Enteropathogenic e.coli sustains iodoacetamide-induced ulcerative colitis-like colitis in rats: modulation of IL-1ß, IL-6, TNF-α, COX-2, and apoptosisi.

Pathogenic or non-pathogenic bacteria from flora may play a key role in inflammatory bowel disease (IBD) pathogenesis. However, a specific infectious agent causing IBD has not been identified. This study assessed the impact of enteropathogenic E. coli (EPEC) on the modulation of IL-1beta, IL-6, TNF- alpha, COX-2, BAX and Bcl-2 expression, in sustaining inflammation of a rat colitis model. Two hundred male Sprague-Dawley rats (4 groups) were inoculated weekly or bi-weekly for 70 days, with 1 percent methylcellulose (MC), (b) 6 percent iodoacetamide (IA) in 1 percent MC, (c) 4x108 CFU of EPEC, and (d) IA+EPEC. After a month, treatment was stopped in half of the animals in each group. IL-1beta, IL-6, TNF-alpha, COX-2, BAX and Bcl-2 expression were measured in colonic mucosa scrapings. IL-1beta, IL-6, TNF-alpha, and COX-2 were significantly increased in colonic mucosa of the IA+EPEC group and to a lesser but significant level in the IA group compared to controls, or EPEC alone, both in continued and discontinued treatment groups. Additionally, the BAX/Bcl-2 ratio decreased, indicating less apoptosis in the IA+EPEC group which exhibited more necrosis. These effects increased with experiment duration. This work provides new arguments favouring the role of bacteria in IBD pathogenesis.

Expression and localisation of insulin receptor substrate 2 in normal intestine and colorectal tumours. Regulation by intestine-specific transcription factor CDX2.

BACKGROUND AND AIMS: Self-renewal and differentiation of intestinal epithelium is a tightly regulated process, whose perturbations are implicated in human colorectal tumourigenesis. The insulin/insulin-like growth factor (IGF) signalling pathway may play an important role in intestinal epithelium homeostasis. Insulin receptor substrate 2 (IRS2) is a poorly characterised component in this pathway. METHODS: Using complementary in vitro and in vivo human and murine models, expression (mRNA and protein levels), localisation (immunohistochemistry) and regulation of IRS2 were investigated in the normal intestine and colorectal tumours. In silico analysis of the human IRS2 promoter was performed together with reporter and chromatin immunoprecipitation assays. RESULTS: Significant IRS2 expression was detected in the intestine, with specific protein localisation in the villus region of the ileum and in the surface epithelium of the colon. In human HT29 and Caco2 cells, IRS2 mRNA levels increased with spontaneous and induced differentiation, together with CDX2 (caudal-related homeobox protein 2), P21 and KLF4 (Krüppel-like factor 4). Adenoviral infection with human CDX2 induced IRS2 expression in APC- (adenomatous polyposis coli) and beta-catenin-mutated cells. On the other hand, IRS2 downregulation was observed in differentiated enterocytes after adenoviral infection with short hairpin CDX2 (shCDX2), in the intestine of CDX2 heterozygous mice and in colorectal tumours of Apc(Min/+) and patients with familial adenomatous polyposis (FAP). The human IRS2 promoter region presents several CDX2-binding sites where CDX2 immunoprecipitated in vivo. IRS2 reporters were functionally activated via CDX2 and blocked via a dominant-negative CDX2 protein. CONCLUSIONS: Combining gain- and loss-of-function approaches, an intriguing scenario is presented whereby IRS2 is significantly expressed in the apical intestinal compartment and is directly controlled by CDX2 in normal intestine and tumours.

Multiple levels of control of the stage- and region-specific expression of rat intestinal lactase.

