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The human TUPLE1 gene encodes a putative transcriptional regulator and maps to chromosome 22, and therefore may play a role in Di-George syndrome (DGS), relo-cardio-facial syndrome (VCFS), or a related pathology. The murine TUPLE1 gene has also been cloned and shows strong sequence similarity to TUPLE1. Comparative mapping is useful in the study of chromosome evolution and is sometimes able to indicate possible mouse mutations that are potential models of human genetic disorders. As TIPLE1 is a candidate gene for the haploinsufficient phenotype in DGS, we mapped TUPLE1 to mouse chromosome 16A-B1. 6 refs., 1 fig.

Polycystin-2 (PC-2), a protein encoded by PKD2 and involved in autosomal dominant polycystic kidney disease (ADPKD), is a non-selective cationic channel recently implicated in the function of primary cilia. We recently constructed a new animal model in the form of a transgenic mouse with a BAC-containing human PKD2 inserted in its genome. Two transgenic mouse lines overexpressing human PKD2 showed mitotic instability. Fibroblasts from these transgenic mouse lines have abnormal chromosomal numbers. These lines also have supernumerary centrosomes. PC-2 overexpression is associated with mitotic instability and centrosome overduplication. PC-2 therefore seems to play a role in centrosome duplication, and this hypothesis is being evaluated in other models.

TBX1 , encoding a T-box containing transcription factor, is the major candidate gene for del22q11.2 or DiGeorge syndrome, characterized by craniofacial and cardiovascular defects including tetralogy of Fallot and common arterial trunk. Mice lacking Tbx1 have severe defects in the development of pharyngeal derivatives including cardiac progenitor cells of the second heart field that contribute to the arterial pole of the heart. The outflow tract of Tbx1 mutant embryos is short and narrow resulting in common arterial trunk. Here we show by a series of genetic crosses using transgene markers of second heart field derived myocardium and coronary endothelial cells that a subdomain of myocardium normally observed at the base of the pulmonary trunk is reduced and malpositioned in Tbx1 mutant hearts. This defect is associated with anomalous coronary artery patterning. Both right and left coronary ostia form predominantly at the right/ventral sinus in mutant hearts, proximal coronary arteries coursing across the normally coronary free ventral region of the heart. We have identified Semaphorin3c as a Tbx1 -dependent gene expressed in subpulmonary myocardium. Our results implicate second heart field development in coronary artery patterning and provide new insights into the association between conotruncal defects and coronary artery anomalies.

SUMO-1, a ubiquitin-like protein, is covalently bound to many proteins, leading to chromatin inactivation and transcriptional repression. The high concentration of SUMO-1 on the XY body in rodents suggests that this protein has an important role in facultative heterochromatin organization. In human, the precise role of SUMO in chromatin/heterochromatin organization remains to be defined. Here we describe the SUMO-1 distribution, during human male meiosis, in relation to the formation of the different types of heterochromatin. We show that, during late pachynema, SUMO-1 appears on the constitutive heterochromatin, but is excluded from the XY body facultative heterochromatin. At the SUMO-1 labelled areas, the presence of HP1alpha protein, as well as of trimethylated H3-K9 and H4-K20 histone modifications, supports a role for SUMO-1 in constitutive heterochromatin organization. We also establish that, on the constitutive heterochromatin, H4-K20me3 staining progressively decreases as SUMO-1 staining increases, suggesting that core histone(s), and histone H4 in particular, are direct targets for sumoylation. Our results also suggest that, in the context of global histone H4 hyperacetylation that precedes the histone-to-protamine transition at postmeiotic stages of spermatogenesis, histone H4 sumoylation may represent an important epigenetic marker replacing methylation on the constitutive heterochromatin.

Unbalanced translocations, that involve the proximal chromosome 15 long arm and the telomeric region of a partner chromosome, result in a karyotype of 45 chromosomes with monosomy of the proximal 15q imprinted region. Here, we present our analysis of eight such unbalanced translocations that, depending on the parental origin of the rearranged chromosome, were associated with either Prader-Willi or Angelman syndrome. First, using FISH with specific BAC clones, we characterized the chromosome 15 breakpoint of each translocation and demonstrate that four of them are clustered in a small 460 kb interval located in the proximal 15q14 band. Second, analyzing the sequence of this region, we demonstrate the proximity of a low-copy repeat 15 (LCR15)-duplicon element that is known to facilitate recombination events at meiosis and to promote rearrangements. The presence, in this region, of both a cluster of translocation breakpoints and a LCR15-duplicon element defines a new breakpoint cluster (BP6), which, to our knowledge, is the most distal breakpoint cluster described in proximal 15q. Third, we demonstrate that the breakpoints for other rearrangements including large inv dup (15) chromosomes do not map to BP6, suggesting that it is specific to translocations. Finally, the translocation breakpoints located within BP6 result in very large proximal 15q deletions providing new informative genotype-phenotype correlations.

We have recently demonstrated that heterochromatin HP1 proteins are aberrantly distributed in lymphocytes of patients with immunodeficiency, centromeric instability and facial dysmorphy (ICF) syndrome. The three HP1 proteins accumulate in one giant body over the 1qh and 16qh juxtacentromeric heterochromatins, which are hypomethylated in ICF. The presence of PML (promyelocytic leukaemia) protein within this body suggests it to be a giant PML nuclear body (PML-NB). The structural integrity of PML-NBs is of major importance for normal cell functioning. Nevertheless, the structural organisation and the functions of these nuclear bodies remain unclear. Here, we take advantage of the large size of the giant body to demonstrate that it contains a core of satellite DNA with proteins being organised in ordered concentric layers forming a sphere around it. We extend these results to normal PML-NBs and propose a model for the general organisation of these structures at the G2 phase. Moreover, based on the presence of satellite DNA and the proteins HP1, BRCA1, ATRX and DAXX within the PML-NBs, we propose that these structures have a specific function: the re-establishment of the condensed heterochromatic state on late-replicated satellite DNA. Our findings that chromatin-remodelling proteins fail to accumulate around satellite DNA in PML-deficient NB4 cells support a central role for PML protein in this cellular function.

We report the case of a girl presenting with an unusual form of multiple joint fusion. Skeletal abnormalities consisted of radioulnar synostosis and vertebral fusions without any carpal, digital or tarsal involvement, and broad ribs and clavicles. Spinal X-rays were available from age 4 to 21, demonstrating that the spinal involvement was progressive and led to a complete anterior and lateral fusion of vertebrae. A complete sequencing of the NOGGIN gene failed to find any mutation. In addition, this girl was carrier of an apparently balanced reciprocal translocation t(10;20)(p11;p13). We investigated the role of the BMP2A gene as a potential candidate gene. Fluorescence in situ hybridization with YAC probes from chromosome 20 showed that the BMP2A gene was not disrupted by the translocation breakpoint.

