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Characterization of Gene Targets Regulated by Lmx1b during Limb Dorsalization
Author(s) -
Feenstra Jennifer M,
Pira Charmaine U,
Kanaya Kohei,
Hoffman Sarah E,
Eppey Rick J,
Oberg Kerby C
Publication year - 2011
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.25.1_supplement.lb22
Subject(s) - limb development , biology , in situ hybridization , homeobox , mesoderm , limb bud , emx2 , anatomy , microarray , microbiology and biotechnology , gene , transcription factor , gene expression , embryonic stem cell , embryo , genetics
Lmx1b is a homeodomain transcription factor that regulates dorsal identity during limb development and Lmx1b knockout (KO) mice develop nearly symmetrical ventral‐ventral limbs with hypoplastic scapulae. Currently, downstream targets of Lmx1b within dorsal mesoderm that impart the limbs' unique dorsal asymmetry are unknown. To identify genes targeted by Lmx1b, we compared gene arrays from Lmx1b KO and wild type (WT) mouse limbs during limb outgrowth and patterning, i.e ., 11.5, 12.5, and 13.5 days post coitum (dpc). Real‐time PCR confirmed microarray results and whole mount in situ hybridization was used to localize and verify differential dorsal‐ventral expression. Microarray analysis identified 23 targets differentially expressed in all three arrays. Real‐time PCR confirmed 17 up‐regulated and 2 down‐regulated targets. Whole mount in situ hybridization of WT 12.5 dpc embryos demonstrated a dorsal‐ventral asymmetric pattern for the validated targets, further categorized as skeletal, soft tissue or other. Skeletal targets, including Emx2, Matrilin1 and Matrilin4, demonstrated a loss of scapular expression in the Lmx1b KO mice. Soft tissue targets which demonstrated dorsal mesodermal staining in WT were drastically reduced in KO limbs. This study provides the most comprehensive characterization of skeletal and soft tissue genes regulated by Lmx1b during limb development to date. Our data also suggest targets which Lmx1b may augment to ensure proper scapular development. Further studies are warranted to determine the mechanism of Lmx1b target regulation during limb dorsalization. NIH HD39421