To elucidate the mechanisms leading to the functional regionalization of the digestive epithelium, lactase expression was analyzed at the protein, mRNA and gene levels, along the intestinal tract at various stages of the rat postnatal development. In the colon of neonates, the transient expression of mRNA and enzyme correlated well with gene transcription. In contrast to the colon, complex patterns were observed in the small intestine. In suckling animals, the mRNA was present at a high level despite the progressive decline of enzyme activity. Crypts were devoid of mRNA and the transcript mainly accumulated in the lower half of the villi. From weaning onwards, a functional regionalization of the epithelium was defined, characterized by the modification of the longitudinal distribution of lactase mRNA. Indeed the transcript remained abundant in the distal duodenum, jejunum and proximal ileum, but decreased in the proximal duodenum and became virtually absent in the distal ileum. Concomitantly, the mRNA and enzyme distribution along the villi changed in the different segments of the small intestine. Patterns similar to those described in sucklings were retained in the adult jejunum. In contrast, mRNA and enzyme could no longer be detected in the distal ileum, while mosaicism appeared in the proximal duodenum. In vitro transcription assays carried out with isolated nuclei suggested that the decay of lactase mRNA in the proximal duodenum at weaning was associated with a decreasing rate of transcription of the gene. However active gene transcription was retained in the nuclei of the adult jejunum and ileum. The loss of mRNA in the adult distal ileum despite the maintenance of active transcription did not result from an intragenic block of pre-RNA elongation, as shown by transcription assays carried out at various positions of the lactase gene. In addition, we looked for the ontogenic decline of lactase protein despite the maintenance of a high amount of mRNA in the jejunum, and it became evident that the fraction of mRNA present in polysomes was constant with age. Taken together, these data indicate that lactase constitutes an unusual marker of development and of functional regionalization of the intestinal tract which exhibits a complex time- and space-specific pattern of gene, mRNA, and protein expression. The distinct patterns occurring in the duodenum, jejunum, ileum, and the colon of pre- and postweaned rats depend on a combination of transcriptional, posttranscriptional, and posttranslational levels of regulation. and are associated with a different mRNA distribution along villi in each intestinal segment.

Fetal endoderm primarily holds the temporal and positional information required for mammalian intestinal development.

In rodents, the intestinal tract progressively acquires a functional regionalization during postnatal development. Using lactase-phlorizin hydrolase as a marker, we have analyzed in a xenograft model the ontogenic potencies of fetal rat intestinal segments taken prior to endoderm cytodifferentiation. Segments from the presumptive proximal jejunum and distal ileum grafted in nude mice developed correct spatial and temporal patterns of lactase protein and mRNA expression, which reproduced the normal pre- and post-weaning conditions. Segments from the fetal colon showed a faint lactase immunostaining 8-10 d after transplantation in chick embryos but not in mice; it is consistent with the transient expression of this enzyme in the colon of rat neonates. Heterotopic cross-associations comprising endoderm and mesenchyme from the presumptive proximal jejunum and distal ileum developed as xenografts in nude mice, and they exhibited lactase mRNA and protein expression patterns that were typical of the origin of the endodermal moiety. Endoderm from the distal ileum also expressed a normal lactase pattern when it was associated to fetal skin fibroblasts, while the fibroblasts differentiated into muscle layers containing alpha-smooth-muscle actin. Noteworthy, associations comprising colon endoderm and small intestinal mesenchyme showed a typical small intestinal morphology and expressed the digestive enzyme sucrase-isomaltase normally absent in the colon. However, in heterologous associations comprising lung or stomach endoderm and small intestinal mesenchyme, the epithelial compartment expressed markers in accordance to their tissue of origin but neither intestinal lactase nor sucrase-isomaltase. A thick intestinal muscle coat in which cells expressed alpha-smooth-muscle actin surrounded the grafts. The results demonstrate that: (a) the temporal and positional information needed for intestinal ontogeny up to the post-weaning stage results from an intrinsic program that is fixed in mammalian fetuses prior to endoderm cytodifferentiation; (b) this temporal and positional information is primarily carried by the endodermal moiety which is also able to change the fate of heterologous mesodermal cells to form intestinal mesenchyme; and (c) the small intestinal mesenchyme in turn may deliver instructive information as shown in association with colonic endoderm; yet this effect is not obvious with nonintestinal endoderms.

Type IV collagen mRNA accumulates in the mesenchymal compartment at early stages of murine developing intestine.

The expression of type IV collagen mRNA during mouse intestinal morphogenesis was examined by in situ hybridization using a cDNA probe corresponding to mRNA for alpha 1 (IV) chain. Type IV collagen mRNA is detected in the embryonic mesenchymal cells at early stages of development (12 d of gestation). A segregation of mesenchymal cells expressing high levels of type IV collagen mRNA in close vicinity of the epithelium occurs just before villus formation. During villus outgrowth, type IV collagen mRNA, still confined to mesenchyme-derived tissues, is progressively restricted to the mucosal connective tissue (the lamina propria) and to a lesser extent to the muscular layers. In the adult, the amount of messenger is quite low as compared to the level found in the developing intestine and the in situ hybridization signal, indistinguishable from the background, is uniform throughout the whole intestinal wall. At all developmental stages no detectable specific hybridization signal is virtually observed over the epithelium cell layer. These results show that high amounts of the type IV collagen messenger are detected during phases of intensive morphogenetic events. Furthermore, they reinforce the notion already gained previously (Simon-Assmann et al. 1988) that the mesenchymal compartment is the principal endogenous source of type IV collagen. They also indicate that the continuous migration of epithelial cells along the basement membrane of intestinal villi in the mature organ is not accompanied by a significant remodeling of the collagen IV network.