We report the case of a boy whose karyotype at birth showed additional material on one chromosome 15. He underwent treatment for unilateral nephroblastoma at 6 years old. At 23 years old, he presented with body asymmetry, facial dysmorphism, arachnodactyly, severe scoliosis, and mental retardation. Molecular cytogenetic analyses of peripheral lymphocytes demonstrated a complex mosaic with three clones. A major cell lineage (68%) showed a chromosome 15 with additional material fused to its telomere long arm that was constituted by an inverted duplicated 15q24.3-qter segment. Therefore, the resulting add(15)(q) harbored an intrachromosomal triplication with the middle segment being inverted in orientation. A minor cell lineage (7%) showed an abnormal chromosome 3 resulting from a telomeric fusion between its short arm and an inverted duplicated 15q24.3-qter segment. The third cell lineage (25%) showed a normal 46,XY constitution. Finally, this resulted in tetrasomy for the distal 15q24.3-qter region in 75% of the patient's lymphocytes. To our knowledge, distal 15q tetrasomy is rare and only eight cases have been reported in the literature, all due to a supernumerary analphoid marker consisting of an inverted duplication. We report here the first observation of distal 15q tetrasomy associated with a 46 chromosomes constitution. We compare the phenotype of our patient to previous cases of distal tetrasomy 15q and discuss the mechanisms underlying this chromosomal rearrangement.

The Chudley-Lowry syndrome (ChLS, MIM 309490) is an X-linked recessive condition characterized by moderate to severe mental retardation, short stature, mild obesity, hypogonadism, and distinctive facial features characterized by depressed nasal bridge, anteverted nares, inverted-V-shaped upper lip, and macrostomia. The original Chudley-Lowry family consists of three affected males in two generations. Linkage analysis had localized the gene to a large interval, Xp21-Xq26 and an obligate carrier was demonstrated to have highly skewed X inactivation. The combination of the clinical phenotype, consistent with that of the patients with ATR-X syndrome, the skewed X-inactivation pattern in a carrier female, as well as the mapping interval including band Xq13.3, prompted us to consider the XNP/ATR-X gene being involved in this syndrome. Using RT-PCR analysis, we screened the entire XNP/ATR-X gene and found a mutation in exon 2 (c.109C > T) giving rise to a stop codon at position 37 (p.R37X). Western blot and immunocytochemical analyses using a specific monoclonal antibody directed against XNP/ATR-X showed the protein to be present in lymphoblastoid cells from one affected male, despite the premature stop codon. To explain these discordant results, we further analyzed the 5' region of the XNP/ATR-X gene and found three alternative transcripts, which differ in the presence or absence of exon 2, and the length of exon 1. Our data suggest that ChLS is allelic to the ATR-X syndrome with its less severe phenotype being due to the presence of some XNP/ATR-X protein.

Collagen IV is a major structural component of basement membranes. In the glomerular basement membrane (GBM) of the kidney, the alpha3, alpha4, and alpha5(IV) collagen chains form a distinct network that is essential for the long-term stability of the glomerular filtration barrier, and is absent in most patients affected with Alport syndrome, a progressive inherited nephropathy associated with mutation in COL4A3, COL4A4, or COL4A5 genes. To investigate, in vivo, the regulation of the expression, assembly, and function of the alpha3alpha4alpha5(IV) protomer, we have generated a yeast artificial chromosome transgenic line of mice carrying the human COL4A3-COL4A4 locus. Transgenic mice expressed the human alpha3 and alpha4(IV) chains in a tissue-specific manner. In the kidney, when expressed onto a Col4a3(-/-) background, the human alpha3(IV) chain restored the expression of and co-assembled with the mouse alpha4 and alpha5(IV) chains specifically at sites where the human alpha3(IV) was expressed, demonstrating that the expression of all three chains is required for network assembly. The co-assembly of the human and mouse chains into a hybrid network in the GBM restores a functional GBM and rescues the Alport phenotype, providing further evidence that defective assembly of the alpha3-alpha4-alpha5(IV) protomer, caused by mutations in any of the three chains, is the pathogenic mechanism responsible for the disease. This line of mice, humanized for the alpha3(IV) collagen chain, will also provide a valuable model for studying the pathogenesis of Goodpasture syndrome, an autoimmune disease caused by antibodies against this chain.

We describe here a patient with intrachromosomal triplication 15q11-q13, a rare chromosomal event associated with severe mental retardation and intractable epilepsy. Cytogenetic studies including FISH on interphasic nuclei showed that the middle segment of the triplication was inverted in orientation. Molecular analyses demonstrated that the rearrangement was of maternal origin. Based on these cytogenetic and molecular data and those of the nine cases reported in the literature, we discuss the mechanistic origins of these triplications. We present several arguments for the mechanism involving two U-type exchanges occurring simultaneously at the pachytene stage of meiosis.

Comparing the genomes of the great apes and human should provide novel information concerning the origins of humankind. Relative to the great apes, the human karyotype has one fewer chromosome pair, as human chromosome 2 derived from the telomeric fusion of two ancestral primate chromosomes. To identify the genomic rearrangements that accompanied human speciation, we initiated a comparative study between human, chimpanzee, and gorilla. Using the HAPPY mapping method, an acellular adaptation of the radiation hybrid method, we mapped a few hundred markers on the human, chimpanzee, and gorilla genomes. This allowed us to identify several chromosome rearrangements, in particular a pericentric inversion and a translocation. We precisely localized the synteny breakpoint that led to the formation of human chromosome 2. This breakpoint was confirmed by FISH mapping.

Mammalian telomeres are composed of long arrays of TTAGGG repeats that form a nucleoprotein complex which protects the chromosome ends. Human telomere function is known to require two TTAGGG repeat factors, TRF1 and TRF2, and several interacting proteins, but the mechanism by which the DNA/protein complex prevents end to end fusion in vivo has not been elucidated. In order to better understand the role of specific telomere-associated proteins in the organisation of chromosome ends, we have studied a patient with a rare chromosome rearrangement that has given rise to an interstitial telomere. Using specific antibodies and immuno-FISH on unfixed metaphase chromosomes, we show that the proteins TRF2 and TIN2 (TIN2 interacts with TRF1) co-localise with the interstitial TTAGGG repeats. Our results demonstrate, for the first time in humans, that TRF2 and TIN2 proteins associate with interstitial duplex TTAGGG repeats, in vivo. They confirm that double stranded-telomeric repeats, even when complexed with specific proteins, are not sufficient to create a functional telomere. Finally, they suggest a possible role for proteins in stabilising interstitial TTAGGG repeats.

