PARENT SESSION
MINISYMPOSIUM VII. Conditional Gene Disruption in the Mouse: Concepts and Applications.
11:00 AM-12:30 PM
Grand Ballroom V
Chair: Adashi, Eli¹, ¹

(M19) USING CONDITIONAL GENE INACTIVATION TO STUDY SIGNALING PATHWAYS DURING LIMB DEVELOPMENT.

Lewandoski, Mark¹, Wilson, Catherine¹, Anderson, Rhonda¹, Mishina, Yuji², Williams, Trevor⁴, ¹ Cancer and Developmental Biology Lab, Frederick, MD² Laboratory of Reproductive and Developmental Toxicology, Research Triangle Park, NC⁴ Dept. of Craniofacial Biology, Denver, CO

ABSTRACT- I will focus on strategies and approaches that maximize the efficacy of tissue-specific knockouts (via Cre/Flp DNA recombinases) while focusing specifically on limb development in the mouse. Limb development provides an excellent system for studying the mechanisms that regulate pattern formation during embryogenesis. From one perspective, the three axis of the limb are regulated by secreted molecules specifically expressed from organizing centers such as the zone of polarizing activity, the apical ectodermal ridge (AER) and the non-ridge ectoderm. Whilst many of the factors crucial to the activity of these organizing centers have been identified, it has been often impossible to study their roles via mouse knockouts because the null homozygotes die too early to form limbs. Therefore tissue-specific gene inactivation studies are required. Such strategies have revealed that signaling through the Fibroblast Growth Factor (FGF) and Bone Morphogenetic Protein (BMP) pathways is essential to normal limb development. AER-specific knockouts of Fgf4 and Fgf8 demonstrate that these ligands play partially redundant roles in determining the proximal-distal axis. Inactivation of the receptor BmpR1A in the AER demonstrates that BMP signaling is also required for normal proximal-distal development and at an earlier stage than any known FGF signal. Inactivation of BmpR1A in the ventral ectoderm (via En1-Cre) results in normal dorsoventral limb patterning. As BmpR1A has been previously shown to control dorsoventral patterning via activation of the En1 gene, these data suggest that BMP signaling is required only at very early stages of limb development for a normal dorsoventral axis. Finally, inactivation of BmpR1A in the limb bud mesenchyme results in digit loss, an equally-sized tibia and fibula as well as trifurcation so that a third skeletal element forms in addition to the tibia and fibula, thus indicating that BMP signaling is required for normal limb patterning.

KEY WORDS: Limb development, Cre/loxP, FGF, BMP