Experimental manipulation in birds has shown that trunk dermis has a double origin: dorsally, it derives from the somite dermomyotome, while ventrally, it is formed by the somatopleure. Taking advantage of an nlacZ reporter gene integrated into the mouse Msx1 locus (Msx1(nlacZ) allele), we detected segmental expression of the Msx1 gene in cells of the dorsal mesenchyme of the trunk between embryonic days 11 and 14. Replacing somites from a chick host embryo by murine Msx1(nlacZ )somites allowed us to demonstrate that these Msx1-(beta)-galactosidase positive cells are of somitic origin. We propose that these cells are dermal progenitor cells that migrate from the somites and subsequently contribute to the dorsalmost dermis. By analysing Msx1(nlacZ) expression in a Splotch mutant, we observed that migration of these cells does not depend on Pax3, in contrast to other migratory populations such as limb muscle progenitor cells and neural crest cells. Msx1 expression was never detected in cells overlying the dermomyotome, although these cells are also of somitic origin. Therefore, we propose that two somite-derived populations of dermis progenitor cells can be distinguished. Cells expressing the Msx1 gene would migrate from the somite and contribute to the dermis of the dorsalmost trunk region. A second population of cells would disaggregate from the somite and contribute to the dermis overlying the dermomyotome. This population never expresses Msx1. Msx1 expression was investigated in the context of the onset of dermis formation monitored by the Dermo1 gene expression. The gene is downregulated prior to the onset of dermis differentiation, suggesting a role for Msx1 in the control of this process.

In myoblast cell cultures, the Msx1 protein is able to repress myogenesis and maintain cells in an undifferentiated and proliferative state. However, there has been no evidence that Msx1 is expressed in muscle or its precursors in vivo. Using mice with the nlacZ gene integrated into the Msx1 locus, we show that the reporter gene is expressed in the lateral dermomyotome of brachial and thoracic somites. Cells from this region will subsequently contribute to forelimb and intercostal muscles. Using Pax3 gene transcripts as a marker of limb muscle progenitor cells as they migrate from the somites, we have defined precisely the somitic origin and timing of cell migration from somites to limb buds in the mouse. Differences in the timing of migration between chick and mouse are discussed. Somites that label for Msx1(nlacZ )transgene expression in the forelimb region partially overlap with those that contribute Pax3-expressing cells to the forelimb. In order to see whether Msx1 is expressed in this migrating population, we have grafted somites from the forelimb level of Msx1(nlacZ )mouse embryos into a chick host embryo. We show that most cells migrating into the wing field express the Msx1(nlacZ )transgene, together with Pax3. In these experiments, Msx1 expression in the somite depends on the axial position of the graft. Wing mesenchyme is capable of inducing Msx1 transcription in somites that normally would not express the gene; chick hindlimb mesenchyme, while permissive for this expression, does not induce it. In the mouse limb bud, the Msx1(nlacZ )transgene is downregulated prior to the activation of the Myf5 gene, an early marker of myogenic differentiation. These observations are consistent with the proposal that Msx1 is involved in the repression of muscle differentiation in the lateral half of the somite and in limb muscle progenitor cells during their migration.

The mouse Msx1 gene is a homologue of the Drosophila msh gene. It is expressed in a spatially and temporally complex pattern which suggests that this gene is involved in several inductive processes between the ectoderm and the mesoderm during embryonic development, To study the regulation of the mouse Msx1gene, we developed a transgenesis analysis relying on ES cells as a vector to transfer transgenic sequences into the mouse genome. Five transgenes were constructed by fusion of a nlacZ reporter gene to 5 kb of DNA from the Msx1 5' upstream region and progressive deletions thereof. These transgenes were transfected into ES cells and their expression was studied during in vitro differentiation.. ES cells carrying the longest transgene were further microinjected into morulae. The resulting chimeric embryos were analysed between embryonic days (E) 9.5 and 12.5 for beta-galactosidase expression. Between E9.5 and E11.5, the expression pattern of the t ransgene mimics closely that of the endogenous gene, apart for expression in the fronto-nasal process mesenchyme. At E12.5, expression is essentially lost from mesenchyme and retained only in the ectoderm. Our results show that ES cells constitute an efficient vector to transfer transgenes into chimeric embryos and analyse their expression and that there is a correlation between expression of a given transgene in differentiated ES cells and in transgenic chimeras. They further demonstrate that a 5 kb promoter region fr om the Msx1 gene is sufficient to provide most of the endogenous gene expression.