Key role of the Cdx2 homeobox gene in extracellular matrix-mediated intestinal cell differentiation.

To explore the role of homeobox genes in the intestine, the human colon adenocarcinoma cell line Caco2-TC7 has been stably transfected with plasmids synthesizing Cdx1 and Cdx2 sense and antisense RNAs. Cdx1 overexpression or inhibition by antisense RNA does not markedly modify the cell differentiation markers analyzed in this study. In contrast, Cdx2 overexpression stimulates two typical markers of enterocytic differentiation: sucrase-isomaltase and lactase. Cells in which the endogenous expression of Cdx2 is reduced by antisense RNA attach poorly to the substratum. Conversely, Cdx2 overexpression modifies the expression of molecules involved in cell-cell and cell-substratum interactions and in transduction process: indeed, E-cadherin, integrin-beta4 subunit, laminin-gamma2 chain, hemidesmosomal protein, APC, and alpha-actinin are upregulated. Interestingly, most of these molecules are preferentially expressed in vivo in the differentiated villi enterocytes rather than in crypt cells. Cdx2 overexpression also results in the stimulation of HoxA-9 mRNA expression, an homeobox gene selectively expressed in the colon. In contrast, Cdx2-overexpressing cells display a decline of Cdx1 mRNA, which is mostly found in vivo in crypt cells. When implanted in nude mice, Cdx2-overexpressing cells produce larger tumors than control cells, and form glandular and villus-like structures. Laminin-1 is known to stimulate intestinal cell differentiation in vitro. In the present study, we demonstrate that the differentiating effect of laminin-1 coatings on Caco2-TC7 cells is accompanied by an upregulation of Cdx2. To further document this observation, we analyzed a series of Caco2 clones in which the production of laminin-alpha1 chain is differentially inhibited by antisense RNA. We found a positive correlation between the level of Cdx2 expression, that of endogenous laminin-alpha1 chain mRNA and that of sucrase-isomaltase expression in these cell lines. Taken together, these results suggest (a) that Cdx1 and Cdx2 homeobox genes play distinct roles in the intestinal epithelium, (b) that Cdx2 provokes pleiotropic effects triggering cells towards the phenotype of differentiated villus enterocytes, and (c) that Cdx2 expression is modulated by basement membrane components. Hence, we conclude that Cdx2 plays a key role in the extracellular matrix-mediated intestinal cell differentiation.

Sprouty2 inhibits BDNF-induced signaling and modulates neuronal differentiation and survival.

Sprouty (Spry) proteins are ligand-inducible inhibitors of receptor tyrosine kinases-dependent signaling pathways, which control various biological processes, including proliferation, differentiation and survival. Here, we investigated the regulation and the role of Spry2 in cells of the central nervous system (CNS). In primary cultures of immature neurons, the neurotrophic factor BDNF (brain-derived neurotrophic factor) regulates spry2 expression. We identified the transcription factors CREB and SP1 as important regulators of the BDNF activation of the spry2 promoter. In immature neurons, we show that overexpression of wild-type Spry2 blocks neurite formation and neurofilament light chain expression, whereas inhibition of Spry2 by a dominant-negative mutant or small interfering RNA favors sprouting of multiple neurites. In mature neurons that exhibit an extensive neurite network, spry2 expression is sustained by BDNF and is downregulated during neuronal apoptosis. Interestingly, in these differentiated neurons, overexpression of Spry2 induces neuronal cell death, whereas its inhibition favors neuronal survival. Together, our results imply that Spry2 is involved in the development of the CNS by inhibiting both neuronal differentiation and survival through a negative-feedback loop that downregulates neurotrophic factors-driven signaling pathways.

Different effects of the Cdx1 and Cdx2 homeobox genes in a murine model of intestinal inflammation.