The HP1 class of chromobox (Cbx) genes encode an evolutionarily conserved family of proteins involved in the packaging of chromosomal domains into a repressive heterochromatic state. The murine Cbx5, Cbxl and Cbx3 genes encode the three mouse HP1 proteins, mHPlot, -β and -γ respectively. Here, we report the cloning of the mouse Cbx31 HPlγ gene and the chromosomal localisation of Cbx3 and three Cfo3-related pseudogenes. The Cbx3 structural gene is located on mouse Chromosome 6, close to the Hoxa cluster. Two Cbx3 processed pseudogenes are separated by just 300 bp and are arranged in a head-to-tail configuration on Chromosome 13 while a third pseu-dogene is found on mouse Chromosome 4. The genomic intron-exon arrangement of Cbx3 is different from the conserved organisation of three other mammalian HP1 genes, Cbxl (mHPip), CBX3 (hHPly), and Cbx5 (mHPla) in that Cbx3 lacks an intron that is present in the others.

Striatin is an intracellular protein characterized by four protein-protein interaction domains, a caveolin-binding motif, a coiled-coil structure, a calmodulin-binding domain, and a WD repeat domain, suggesting that it is a signaling or a scaffold protein. Down-regulation of striatin, which is expressed in a few subsets of neurons, impairs the growth of dendrites as well as rat locomotor activity (Bartoli, M., Ternaux, J. P., Forni, C., Portalier, P., Salin, P., Amalric, M., and Monneron, A. (1999) J. Neurobiol. 40, 234-243). Zinedin, a "novel" protein described here, and SG2NA share with striatin identical protein-protein interaction domains and the same overall domain structure. A phylogenetic analysis supports the hypothesis that they constitute a multigenic family deriving from an ancestral gene. DNA probes and antibodies raised against specific domains of each protein showed that zinedin is mainly expressed in the central nervous system, whereas SG2NA, of more widespread occurrence, is mainly expressed in the brain and muscle. All three proteins are both cytosolic and membrane-bound. All three bind calmodulin in the presence of Ca(2+). In rat brain, SG2NA and striatin are generally not found in the same neurons. Both localize to the soma and dendrites, suggesting that they share a similar type of addressing and closely related functions.

Chemokines are key regulators of migration in lymphoid tissues. In the thymus, maturing thymocytes move from the outer capsule to the inner medulla and thereby interact with different types of stromal cells that control their maturation and selection. In the process of searching for molecules specifically expressed at different stages of mouse thymic differentiation, we have characterized the cDNA coding for the thymus-expressed chemokine (TECK) and its receptor CCR9. The TECK receptor gene was isolated and shown to be localized on the mouse chromosome 9F1-F4. Thymic dendritic cells have been initially thought to be a prevalent source of TECK. In contrast, our results indicate that thymic epithelial cells constitute the predominant source of TECK. Consistent with the latter distribution, the TECK receptor is highly expressed by double-positive thymocytes, and TECK can chemoattract both double-positive and single-positive thymocytes. The TECK transcript is also abundantly expressed in the epithelial cells lining the small intestine. In conclusion, the interplay of TECK and its receptor CCR9 is likely to have a significant role in the recruitment of developing thymocytes to discrete compartments of the thymus.

We used multicolour fluorescence in-situ hybridization on air-dried pachytene nuclei to analyse the structural and functional domains of the sex vesicle (SV) in human, chimpanzee and mouse. The same technology associated with 3-dimensional analysis was then performed on human and mouse pachytene nuclei from cytospin preparations and tissue cryosections. The human and the chimpanzee SVs were very similar, with a consistently small size and a high degree of condensation. The mouse SV was most often seen to be large and poorly condensed, although it did undergo progressive condensation during pachynema. These results suggest that the condensation of the sex chromosomes is not a prerequisite for the formation of the mouse SV, and that a different specific mechanism could be responsible for its formation. We also found that the X and Y chromosomes are organized into two separate and non-entangled chromatin domains in the SV of the three species. In each species, telomeres of the X and Y chromosomes remain clustered in a small area of the SV, even those without a pseudoautosomal region. The possible mechanisms involved in the organization of the sex chromosomes and in SV formation are discussed.

