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1.
Development ; 128(18): 3595-607, 2001 Sep.
Article in English | MEDLINE | ID: mdl-11566863

ABSTRACT

During development of the vertebrate hindbrain, Hox genes play multiple roles in the segmental processes that regulate anteroposterior (AP) patterning. Paralogous Hox genes, such as Hoxa3, Hoxb3 and Hoxd3, generally have very similar patterns of expression, and gene targeting experiments have shown that members of paralogy group 3 can functionally compensate for each other. Hence, distinct functions for individual members of this family may primarily depend upon differences in their expression domains. The earliest domains of expression of the Hoxa3 and Hoxb3 genes in hindbrain rhombomeric (r) segments are transiently regulated by kreisler, a conserved Maf b-Zip protein, but the mechanisms that maintain expression in later stages are unknown. In this study, we have compared the segmental expression and regulation of Hoxa3 and Hoxb3 in mouse and chick embryos to investigate how they are controlled after initial activation. We found that the patterns of Hoxa3 and Hoxb3 expression in r5 and r6 in later stages during mouse and chick hindbrain development were differentially regulated. Hoxa3 expression was maintained in r5 and r6, while Hoxb3 was downregulated. Regulatory comparisons of cis-elements from the chick and mouse Hoxa3 locus in both transgenic mouse and chick embryos have identified a conserved enhancer that mediates the late phase of Hoxa3 expression through a conserved auto/cross-regulatory loop. This block of similarity is also present in the human and horn shark loci, and contains two bipartite Hox/Pbx-binding sites that are necessary for its in vivo activity in the hindbrain. These HOX/PBC sites are positioned near a conserved kreisler-binding site (KrA) that is involved in activating early expression in r5 and r6, but their activity is independent of kreisler. This work demonstrates that separate elements are involved in initiating and maintaining Hoxa3 expression during hindbrain segmentation, and that it is regulated in a manner different from Hoxb3 in later stages. Together, these findings add further strength to the emerging importance of positive auto- and cross-regulatory interactions between Hox genes as a general mechanism for maintaining their correct spatial patterns in the vertebrate nervous system.


Subject(s)
Avian Proteins , Body Patterning/genetics , Homeodomain Proteins/genetics , Oncogene Proteins , Rhombencephalon/embryology , Xenopus Proteins , Animals , Base Sequence , Binding Sites , Biological Evolution , Chick Embryo , Conserved Sequence , DNA-Binding Proteins , Enhancer Elements, Genetic , G-Box Binding Factors , Gene Expression Regulation, Developmental , MafB Transcription Factor , Mice , Mice, Transgenic , Models, Genetic , Regulatory Sequences, Nucleic Acid , Sequence Homology, Nucleic Acid , Species Specificity , Transcription Factors
2.
Dev Biol ; 224(2): 238-49, 2000 Aug 15.
Article in English | MEDLINE | ID: mdl-10926763

ABSTRACT

The Polycomb group genes are required for the correct expression of the homeotic complex genes and segment specification during Drosophila embryogenesis and larval development. In mouse, inactivation studies of several Polycomb group genes indicate that they are also involved in Hox gene regulation. We have used our previously generated M33 mutants to study the function of M33, the mouse homologue of the Polycomb gene of Drosophila. In this paper, we show that in the absence of M33, the window of Hoxd4 retinoic acid (RA) responsiveness is opened earlier and that Hoxd11 gene expression is activated earlier in development This indicates that M33 antagonizes the RA pathway and has a function in the establishment of the early temporal sequence of activation of Hox genes. Despite the early activation, A-P boundaries are correct in later stages, indicating a separate control mechanism for early aspects of Hox regulation. This raises a number of interesting issues with respect to the roles of both Pc-G proteins and Hox regulatory mechanisms. We propose that a function of the M33 protein is to control the accessibility of retinoic acid response elements in the vicinity of Hox genes regulatory regions by direct or indirect mechanisms or both. This could provide a means for preventing ectopic transactivation early in development and be part of the molecular basis for temporal colinearity of Hox gene expression.


Subject(s)
Gene Expression Regulation, Developmental/drug effects , Genes, Homeobox , Repressor Proteins/physiology , Tretinoin/pharmacology , Animals , Bone and Bones/abnormalities , Bone and Bones/embryology , Mice , Mice, Mutant Strains , Polycomb Repressive Complex 1 , Polycomb-Group Proteins , Repressor Proteins/genetics
3.
Nat Cell Biol ; 1(8): E203-7, 1999 Dec.
Article in English | MEDLINE | ID: mdl-10587659

ABSTRACT

Efficient gene transfer by electroporation of chick embryos in ovo has allowed the development of new approaches to the analysis of gene regulation, function and expression, creating an exciting opportunity to build upon the classical manipulative advantages of the chick embryonic system. This method is applicable to other vertebrate embryos and is an important tool with which to address cell and developmental biology questions. Here we describe the technical aspects of in ovo electroporation, its different applications and future perspectives.


Subject(s)
Chick Embryo/metabolism , Chickens/genetics , Chickens/metabolism , Electroporation/methods , Gene Expression Regulation, Developmental , Oocytes/metabolism , Animals , Embryo, Mammalian/metabolism , Embryo, Nonmammalian , Gene Transfer Techniques , Genes, Reporter/genetics , Mice , Organ Specificity
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