AIMS: The CDX1 and CDX2 homeoproteins are intestine-specific transcription factors regulating homeostasis. We investigated their relevance in experimentally-induced intestinal inflammation. METHODS: The response to intestinal inflammation induced by dextran sodium sulfate (DSS) was compared in wild type, Cdx1(-/-) and Cdx2(+/-) mice. Intestinal permeability was determined in wild type and Cdx2(+/-) mice. Protein-protein interactions were investigated by co-immunoprecipitation and GST-pulldown, and their functional consequences were assessed using Luciferase reporter systems. RESULTS: Heterozygous Cdx2(+/-) mice, but not Cdx1(-/-) mice, were hypersensitive to DSS-induced acute inflammation as all these mice showed blood in the stools at day 1 of DSS treatment. Hypersensitivity was associated to a 50% higher intestinal permeability. In Cdx2(+/-) mice, the colonic epithelium was repaired during the week after the end of DSS treatment, whereas two weeks were required for wild type animals. Subsequently, no colonic tumour was observed in Cdx2(+/-) mice subjected to 5 repeated cycles of DSS, in contrast to the 2.7 tumours found per wild type mouse. Based on the fact that Smad3(+/-) mice, like Cdx2(+/-) mice, better repair the damaged intestinal epithelium, we found that the CDX2 protein interacts with SMAD3, independently of SMAD4, resulting in a 5-fold stimulation of SMAD3 transcriptional activity. CDX1 also interacted with SMAD3 but it inhibited by 10-fold the SMAD3/SMAD4-dependent transcription. CONCLUSION: The Cdx1 and Cdx2 homeobox genes have distinct effects on the outcome of a pro-inflammatory challenge. This is mirrored by different functional interactions of the CDX1 and CDX2 proteins with SMAD3, a major element of the TGFbeta signalling pathway.

Functional interference between thyroid hormone receptor alpha (TRalpha) and natural truncated TRDeltaalpha isoforms in the control of intestine development.

Thyroid hormone is known to participate in the control of intestine maturation at weaning. Its action is mediated by the thyroid hormone nuclear receptors, encoded by the TRalpha and TRbeta genes. Since previous studies have shown that TRbeta plays a minor role in the gut, we focused here our analysis on the TRalpha gene. The TRalpha locus generates the TRalpha1 receptor together with the splicing variant TRalpha2 and the truncated products TRDeltaalpha1 and TRDeltaalpha2, which all lack an intact ligand binding domain. The TRDeltaalpha isoforms are transcribed from an internal promoter located in intron 7, and their distribution is restricted to a few tissues including those of the intestine. In order to define the functions of the different isoforms encoded by the TRalpha locus in the intestinal mucosa, we produced mice either lacking all known TRalpha products or harboring a mutation which inactivates the intronic promoter. We performed a detailed analysis of the intestinal phenotypes in these mice and compared it to that of the previously described TRalpha(-/-) mice, in which TRalpha isoforms are abolished but the TRDeltaalpha isoforms remain. This comparative analysis leads us to the following conclusions: (i) the TRalpha1 receptor mediates the T3-dependent functions in the intestine at weaning time and (ii) the TRDeltaalpha products negatively control the responsiveness of the epithelial cells to T3. Moreover, we show that TRDeltaalpha proteins can interfere with the transcription of the intestine-specific homeobox genes cdx1 and cdx2 and that their activity is regulated by TRalpha1. Altogether these data demonstrate that cooperation of TRalpha and TRDeltaalpha products is essential to ensure the normal postnatal development of the intestine and that mutations in the TRalpha locus can generate different phenotypes caused by the disruption of the equilibrium between these products.

Production of low-lactose milk by ectopic expression of intestinal lactase in the mouse mammary gland.

We have investigated, in mice, an in vivo method for producing low-lactose milk, based on the creation of transgenic animals carrying a hybrid gene in which the intestinal lactase-phlorizin hydrolase cDNA was placed under the control of the mammary-specific alpha-lactalbumin promoter. Transgenic females expressed lactase protein and activity during lactation at the apical side of mammary alveolar cells. Active lactase was also secreted into milk, anchored in the outer membrane of fat globules. Lactase synthesis in the mammary gland caused a significant decrease in milk lactose (50-85%) without obvious changes in fat and protein concentrations. Sucklings nourished with low-lactose milk developed normally. Hence, these data validate the use of transgenic animals expressing lactase in the mammary gland to produce low-lactose milk in vivo, and they demonstrate that the secretion of an intestinal digestive enzyme into milk can selectively modify its composition.

