The long-term goal of this application is to understand the molecular mechanisms underlying the regulation of early mammalian embryogenesis, specifically gastrulation. Mammalian development is controlled by genes which are differentially expressed in a temporal and spatial manner. The goal of this proposal is to identify and isolate genes which control developmental processes during gastrulation, using the mouse as a model system. To achieve this goal, the applicant proposes to employ a genetic screen using """"""""gene trap"""""""" retroviral vectors in pluripotent mouse embryonic stem cells (ES cells). These vectors contain a reporter gene, Escherichia lacZ, whose expression is dependent on transcription initiated from cis- acting regulatory sequences of a cellular """"""""tagged"""""""" gene. Therefore, genes potentially important for developmental processes can be identified based upon the patterns of the reporter gene expression disrupting the normal expression of the cellular gene, resulting in abnormal developmental process. Finally, the integrated provirus serves as a """"""""tag"""""""" to isolate the disrupted cellular gene. To use this strategy effectively, for genetic screening in developing mouse embryos, the applicant proposes to pre-screen a large number of ES cell clones for desired retroviral insertions in gastrulation- specific genes by monitoring the pattern of lacZ expression (i) upon differentiation of ES cells in vitro into embryoid bodies, and (ii) in vivo in developing chimeric embryos. Transgenic mice will be generated and analyzed for developmental abnormalities associated with the mutated gene(s). Finally, the investigator proposes to isolate the disrupted endogenous gene(s). Successful identification of gastrulation-specific genes will be crucial for elucidating the regulation of early mammalian embryogenesis and, more generally, the molecular basis of developmental abnormalities in humans.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
5R29HD031534-03
Application #
2025533
Study Section
Human Embryology and Development Subcommittee 1 (HED)
Project Start
1994-12-01
Project End
1999-11-30
Budget Start
1996-12-01
Budget End
1997-11-30
Support Year
3
Fiscal Year
1997
Total Cost
Indirect Cost
Name
Mount Sinai School of Medicine
Department
Biochemistry
Type
Schools of Medicine
DUNS #
114400633
City
New York
State
NY
Country
United States
Zip Code
10029
Campagnolo, Luisa; Leahy, Amy; Chitnis, Smita et al. (2005) EGFL7 is a chemoattractant for endothelial cells and is up-regulated in angiogenesis and arterial injury. Am J Pathol 167:275-84
Kuhnert, Frank; Campagnolo, Luisa; Xiong, Jing-Wei et al. (2005) Dosage-dependent requirement for mouse Vezf1 in vascular system development. Dev Biol 283:140-56
Kuhnert, Frank; Stuhlmann, Heidi (2004) Identifying early vascular genes through gene trapping in mouse embryonic stem cells. Curr Top Dev Biol 62:261-81
Stuhlmann, Heidi (2003) Gene trap vector screen for developmental genes in differentiating ES cells. Methods Enzymol 365:386-406
Xiong, J W; Leahy, A; Stuhlmann, H (1999) Retroviral promoter-trap insertion into a novel mammalian septin gene expressed during mouse neuronal development. Mech Dev 86:183-91
Xiong, J W; Leahy, A; Lee, H H et al. (1999) Vezf1: A Zn finger transcription factor restricted to endothelial cells and their precursors. Dev Biol 206:123-41
Xiong, J W; Battaglino, R; Leahy, A et al. (1998) Large-scale screening for developmental genes in embryonic stem cells and embryoid bodies using retroviral entrapment vectors. Dev Dyn 212:181-97