A degenerate polymerase chain reaction (PCR) homology screening procedure was applied to rat brain cDNA in order to identify novel genes belonging to the amiloride-sensitive Na+ channel and degenerin (NaC/DEG) family of ion channels. A single gene was identified that encodes a protein related to but clearly different from the already cloned members of the family (18-30 % amino acid sequence identity). Phylogenetic analysis linked this protein to the group of ligand-gated channels that includes the mammalian acid-sensing ion channels and the Phe-Met-Arg-Phe-amide (FMRFamide)-activated Na+ channel. Expression of gain-of-function mutants after cRNA injection into Xenopus laevis oocytes or transient transfection of COS cells induced large constitutive currents. The activated channel was amiloride sensitive (IC50, 1.31 μm) and displayed a low conductance (9-10 pS) and a high selectivity for Na+ over K+ (ratio of the respective permeabilities, PNa+/PK+≥ 10), all of which are characteristic of NaC/DEG channel behaviour. Northern blot and reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed a predominant expression of its mRNA in the small intestine, the liver (including hepatocytes) and the brain. This channel has been called the brain-liver-intestine amiloride-sensitive Na+ channel (BLINaC). Corresponding gain-of-function mutations in Caenorhabditis elegans degenerins are responsible for inherited neurodegeneration in the nematode. Besides the BLINaC physiological function that remains to be established, mutations in this novel mammalian degenerin-like channel might be of pathophysiological importance in inherited neurodegeneration and liver or intestinal pathologies. The amiloride-sensitive Na+ channel and degenerin family (NaC/DEG) is an expanding family of cationic channels associated with very diverse functions in many organisms (Barbry & Hofman, 1997; Horisberger, 1998). Despite this functional diversity, all these channels share common properties including permeability to Na+, inhibition by the diuretic amiloride and voltage-independent gating. The NaC/DEG family includes channels that are constitutively active, like the epithelial Na+ channel (ENaC) involved in taste perception and Na+ homeostasis (Garty & Palmer, 1997) or the Drosophila gonad-specific amiloride-sensitive Na+ channel (dGNaC1) (Darboux et al. 1998a), also named ripped pocket (RPK) (Adams et al. 1998a), suggested to participate in gametogenesis or early embryonic development. The NaC/DEG family also comprises ligand-gated channels like the Phe-Met-Arg-Phe-NH2 (FMRFamide) peptide-gated Na+ channel (FaNaC) cloned from the snail Helix aspersa nervous system (Lingueglia et al. 1995; Cottrell, 1997; Zhainazarov & Cottrell, 1998) where it participates in neuromodulation or the mammalian acid-sensing ion channels (ASICs) recently characterized from mammalian brain and sensory neurons (Waldmann et al. 1997b; Waldmann & Lazdunski, 1998) where they are thought to play an important role in the pain accompanying tissue acidosis. Stretch is another stimulus that has been proposed to activate members of this ion channel family cloned from the nematode Caenorhabditis elegans and from Drosophila melanogaster (Adams et al. 1998a; Darboux et al. 1998b). MEC-4 (Driscoll & Chalfie, 1991) and MEC-10 (Huang & Chalfie, 1994) are expressed in the mechanosensory neurons of Caenorhabditis elegans where they are involved in mechanoperception. Recently, novel members that are required for co-ordinated movement have been identified in motor neurons (UNC-8, DEL-1; Tavernarakis et al. 1997) and muscle (UNC-105; Liu et al. 1996). However, the modes of activation of these channels remain uncertain since only an heterologous expression of gain-of-function mutants of UNC-105 has been described to date (Garcia-Anoveros et al. 1998). One interesting feature of these proteins is that gain-of-function mutations in their genes (deg-1, mec-4, mec-10, unc-8) cause the degeneration of some or all the neurons in which they are expressed, hence the name degenerin. Remarkably, extrapolation in some mammalian members of this ion channel family (i.e. ASIC1, Bassilana et al. 1997, and ASIC2, previously named MDEG1, Waldmann et al. 1996) of a particular point mutation associated with gain-of-function and neuronal degeneration in Caenorhabditis elegans degenerins, also causes constitutive channel activation. Taken together, the implication of degenerin gain-of-function mutations in inherited neurodegeneration in the nematode Caenorhabditis elegans and the existence of gain-of-function mutations in mammalian neuronal NaC/DEG channels that can cause cell death (Waldmann et al. 1996) suggest that degenerin-like channels might also be involved in mammalian forms of neurodegeneration or muscle pathologies. The number of mammalian NaC/DEG genes remains quite low compared with the more than 15 as yet uncharacterized degenerin homologues found in the Caenorhabditis elegans genome. In the present study, we report the cloning of a novel degenerin-like ion channel gene in rat and mouse and determine its tissue distribution and the biophysical properties of gain-of-function mutants.

International audience; The mouse heat shock factor 2 (HSF2) cDNA was previously cloned by homology to HSF1, the heat shock factor involved in the cellular response to stress [Sarge, K.D., Zimarino, V., Holm, K., Wu, C., Morimoto, R.I., Cloning and characterization of two mouse heat shock factors with distinct inducible and constitutive DNA-binding ability. Genes Dev. 5 (1991) 1902-1911]. HSF2 is active in restricted cell types during pre- and post-implantation stages of development, and only in male germ cells of adult mice. However, the function of this factor remains elusive. We report here the cloning of the mouse Hsf2 gene and its genomic structure. We show that the gene is composed of 13 exons of variable sizes spanning at least 43kb in the genome. The transcription start site has been determined, and upstream sequences with promoter activity have been identified by their ability to direct the expression of a luciferase reporter gene in transfected cells. A preliminary analysis of the proximal promoter sequence determined that the TATA box is absent, but that a GC-rich region with several potential binding sites for transcription factors is present. The gene has been mapped to mouse chromosome 10 by in-situ hybridization on metaphase chromosomes.

Using fluorescent in-situ hybridization, we investigated the positioning of different human bivalents at the pachytene stage of normal male meiosis. We showed that, in about 35% of nuclei, the pericentromeric region of bivalent 15 is closely associated with the sex vesicle (SV). This behaviour may be linked to the presence of three domains in the pericentromeric region of chromosome 15: a large imprinted domain, a nucleolar organizing region (NOR), and a heterochromatic block. In order to define the domains of chromosome 15 involved in this association, we analysed the meiotic behaviour of other bivalents with similar domains: human bivalent 11 and mouse bivalent 7, bearing imprinted domains, other human acrocentric bivalents bearing a NOR, and the human bivalents 1, 9 and 16 containing a heterochromatic region. None of these bivalents were as frequently associated with the SV as the human bivalent 15. Nevertheless, we suggest that the bivalent 15 heterochromatin may be responsible for the association because of two properties: its telomeric location on chromosome 15 and its strong sequence homology with the Yq heterochromatin. This phenomenon could explain the high frequency of translocations between the chromosome 15 and the X or Y chromosomes.

D52 proteins are emerging as signalling molecules which may be regulators of cell proliferation. Having previously reported the existence of the human D52 gene family, comprising the hD52 and hD53 genes expressed in human breast carcinoma, we report the identification of a novel human gene hD54 ( TPD52L2 ), which represents a third D52 gene family member. In situ mapping placed the hD54 gene on human chromosome 20q13.2–q13.3, a localization distinct from those of both hD52 and hD53 genes. The identified hD54 cDNAs predicted three hD54 isoforms, suggesting that alternatively-spliced transcripts may be produced from D52-like genes. This was confirmed by directly sequencing reverse transcriptase–polymerase chain reaction (RT–PCR) products amplified from D52-like gene transcripts expressed in developing and adult rat tissues, and by performing sequence analyses of the expressed sequence tag divisions of nucleotide databases. Alternative splicing of sequences encoding two regions, termed ins2 and ins3, was identified in one or more D52-like genes, with these alternative splicing events being differentially regulated. The functional consequences of alternative splicing were examined by characterizing the protein–protein interactions mediated by a truncated hD53 isoform within the yeast two-hybrid system. This hD53 isoform displayed altered interaction capabilities with respect to those of full-length hD53, suggesting that alternative splicing within the D52 gene family functions in part to alter the protein–protein interaction capabilities of encoded isoforms.