Downregulation of the colon tumour-suppressor homeobox gene Cdx-2 by oncogenic ras.

Downregulation of the colon tumour-suppressor homeobox gene Cdx-2 by oncogenic ras Constitutive activation of the ras proto-oncogene is a frequent and early event in colon cancers, but the downstream nuclear targets are not fully understood. The Cdx-1 and Cdx-2 homeobox genes play crucial roles in intestinal cell proliferation and differentiation. In addition, Cdx-2 is a colonic tumour-suppressor gene, whereas Cdx-1 has oncogenic potential. Here, we show that constitutive activation of ras alters Cdx-1 and Cdx-2 expression in human colonic Caco-2 and HT-29 cells that harbour a normal ras proto-oncogene. Oncogenic ras downregulates Cdx-2 through activation of the PKC pathway and a decline in activity of the Cdx-2 promoter AP-1 site. This decline results from a PKC-dependent decrease in the relative expression of c-Jun, an activator of Cdx-2 transcription, compared to c-Fos, an inhibitor of Cdx-2. Unlike Cdx-2, Cdx-1 is upregulated by oncogenic ras and this effect is mediated by activation of the MEK1 pathway. These results indicate that oncogenic ras activation has opposite effects on Cdx-1 and Cdx-2 expression through distinct signalling pathways and they provide the first evidence for a functional link between ras activation and the downregulation of the Cdx-2 tumour-suppressor gene in colon cancer cells.

The rudimentary gene of Drosophila melanogaster encodes four enzymic functions.

We have determined the 7168 nucleotide DNA sequence corresponding to the messenger RNA of the rudimentary gene of Drosophila melanogaster. By sequence comparison with genes involved in the pyrimidine pathway of prokaryotes and lower eukaryotes, we conclude that the rudimentary gene encodes four enzymically different functions. Each function is restricted to a specific coding domain but in an order different from that previously defined by genetic data. We have found that the corresponding mammalian gene, the CAD gene, exhibits a similar functional organization, and we propose schemes for the evolution of the corresponding coding sequences.

Molecular organization of the rudimentary gene of Drosophila melanogaster.

We have determined the molecular structure of the rudimentary gene of Drosophila melanogaster. The transcription unit, which extends over 14,000 bases, is split into seven exons and six introns. Sequences presenting homologies with a TATA box and a CAT box are located upstream from the transcription initiation site, and there is a consensus polyadenylation signal in the vicinity of the 3' end of the messenger RNA coding sequence. Short intronic sequences fit well with consensus signals found in other eukaryotic genes and involved in the splicing of the precursor messenger RNA. Partial genomic and complementary DNA sequencing has revealed stretches of amino acid sequence homologies with the corresponding enzymes in Saccharomyces cerevisiae and Escherichia coli. This is in good agreement with the genetic map of the locus, which indicates that the enzymic activities encoded by the gene are organized linearly along the DNA.

Broader expression of the mouse platelet factor 4-cre transgene beyond the megakaryocyte lineage.

BACKGROUND: Transgenic mice expressing cre recombinase under the control of the platelet factor 4 (Pf4) promoter, in the context of a 100-kb bacterial artificial chromosome, have become a valuable tool with which to study genetic modifications in the platelet lineage. However, the specificity of cre expression has recently been questioned, and the time of its onset during megakaryopoiesis remains unknown. OBJECTIVES/METHODS: To characterize the expression of this transgene, we used double-fluorescent cre reporter mice. RESULTS: In the bone marrow, Pf4-cre-mediated recombination had occurred in all CD42-positive megakaryocytes as early as stage I of maturation, and in rare CD42-negative cells. In circulating blood, all platelets had recombined, along with only a minor fraction of CD45-positive cells. However, we found that all tissues contained recombined cells of monocyte/macrophage origin. When recombined, these cells might potentially modify the function of the tissues under particular conditions, especially inflammatory conditions, which further increase recombination in immune cells. Unexpectedly, a subset of epithelial cells from the distal colon showed signs of recombination resulting from endogenous Pf4-cre expression. This is probably the basis of the unexplained colon tumors developed by Apc(flox/flox) ;Pf4-cre mice, generated in a separate study on the role of Apc in platelet formation. CONCLUSION: Altogether, our results indicate early recombination with full penetrance in megakaryopoiesis, and confirm the value of Pf4-cre mice for the genetic engineering of megakaryocytes and platelets. However, care must be taken when investigating the role of platelets in processes outside hemostasis, especially when immune cells might be involved.