We have generated a null allele of the mouse Msx1 homeobox gene by insertion of an nlacZ reporter gene into its homeobox. The sensitivity of β-galactosidase detection permitted us to reveal novel aspects of Msx1 gene expression in heterozygous embryos, in particular in ectoderm and mesoderm during gastrulation, and in migrating neural crest cells. Homozygous mutant mice die at birth with facial defects (see Satokata, I. and Maas, R. (1994) Msx1 deficient mice exhibit cleft palate and abnormalities of craniofacial and tooth development. Nat. Genet. 6, 348–356). To investigate the reason for this limited phenotype, we compared the pattern of Msx1 expression with that of the closely related Msx2 gene in wild type embryos and in Msx1-/- mutants. Notably, whereas the expression of Msx1 and Msx2 overlap in the developing limb, this is not the case in the facial regions most affected in the mutant.

The Msx1 homeobox locus has been mapped in relation to the mutations hammer-toe (Hm) and hemimelic extra toes (Hx). Msx1 is expressed in the developing limb, while limb development is affected by the Hm and Hx mutations. Hm and Hx are very tightly linked loci. In interspecific crosses, the segregation of either mutation was followed in relation to polymorphic alleles of Msx1, Il6, and En2, to give a fine map around the mutant loci. Our results show that Msx1 is not allelic to either of the mutations, but is located about 3 cM from them. Il6 did not recombine with either Hm or Hx and, therefore, provides a point of access for the analysis of these mutations at the molecular level.

In tetrapod vertebrates, limbs are formed as a result of inductive interactions between ectoderm and mesoderm. The mesoderm from the limb field induces the formation, in the ectoderm, of a pseudo-stratified epithelium, the apical ectodermal ridge, which in turn is required for limb mesoderm outgrowth and patterning. Homeobox genes from the msh family are expressed in the apical region of limb bud mesoderm. Using the potential of chick experimental embryology, we have demonstrated that these genes respond to ecto-mesodermal induction at this site and may be implicated in the response of the mesoderm to the ectodermal inductive activity. This property appears to be more general for the sites in the embryo which grow and are patterned as a result of interactions between ectoderm and subjacent mesoderm, since many of them are places for the expression of the msh-related genes (e.g. fronto-nasal and maxillary processes, tooth germ, genital tubercle). These genes might be implicated in patterning events at these sites through the activation of other genes directly involved in the definition of positional information, such as Hox genes.

In tetrapod vertebrates, limbs are formed as a result of inductive interactions between ectoderm and mesoderm. The mesoderm from the limb field induces the formation, in the ectoderm, of a pseudo-stratified epithelium, the apical ectodermal ridge, which in turn is required for limb mesoderm outgrowth and patterning. Homeobox genes from the msh family are expressed in the apical region of limb bud mesoderm. Using the potential of chick experimental embryology, we have demonstrated that these genes respond to ecto-mesodermal induction at this site and may be implicated in the response of the mesoderm to the ectodermal inductive activity. This property appears to be more general for the sites in the embryo which grow and are patterned as a result of interactions between ectoderm and subjacent mesoderm, since many of them are places for the expression of the msh-related genes (e.g. fronto-nasal and maxillary processes, tooth germ, genital tubercle). These genes might be implicated in patterning events at these sites through the activation of other genes directly involved in the definition of positional information, such as Hox genes.

The utrophin (UTRN) locus is the autosomal homologue of the DMD (Duchenne muscular dystrophy) gene and encodes a protein, utrophin which is thought to be upregulated in the absence of dystrophin. In this study the spatial and temporal expression of the UTRN gene has been examined during mouse embryogenesis and compared with that of the DMD gene. The patterns of expression of these two genes are very different. Whilst DMD is expressed largely in mesodermal derivatives such as cardiac and striated muscle, UTRN shows a more widespread distribution and is expressed in neural tube, tissues which originate from neural crest and a variety of other sites of non‐neural origin. In early embryos UTRN transcripts initially accumulate in the mid‐neural plate and thereafter in the caudal neural tube. UTRN mRNA then becomes abundant in a subset of neural crest cell derived tissues, in particular the spinal and facial ganglia and ossifying facial cartilages. UTRN is also expressed in a variety of other sites and organs such as the tendon primordia in the digits, the pituitary, thyroid and adrenal glands, cardiac muscle, kidney and lung, follicies of the vibrissae and the outflow tract of the heart. Several patterns of UTRN expression are apparent and we discuss the possibility that these can be ascribed to a family of mRNAs transcribed from the UTRN gene using alternative promoters. © 1993 Wiley‐Liss, Inc.