We report here the identification of two distinct human cDNAs, called VNN1 and VNN2, related to the recently described mouse Vanin-1 molecule involved in lymphocyte migration (M. Aurrand-Lions et al., 1996, Immunity 5: 391-405). Tissue distribution of the expression of these two human Vanin-like genes is differential. Since Vanin-1 shares significant homologies with human biotinidase (BTD), we describe here a new family of related genes including at least four members: mouse Vanin-1, VNN1, VNN2, and BTD. We have mapped the murine locus encompassing the Vanin-1 gene on mouse chromosome 10 in position A2B1. The two human Vanin-like genes are closely linked, since they were found on the same YAC clone and colocalized on human chromosome 6q23-q24 known to contain several genetic alterations linked to the progression of metastatic human cancers.

We have identified a novel lysosome-associated membrane glycoprotein localized on chromosome 3q26.3-q27, DC-LAMP, which is homologous to CD68. DC-LAMP mRNA is present only in lymphoid organs and DC. A specific MAb detects the protein exclusively in interdigitating dendritic cells. Expression of DC-LAMP increases progressively during in vitro DC differentiation, but sharply upon activation with LPS, TNFα, or CD40L. Confocal microscopy confirmed the lysosomal distribution of the protein. Furthermore, DC-LAMP was found in the MHC class II compartment immediately before the translocation of MHC class II molecules to the cell surface, after which it concentrates into perinuclear lysosomes. This suggests that DC-LAMP might change the lysosome function after the transfer of peptide–MHC class II molecules to the surface of DC.

The murine epididymal retinoic acid- binding protein (mE-RABP) is specifically synthesized in the mouse mid/distal caput epididymidis and secreted in the lumen. In this report, we have demonstrated by Southern blot analysis of genomic DNA that mE-RABP is encoded by a single-copy gene. A mouse 129/SvJ genomic bacterial artificial chromosome (BAC) library was screened using a cDNA encoding the minor form of mE-RABP. One positive BAC clone was characterized and sequenced to determine the nucleotide sequence of the entire mE-RABP gene. The molecular cloning of the mE-RABP gene completes the characterization of the 20.5-kDa-predicted preprotein lead- ing to the minor and major forms of mE-RABP. Comparison of the DNA sequence of the promoter and coding regions with that of the rat epididymal secretory protein I (ESP I) gene showed that the mE-RABP gene is the orthologue of the ESP I gene that encodes a rat epididymal retinoic acid-binding protein. Several regulatory elements, including a putative androgen receptor binding site, ''CACCC-boxes,'' NF-1, Oct-1, and SP-1 recognition sites, are conserved in the proximal promoter. Analysis of the nucleotide sequence of the mE-RABP gene revealed the presence of seven exons and showed that the genomic organization is highly related to other genes encoding lipocalins. The mE-RABP gene was mapped by fluorescent in situ hybridization to the (A3-B) region of the murine chromosome 2. Our data, combined with that of others, suggest that the proximal segment of the mouse chromosome 2 may be a rich region for genes encoding lipocalins with a genomic organization highly related to the mE-RABP gene. Mol. Reprod. Dev. 50:387-395, 1998. r 1998 Wiley-Liss, Inc.

In mammals, anti-Mullerian hormone (AMH) is produced by Sertoli cells from the onset of testicular differentiation and by granulosa cells only after birth. SOX9, a transcription factor related to the testis-determining factor SRY, is expressed in mouse testis 1 day before AMH. To determine the relationship between AMH and SOX9 in birds, we cloned the AMH promoter in search of SOX9 response elements, and we compared the expression of AMH and SOX9 in the gonads of chick embryos using in situ hybridization. Potential SOX response elements were found in the AMH promoter; however, AMH is expressed in both sexes at stage 25, 1 day before the first SOX9 transcripts appear in the male gonads. SOX9 is never expressed in the female. These results do not support the hypothesis that SOX9 could trigger the expression of testicular AMH in the chick but does not exclude a later role in testis development.

Rat striatin, a recently discovered calmodulin-binding protein belonging to the WD repeat family, is expressed in neurons, mostly in the striatum and motor and olfactory systems. Striatin is localized in the somato-dendritic compartment of neurons, mainly in the spines. It may play a role in dendritic Ca2+signaling. Here we report the cloning and sequencing of human striatin cDNA (HGMW-approved symbol STRN), the localization of the gene to chromosome 2p22–p21, and its preferential expression in brain. The human cDNA sequence is predicted to encode a 780-amino-acid protein possessing eight WD repeats. Striatin is highly conserved between rat and human with 96% identity and 98% similarity at the amino acid level. Since theCaenorabditis elegansgenome also contains a closely related striatin coding sequence, the function of striatin is likely to be well conserved.

The ADAM (A Disintegrin And Metalloprotease) family of cell-surface proteins may have an important role in cellular interactions and in modulating cellular responses. In this report we describe a novel, secreted form of human ADAM 12 (meltrin alpha), designated ADAM 12-S (S for short), and a larger, membrane-bound form designated ADAM 12-L (L for long form). These two forms arise by alternative splicing of a single gene located on chromosome 10q26. Northern blotting demonstrated that mRNAs of both forms are abundant in human term placenta and are also present in some tumor cell lines. The ADAM 12-L transcript can also be detected in normal human adult skeletal, cardiac, and smooth muscle. Human A204 embryonal rhabdomyosarcoma cells that do not differentiate into muscle cells and do not express any form of ADAM 12 were stably transfected with an ADAM 12-S minigene encoding the disintegrin domain, the cysteine-rich domain, and the unique 34 amino acid carboxyl terminus. Nude mouse tumors derived from these transfected cells contained ectopic muscle cells of apparent mouse origin as shown by species-specific markers. These results may have potential applications in the development of muscle-directed gene and cell therapies.

The human Lasp-1 (LIM and SH3 protein) gene was previously identified by differential screening of a breast cancer-derived metastatic lymph node cDNA library. It was located on the q12–q21 region of human chromosome 17 and was shown to be amplified and overexpressed in 12% of breast tumours. Lasp-1 defines a new LIM-protein subfamily, as it associates a C-terminal Src homology 3 (SH3) domain to a N-terminal LIM motif. In this study, the isolation and characterization of the cDNA encoding the mouse Lasp-1 protein are described, and it is shown to be highly conserved with its human counterpart. In addition to the LIM and SH3 domains, both human and mouse Lasp-1 contain an actin-binding domain. The mouse gene was mapped by in situ hybridization to the 11C–11D region of chromosome 11. Northern blot analysis shows that this gene is expressed from 7.5 to 17.5 days post-coitum of mouse embryogenesis and in almost all adult tissues.

