Identification of novel genes affecting mesoderm formation and morphogenesis through an enhanced large scale functional screen in Xenopus.
The formation of mesoderm is an important developmental process of vertebrate embryos, which can be broken down into several steps; mesoderm induction, patterning, morphogenesis and differentiation. Although mesoderm formation in Xenopus has been intensively studied, much remains to be learned about the molecular events responsible for each of these steps. Furthermore, the interplay between mesoderm induction, patterning and morphogenesis remains obscure. Here, we describe an enhanced functional screen in Xenopus designed for large-scale identification of genes controlling mesoderm formation. In order to improve the efficiency of the screen, we used a Xenopus tropicalis unique set of cDNAs, highly enriched in full-length clones. The screening strategy incorporates two mesodermal markers, Xbra and Xmyf-5, to assay for cell fate specification and patterning, respectively. In addition we looked for phenotypes that would suggest effects in morphogenesis, such as gastrulation defects and shortened anterior-posterior axis. Out of 1728 full-length clones we isolated 82 for their ability to alter the phenotype of tadpoles and/or the expression of Xbra and Xmyf-5. Many of the clones gave rise to similar misexpression phenotypes (synphenotypes) and many of the genes within each synphenotype group appeared to be involved in similar pathways. We determined the expression pattern of the 82 genes and found that most of the genes were regionalized and expressed in mesoderm. We expect that many of the genes identified in this screen will be important in mesoderm formation.
Chen
Jun-An
JA
Wellcome Trust/Cancer Research UK Gurdon Institute, Tennis Court Road, Cambridge CB2 1QN, UK.
Voigt
Jana
J
Gilchrist
Mike
M
Papalopulu
Nancy
N
Amaya
Enrique
E
D000818:Animals; D017209:Apoptosis; D019521:Body Patterning; D019070:Cell Lineage; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D047108:Embryonic Development; D020224:Expressed Sequence Tags; D005697:Galactosides / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D005821:Genetic Techniques; D005820:Genetic Testing; D017403:In Situ Hybridization; D007211:Indoles / Q000378:metabolism; D008648:Mesoderm / Q000378:metabolism*; D008954:Models, Biological; D005810:Multigene Family; D009124:Muscle Proteins / Q000378:metabolism; D051787:Myogenic Regulatory Factor 5; D010641:Phenotype; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020825:T-Box Domain Proteins / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism; D014158:Transcription, Genetic; D014981:Xenopus / Q000235:genetics*; D029867:Xenopus Proteins
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D005697:Galactosides; D007211:Indoles; D009124:Muscle Proteins; C497419:Myf5 protein, Xenopus; D051787:Myogenic Regulatory Factor 5; D012333:RNA, Messenger; D020825:T-Box Domain Proteins; C116765:TBXT protein, Xenopus; D015534:Trans-Activators; D029867:Xenopus Proteins; D012313:RNA; C044888:5-bromo-4-chloro-3-indolyl beta-galactoside