SHF-mediated cardiogenesis that may lead to improved diagnostics/therapeutics and potentially identify novel genes underlying conotruncal birth defects.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32HL110627-02
Application #
8335578
Study Section
Special Emphasis Panel (ZRG1-F10A-S (20))
Program Officer
Meadows, Tawanna
Project Start
2011-07-22
Project End
2014-07-21
Budget Start
2012-07-22
Budget End
2013-07-21
Support Year
2
Fiscal Year
2012
Total Cost
$53,942
Indirect Cost
Name
Massachusetts General Hospital
Department
Type
DUNS #
073130411
City
Boston
State
MA
Country
United States
Zip Code
02199
Nevis, Kathleen; Obregon, Pablo; Walsh, Conor et al. (2013) Tbx1 is required for second heart field proliferation in zebrafish. Dev Dyn 242:550-9
Paffett-Lugassy, Noelle; Singh, Reena; Nevis, Kathleen R et al. (2013) Heart field origin of great vessel precursors relies on nkx2.5-mediated vasculogenesis. Nat Cell Biol 15:1362-9