Members of the Rho GTPase family regulate the organization of the actin cytoskeleton in response to extracellular growth factors. We have identified three proteins that form a distinct branch of the Rho family: Rnd1, expressed mostly in brain and liver; Rnd2, highly expressed in testis; and Rnd3/RhoE, showing a ubiquitous low expression. At the subcellular level, Rnd1 is concentrated at adherens junctions both in confluent fibroblasts and in epithelial cells. Rnd1 has a low affinity for GDP and spontaneously exchanges nucleotide rapidly in a physiological buffer. Furthermore, Rnd1 lacks intrinsic GTPase activity suggesting that in vivo, it might be constitutively in a GTP-bound form. Expression of Rnd1 or Rnd3/RhoE in fibroblasts inhibits the formation of actin stress fibers, membrane ruffles, and integrin-based focal adhesions and induces loss of cell–substrate adhesion leading to cell rounding (hence Rnd for “round”). We suggest that these proteins control rearrangements of the actin cytoskeleton and changes in cell adhesion.

Dividing cells expressing the Herpes simplex type 1 thymidine kinase (TK) can be killed upon ganciclovir treatment. Likewise, conditional cell knock-out can be obtained in transgenic mice expressing a TK gene placed under the control of tissue-specific regulatory sequences. Such animals provide powerful experimental systems for assessing the functional role of specific cell populations through their time-controlled ablation. However, whatever the regulatory sequences used, a leaky toxic overexpression of TK in testis renders male TK-transgenic mice sterile and prevents the generation of homozygous TK-expressing animals. To solve this problem, we designed a truncated TK variant (ΔTK) not expressed in the testis. We generated transgenic mice expressing ΔTK under the control of lymphocyte-specific regulatory sequences derived from the CD4 gene. The ΔTK protein expressed in T-lymphocytes allowed the conditional ablation of activated T-cells in vitro and in vivo. Importantly, for one transgenic line we could generate fertile homozygous mice harboring a functional ΔTK transgene. ΔTK should thus dramatically facilitate the development of transgenic mice expressing a conditional suicide gene.

Autoimmune diabetes in both the human and the nonobese diabetic mouse has elaborate genetics; in the latter case, the disease is influenced by at least 15–20 loci. We anticipated that the genetics would be simpler in the BDC2.5 T cell receptor transgenic mouse model of diabetes, wherein many T cells express a particular diabetogenic specificity. Initiation of insulitis in this model was the same on the two genetic backgrounds analyzed, but the kinetics and penetrance of diabetes were strikingly different, permitting us to focus on genetic influences during a defined window of disease progression. The differences correlated with variations in five genomic intervals, certain ones of which have been previously implicated in susceptibility to autoimmune disease. This reductionist approach indeed simplified the analysis of diabetes susceptibility loci.

Two novel mouse genes, Ebf2 and Ebf3, have been identified which show high similarity to the rodent Ebf/Olf-1 and the Drosophila collier genes. The strong conservation of the protein regions corresponding to the DNA binding and dimerisation domains previously defined in Ebf strongly suggests that Ebf2 and Ebf3 also constitute DNA sequence-specific transcription factors. Determination of the chromosomal locations of the two genes indicated that the different members of this novel mouse multigene family are not clustered. A detailed analysis of the expression of each of the three Ebf genes in the developing central nervous system revealed partially overlapping patterns with two salient features: 1) In the region extending from the midbrain to the spinal cord, the expression of the three genes correlated with neuronal maturation, with a general activation in early post-mitotic cells, followed by specific patterns of extinction also consistent with the neurogenic gradient. 2) In the forebrain area, although the patterns of expression of the Ebf genes also reflected neuronal maturation, they appeared in addition to be region specific. These data suggest that Ebf genes may be involved in the control of neuronal differentiation in the CNS and in enforcing regional diversity in populations of post-mitotic forebrain neurons. Dev. Dyn. 1997;210:191–205. © 1997 Wiley-Liss, Inc.

The murine Sycp1 gene is expressed at the early stages of meiosis. We show that it is composed of a number of small exons and localized on mouse chromosome 3. In the laboratory strains, two retrogenes were also identified. The first one (Sycp1-ps1), on chromosome 7, has accumulated point mutations and deletions and is not transcribed. A second retrogene (Sycp1-ps2), on chromosome 8, is inserted within the continuity of a moderately repeated element, in an intron of another gene (Cad11). The two retroposition events can be dated to distinct periods in the evolution of the Muridae. Sycp1-ps2 has kept features indicative of a relatively recent origin, namely a nearly intact coding region, a poly(A) tail, and 14-bp terminal repeats. Its recent origin was confirmed by the fact that it is found in all the laboratory strains of mice, but neither in a recent isolate from Mus musculus domesticus wild stocks nor in the closely related subspecies M. musculus musculus, M. m. molossinus, M. m. castaneus, and M. m. bactrianus. Appearance of the more ancient Sycp1-ps1 retrogene is concomitant with the radiation of the genus. It is present in various Mus species (M. spretus, M. spicilegus, M. macedonicus, and M. cookii), but neither in the rat nor in the more closely related Pyromis genus. Transposition of retrotranscripts during meiosis and their hereditary establishment thus appear to occur relatively frequently. They may, therefore, play a significant role in the evolutionary process.

RP105 is a 105-kDa type I membrane protein of the leucine-rich repeat (LRR) family. Anti-RP105 sensitizes B cells to antigen-receptor-mediated apoptosis, but protects B cells from radiation-induced apoptosis and stimulates B cell proliferation. The sequence of the mouse RP105 has been reported. Here, we report the characterization of the human RP105. The 2.6-kb cDNA encodes a protein of 661 amino acids which displays 78% homology with mouse RP105. The 22 LRR and the 9 potential N-linked glycosylation sites within the extracellular region are conserved. While previous studies have shown that RP105 is expressed on surface IgM+IgD+2 B cells in mice, human RP105 was shown to be expressed on all subsets of mature B cells and dendritic cells. Human RP105 gene was mapped to the long arm of chromosome 5, where numerous cytokines and receptors have been localized.