Using isogene specific probes and in situ hybridization on sections, we have examined the expression of structural and regulatory genes in the mouse embryo during the formation of cardiac and skeletal muscle. The temporal and spatial information thus obtained about the onset of expression of muscle genes provides insight into the regulation of myogenesis in vivo. Actin and myosin sequences present in different compartments of the adult heart are initially all co-expressed in the cardiac tube (between 7-8 days). The process of spatial restriction to atrial or ventricular compartments of the heart takes place asynchronously later. In contrast, the onset of expression of actin and myosin genes in the first skeletal muscle, the myotome, which corresponds to the central compartment of the somite, as well as their subsequent down-regulation in different skeletal muscle masses, takes place very asynchronously. One might predict that factor(s) responsible for the transcriptional activation of these genes are present in sufficient quantity in the cardiac tube, whereas in skeletal muscle individual genes are responding to variable levels of factor(s). In fact the four myogenic regulatory sequences present in the mouse - MyoD1, myogenin, myf-5 and myf-6 - do show distinct patterns of expression during the development of skeletal muscle. None of these sequences have been detected in the heart. In the myotome there is no general correlation between the appearance of a particular myogenic sequence and the activation of a particular structural gene. A striking example of this is provided by the muscle isoform of creatine phosphokinase. We would propose that each muscle structural gene has a different threshold of activation, depending on the quantity and nature of the myogenic factor present. We have also examined the onset of expression of the X-linked dystrophin gene known to be expressed in adult heart and skeletal muscle. In the myotome dystrophin transcripts are first detected at the time when myosin heavy chains first accumulate and muscular contraction is initiated. In contrast in the cardiac tube dystrophin transcripts are not detected initially, at a time (from 8 days) when the heart contracts. This observation can be correlated with the pathology of the disease which points to a more essential role of dystrophin in skeletal muscle. No muscle structural gene examined is expressed in the somite prior to myotome formation. If the myogenic regulatory sequences are implicated in muscle cell determination then they should be expressed in the dermomyotome of the immature somite which gives rise to muscle precursor cells.(ABSTRACT TRUNCATED AT 400 WORDS)

The spatial and temporal expression of the dystrophin gene has been examined during mouse embryogenesis, using in situ hybridization on tissue sections with a probe from the 5' end of the dystrophin coding sequence. In striated muscle, dystrophin transcripts are detectable from about 9 d in the heart and slightly later in skeletal muscle. However, there is an important difference between the two types of muscle: the heart is already functional as a contractile organ before the appearance of dystrophin transcripts, whereas this is not the case in skeletal muscle, where dystrophin and myosin heavy chain transcripts are first detectable at the same time. In the heart, dystrophin transcripts accumulate initially in the outflow tract and, at later stages, in both the atria and ventricles. In skeletal muscle, the gene is expressed in all myocytes irrespective of fiber type. In smooth muscle dystrophin transcripts are first detectable from 11 d post coitum in blood vessels, and subsequently in lung bronchi and in the digestive tract. The other major tissue where the dystrophin gene is expressed is the brain, where transcripts are clearly detectable in the cerebellum from 13 d. High-level expression of the gene is also seen in particular regions of the forebrain involved in the regulation of circadian rhythms, the endocrine system, and olfactory function, not previously identified in this context. The findings are discussed in the context of the pathology of Duchenne muscular dystrophy.

We report results from a study of Hox-7 expression during mouse embryonic and fetal development and compare the localization of Hox-7 transcripts with those of the retinoic acid receptors. Transcripts were detected by in situ hybridization. Hox-7 expression occurs in (1) cephalic neural crest and its derivatives, (2) sites of ectomesodermal interaction, (3) extraembryonic tissues, and (4) endocardial cells. Hox-7 does not seem to be involved in defining rostrocaudal boundaries, but instead appears to be expressed along the proximodistal axes at these sites. We further investigated the active sites of morphogenesis, which involve an ectomesodermal interaction (e.g., limb buds, visceral arches), including genital tubercle and tail ridge. These are regions highly positive for Hox-7 transcripts, and many are known to be sites for the expression of gamma-retinoic acid receptors (RARs) and cellular retinoic acid binding proteins. Most regions that express Hox-7 are subregions of gamma-RAR expression. In the developing limb bud, expression of Hox-7 takes place in the interdigital region, where it overlaps areas of beta-RAR expression.