Sequence of the precursor of intestinal lactase-phlorizin hydrolase from fetal rat.

The nucleotide sequence of the cDNA corresponding to the precursor of fetal rat intestinal lactase-phlorizin hydrolase was determined. The precursor consists of four tandemly organized homologous domains flanked by a signal peptide in the N-terminal region and by a transmembrane peptide in the C-terminal region.

The cis-element CE-LPH1 of the rat intestinal lactase gene promoter interacts in vitro with several nuclear factors present in endodermal tissues.

We have shown by electrophoretic mobility shift assays that the nucleotide sequence CE-LPH1, centred at position -49 with respect to the transcription start site of the rat gene encoding intestinal lactase-phlorizin hydrolase, interacts in vitro with nuclear proteins present in the jejunum of suckling animals. Proteins binding to this element were also found in organs of endodermal origin that do not (or no longer) express lactase-phlorizin hydrolase, i.e. the colon, lung and the liver, but not in the brain. However, a DNA-protein interaction was hardly detected with nuclear extracts prepared from adult tissues, although typical factors binding to the Sp1 binding site were detected at the adult stage as in the sucklings. Southwestern blotting experiments conducted with nuclear extracts prepared from the tissues of suckling rats indicated that CE-LPH1 interacts with several factors in the jejunum, colon, lung and the liver. Some of these DNA-binding proteins are specifically expressed in the jejunum or in the liver, whereas others seem to be shared with the colon and the lung. Hence, the cis-element CE-LPH1 located in close vicinity to the pseudo-TATA-box of the intestinal lactase-phlorizin hydrolase gene promoter interacts in vitro with a family of nuclear proteins which may represent markers of the endodermal lineage predominantly expressed prior to weaning.

Discrepancy between the intestinal lactase enzymatic activity and mRNA accumulation in sucklings and adults. Effect of starvation and thyroxine treatment.

The accumulation profile of intestinal lactase mRNA was investigated in suckling and adult rats and pigs. We found no correlation between the lactase enzymatic activity and the accumulation of the messenger at both developmental stages. Modulation of lactase activity by starvation or thyroxine treatment had no effect on lactase mRNA accumulation in the rat intestine. These results confirm that thyroxine modulates lactase expression essentially at the post-transcriptional level.

Gradient expression of Cdx along the rat intestine throughout postnatal development.

Rat genomic DNA was isolated by homology with Cdx1, a murine homeogene selectively expressed in intestinal cells of endodermal origin. Southern blot analysis indicated that the rat genome contains a single or a small number of closely related Cdx gene(s). A major 1.7 kb Cdx mRNA was detected in neonate, suckling and adult rats whereas a 6.5 kb mRNA was restricted to sucklings and adults. Both transcripts showed decreasing concentration from the colon towards the proximal part of the small intestine. No obvious correlation could be established with the patterns of expression of transcripts corresponding to markers of cell proliferation and cell differentiation during postnatal development.

Intestinal epithelial-mesenchymal cell interactions.

Intestinal morphogenesis, as well as maintenance of the stem cell population and of the steady state between cell proliferation and differentiation, results from controlled cell interactions. There is growing evidence that the mesenchymal cells control epithelial cell behavior via their own expression and induction in the epithelial cells of key regulatory genes. This heterologous cross talk involves basement membrane molecules and paracrine factors. New in vitro/in vivo cellular models allowed us to analyze various mesenchymal cell phenotypes and to show that they exhibit different inductive properties on epithelial cells and that their proliferation and metabolic properties are differentially modulated by cytokines. Finally the epithelial-mesenchymal unit is controlled by hormonal and exogenous factors.

The rat LPH gene 5' region: comparative structure with the human gene.

We have determined the sequence of a 2784 bp rat genomic fragment originating from the 5' region of the gene coding for intestinal lactase-phlorizin hydrolase. The fragment overlaps the gene exon 1, part of the intron 1 and the 5'-upstream segment including a TATA-like box. Over 155 bp, the upstream segment shows 72% similarity with the corresponding sequence in human. Far upstream, the rat sequence exhibits a Calcium Responsive Element and putative binding sites for AP2, C/EBP, and CTF/NF. The intron contains a T-rich sequence that may cause DNA helix distortion.