Secretory phospholipases A2(sPLA2s) represent a rapidly expanding family of structurally related enzymes found in mammals as well as in insect and snake venoms. In this report, a cDNA coding for a novel sPLA2 has been isolated from human fetal lung, and its gene has been mapped to chromosome 16p13.1-p12. The mature sPLA2protein has a molecular mass of 13.6 kDa, is acidic (pI 5.3), and made up of 123 amino acids. Key structural features of the sPLA2include: (i) a long prepropeptide ending with an arginine doublet, (ii) 16 cysteines located at positions that are characteristic of both group I and group II sPLA2s, (iii) a C-terminal extension typical of group II sPLA2s, (iv) and the absence of elapid and pancreatic loops that are characteristic of group I sPLA2s. Based on these structural properties, this sPLA2 appears as a first member of a new group of sPLA2s, called group X. A 1.5-kilobase transcript coding for the human group X (hGX) sPLA2 was found in spleen, thymus, and peripheral blood leukocytes, while a less abundant 0.8-kilobase transcript was detected in the pancreas, lung, and colon. When the hGX sPLA2cDNA was expressed in COS cells, sPLA2 activity preferentially accumulated in the culture medium, indicating that hGX sPLA2 is an actively secreted enzyme. It is maximally active at physiological pH and with 10 mm Ca2+. hGX sPLA2 prefers phosphatidylethanolamine and phosphatidylcholine liposomes to those of phosphatidylserine.

Resume Les principales caracteristiques du syndrome de Smith-Magenis sont des criteres mineurs faciaux, une brachydactylie, un retard psychomoteur avec retard de langage et des troubles comportementaux. Ce syndrome est du a une microdeletion du chromosome 17p11.2. Observation. — Une enfant a consulte a l'âge de 7 ans pour un retard psychomoteur. L'examen notait une dysmorphie faciale discrete (brachycephalie, hypoplasie de l'etage moyen de la face, elargissement de la racine du nez, hypoplasie malaire), une brachydactylie, une hyporeflexie osteotendineuse et une voix rauque. Une surdite moderee, des troubles du comportement et des anomalies du sommeil etaient notes. L'etude chromosomique sur lymphocytes a montre une deletion de la sous-bande 17p11.2. L'analyse moleculaire a confirme la deletion par perte de l'allele maternel. Conclusion. — Le syndrome de Smith-Magenis associe des signes cliniques discrets et reste probablement volontiers meconnu. Son diagnostic repose sur l'etude chromosomique.

A subtractive cloning procedure was used to characterize the molecular changes involved in transformation of normal myoblasts to rhabdomyosarcoma (RMS) cells. Here we describe the cloning of DRAL, a novel LIM-domain protein expressed in primary myoblasts but down-regulated in the RMS cell line RD. DRAL is a LIM-only protein with five LIM domains whereby one LIM domain consists only of the second half of the consensus motif. Interestingly, down-regulation of DRAL was not confined to the RD RMS cells, but was a phenomenon extended to other RMS cell lines of both embryonal and alveolar subtype, and to some breast cancer cell lines. Analysis of the expression pattern in normal human tissues revealed that DRAL is expressed at high levels in the heart, suggesting an important function in the specification of the terminally differentiated phenotype of heart muscle cells. Immunofluorescence studies using an antibody directed against recombinant DRAL localized the protein predominantly in the nucleus of cultured cells. On the basis of these results, we conclude that down-regulation of DRAL correlates with the tumor phenotype of RMS cells.

In a 15 year old girl, referred for growth retardation, conventional cytogenetic analysis detected an abnormal 15q+ chromosome with extra material in the proximal region, inherited from her father and grandfather. Using various molecular cytogenetic techniques, including comparative genomic hybridisation, we showed that the extra chromatin resulted from in situ amplification of DNA sequences located between the loci D15Z1 and D15S18. On the basis of the clinical features of our patient and the late replication of the large amplified region, we searched for functional modifications in the adjacent Prader-Willi syndrome region.

Several lines of evidence indicate a crucial role for unconventional myosins in the function of the sensory hair cells of the inner ear. We report here the characterization of the cDNAs encoding two unconventional type I myosins from a mouse cochlear cDNA library. The first cDNA encodes a putative protein named Myo1c, which is likely to be the murine orthologue of the bullfrog myosin I{beta} and which may be involved in the gating of the mechanotransduction channel of the sensory hair cells. This myosin belongs to the group of short-tailed myosins I, with its tail ending shortly after a polybasic, TH-1-like domain. The second cDNA encodes a novel type I myosin Myo1f which displays three regions: a head domain with the conserved ATP- and actin-binding sites, a neck domain with a single IQ motif, and a tail domain with the tripartite structure initially described in protozoan myosins I. The tail of Myo1f includes (1) a TH-1 region rich in basic residues, which may interact with anionic membrane phospholipids; (2) a TH-2 proline-rich region, expected to contain an ATP-insensitive actin-binding site; and (3) an SH-3 domain found in a variety of cytoskeletal and signaling proteins. Northern blot analysis indicated that themore » genes encoding Myo1c and Myo1f display a widespread tissue expression in the adult mouse. Myo1c and Myo1f were mapped by in situ hybridization to the chromosomal regions 11D-11E and 17B-17C, respectively. The human orthologuous genes MYO1C and MYO1F were also characterized, and mapped to the human chromosomal regions 17p13 and 19p13.2- 19p1.3.3, respectively. 45 refs., 5 figs., 2 tabs.« less

Using antibody 2H9 from our heterogeneous nuclear ribonucleoproteins (anti-hnRNP) monoclonal antibody library, we previously showed in HeLa cells that a 35-37-kDa protein doublet switches from the hnRNP complexes to the nuclear matrix following a 10-min heat shock at 45°C (1 Lutz, Y., Jacob, M., and Fuchs, J. P. (1988) Exp. Cell Res. 175, 109-124). cDNA cloning and sequencing revealed an hnRNP protein (2H9) which is a new member of the hnRNP F, H/H′ family. Protein 2H9 displays two consensus sequence-type RNA binding domains (CS-RBD) showing 80-90% homology with two of the three CS-RBDs of hnRNP F and H/H′. Another common feature is the presence of two glycine/tyrosine-rich auxiliary domains located at the C terminus and between the two CS-RBDs. At the functional level we show that specific anti-2H9 peptide antibodies can directly inhibit an in vitro splicing system. Moreover, the 2H9 protein doublet is no more present in nuclear extracts from such briefly stressed cells, which interestingly correlates with the inability of these extracts to catalyze in vitro splicing reactions. Taken together, our data suggest that these proteins are involved in the splicing process and also participate in early heat shock-induced splicing arrest by transiently leaving the hnRNP complexes. These 2H9 proteins, which are encoded by a single gene located on human chromosome 10, were also found to be associated with nuclear bodies in situ.

Prader-Willi syndrome (PWS) is a neurogenetic disorder resulting from the loss of paternal expression of gene(s) localized in the 15q11-q12 region. A new human gene encoding a putative protein with high homology to the mouse NECDIN protein has recently been characterized and mapped to chromosome 15q11-q12. It is expressed from the paternal allele only, suggesting its potential involvement in PWS. We now report the localization of the mouse Necdin gene in a region of conserved synteny to the human PWS region. We demonstrate the paternal specific expression of Necdin in the mouse central nervous system, and show that parental alleles display a differential methylation profile in the coding region. Finally, fluorescence in situ hybridization analysis reveals an asynchronous pattern of replication at the Necdin locus. These results clearly demonstrate imprinting of the mouse Necdin gene. Mouse models will be powerful tools in the study of human PWS phenotype and imprinting mechanisms.

We have performed a comprehensive analysis of cell lines and tissues to compare and contrast the expression patterns of Flt3 ligand (FL), c-Kit ligand (KL), and macrophage colony-stimulating factor as well as their receptors, Flt3, c-Kit, and c-Fms. The message for FL is unusually ubiquitous, whereas that of its receptor is quite restricted, apparently limiting the function of the ligand to fetal development and early hematopoiesis. We have also sequenced a mouse FL genomic clone, revealing how the three splice variant FL mRNAs that we have isolated arise. The chromosomal location of the FL gene has been mapped, by in situ hybridization, to chromosome 7 in mouse and chromosome 19 in human. Natural FL protein has been purified from a stromal cell line and shown to be a 65 kD nondisulfide-linked homodimeric glycoprotein comprised of 30 kD subunits, each containing 12 kD of N- and O-linked sugars. Pulse-chase experiments show that one of the splice variants (T110) is responsible for producing the bulk of soluble FL, but only after it has first been expressed at the cell surface as a membrane-bound form. The other splice-variant forms produce molecules that are either obligatorily soluble (T169) or membrane-bound but released only very slowly (T118). Finally, even though most cell lines express some amount of FL mRNA, we found that very little FL protein is actually made, with T cells and stromal cells being the major producers. The data suggests that FL plays its roles over very short distances, perhaps requiring cell-cell contact.

While constructing a cDNA library of human embryos, we have isolated a clone homologous to jumonji, a mouse gene required for neural tube formation. We have determined the complete coding sequence of the human homologue (JMJ) and deduced the amino acid sequence of the putative protein. We show here that human and mouse jumonji putative proteins are homologous and present 90% identity. During human embryogenesis, JMJ mRNAs are predominantly expressed in neurons and particularly in dorsal root ganglion cells. They are also expressed in neurons of human adult cerebral cortex. In view of these observations, we propose JMJ as a candidate gene for developmental defects of the central nervous system in the human. The human JMJ gene maps at position 6p24-6p23.

International audience; Integrated hepatitis B virus (HBV) DNA is found in the great majority of human hepatocellular carcinomas, suggesting that these viral integrations may be implicated in liver oncogenesis. Besides the insertional mutagenesis characterized in a few selected cases and the contribution of viral transactivators to cell transformation to malignancy, HBV has been shown to generate gross chromosomal rearrangements potentially involved in carcinogenesis. Here, we report a t(3;8) chromosomal translocation present in a hepatocellular carcinoma developed in noncirrhotic liver tissue. One side of the translocation, in 8p23, is shown to be in the vicinity of the carboxypeptidase N gene, a locus that is heavily transcribed in liver tissue and frequently deleted in hepatocellular carcinomas and other epithelial tumors. The other side of the translocation, in 3q27-29, is widely implicated in several types of translocations occurring in different malignancies, such as large-cell lymphomas. The present data strongly support a model in which HBV-induced chromosomal rearrangements play a key role during multistep liver oncogenesis.

Two new members of the Ig superfamily, the Lutheran (Lu) blood group glycoprotein and the B-cell adhesion molecule (B-CAM) epithelial cancer antigen, have been recently cloned from human placenta and colon cancer HT29 cell line, respectively. Although amino acid sequences deduced from cDNA analysis suggested that B-CAM should represent an abridged form of the Lu glycoprotein lacking the last 40 amino acids of the putative cytoplasmic tail, the relationship between the genes encoding these polypeptides has not been determined. In the present report, we showed by Southern blot analysis that the Lu and B-CAM cDNAs derived from a unique LU gene which exhibited an HindIII RFLP associated with the Lua/Lub blood group polymorphism. Accordingly, in situ hybridization of the Lu cDNA probe confirmed the localization of the Lutheran blood group locus to chromosome 19 q13.2–13.3, as previously shown for a B-CAM DNA probe. Sequence comparison between cDNA and genomic PCR fragments indicated that the Lu and B-CAM transcripts previously isolated are generated through the alternative use of internal splice donor and acceptor sites within an exon located at the 3′ end of the LU gene. These spliceoforms corresponded to 2.5 kb and 4.0 kb mRNA species detectable by Northern blot in all tissues and cell lines in which the LU gene is expressed; their primary structures are consistent with the presence of both the Lu and B-CAM antigens on two glycoprotein isoforms. However, the 4.0 kb transcript was very poorly expressed as compared to the 2.5 kb species except in the colon carcinoma HT29 cell line, suggesting a differential regulation of the Lu/B-CAM messenger RNA in some tumor tissues.

We have previously shown that c-Krox is a zinc finger protein that can increases the transcriptional activity of the mouse α1(I) collagen promoter through its binding to two GC-rich sequences (Galera, P., Musso, M., Ducy, P., and Karsenty, G. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 9372-9376). In this report we show that c-Krox can bind to an additional site in the promoter of the α1(I) collagen gene and to three sites in the promoter of the α2(I) collagen gene, the other gene coding for type I collagen. One of the binding sites present in both promoters is adjacent to the CCAAT box. We have performed a structure/function analysis of c-Krox locating the transactivation domain in the zinc finger and C-terminal domains and the dimerization domain in the C-terminal end of the protein. We also demonstrate that c-Krox is an early response gene, whose expression is detectable as early as 9.5-day postcoitum in mouse embryos. Whole-mount in situ hybridization shows that c-Krox is expressed in dermatomes, the somite derivatives that generate dermis, and section in situ hybridization shows that c-Krox and α1(I) collagen mRNAs colocalized in skin but not in bone during development. This result is consistent with the predominant expression of c-Krox in skin in postnatal life. Thus, our findings suggest that c-Krox is one transcription factor controlling the coordinated expression of the two type I collagen genes in skin.