Promoter assay using transgenic mice reveals the presence of cis-acting elements governing tissue- and site-specific expression of Col11a2

1996 ◽  
Vol 15 (3) ◽  
pp. 194-195
Author(s):  
N. Tsumaki ◽  
M. Sugimoto ◽  
Y. Matsui ◽  
K. Nakata ◽  
T. Ochi ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Specific Expression ◽  
Promoter Assay ◽  
Site Specific ◽  
Cis Acting

scholarly journals Reconstitution of human Fc gamma RIII cell type specificity in transgenic mice.

1996 ◽  
Vol 183 (3) ◽  
pp. 1259-1263 ◽  
Author(s):  
M Li ◽  
U Wirthmueller ◽  
J V Ravetch
Keyword(s):  
Transgenic Mice ◽  
Nk Cells ◽  
Gene Promoter ◽  
Homologous Region ◽  
Cell Type ◽  
Cell Type Specificity ◽  
Specific Expression ◽  
Cis Acting ◽  
Sequence Elements ◽  
Flanking Sequences

The human low affinity receptors for the Fc domain of immunoglobulin G, Fc gamma RIII, are encoded by two genes (IIIA and IIIB) which share >95% sequence identity in both coding and flanking sequences. Despite this extraordinary sequence conservation, IIIA is expressed in natural killer (NK) cells and macrophages and is absent in neutrophils, whereas IIIB is expressed only in neutrophils. To determine the molecular basis for this differential expression, we have generated transgenic mice using the genomic sequences of IIIA and IIIB. IIIA and IIIB transgenic mice show faithful reconstitution of this human pattern of cell type specificity. To determine the cis acting sequence elements that confer this specificity, we constructed chimeric genes in which 5.8 kb of 5' sequences of the IIIB gene has been replaced with a homologous region from the IIIA gene, and conversely, IIIA 5' sequences have been substituted for the analogous region of the IIIB gene. Promoter swap transgenic mice that carry IIIA 5' flanking sequences express Fc gamma RIII in macrophages and NK cells. In contrast, promoter swap transgenic mice that contain IIIB 5' sequences express Fc gamma RIII in neutrophils only. These studies define the elements conferring the cell type-specific expression of the human Fc gamma RIII genes within the 5' flanking sequences and first intron of the human Fc gamma RIIIA and Fc gamma RIIIB genes.

scholarly journals Tissue-specific, developmental, hormonal, and dietary regulation of rat phosphoenolpyruvate carboxykinase-human growth hormone fusion genes in transgenic mice.

1992 ◽  
Vol 12 (3) ◽  
pp. 1007-1020 ◽  
Author(s):  
M K Short ◽  
D E Clouthier ◽  
I M Schaefer ◽  
R E Hammer ◽  
M A Magnuson ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Transgenic Mice ◽  
Human Growth Hormone ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Human Growth ◽  
Fusion Genes ◽  
Specific Expression ◽  
Tissue Specific ◽  
Cis Acting ◽  
Specific Manner

The cytosolic phosphoenolpyruvate carboxykinase (PEPCK) gene is expressed in multiple tissues and is regulated in a complex tissue-specific manner. To map the cis-acting DNA elements that direct this tissue-specific expression, we made transgenic mice containing truncated PEPCK-human growth hormone (hGH) fusion genes. The transgenes contained PEPCK promoter fragments with 5' endpoints at -2088, -888, -600, -402, and -207 bp, while the 3' endpoint was at +69 bp. Immunohistochemical analysis showed that the -2088 transgene was expressed in the correct cell types (hepatocytes, proximal tubular epithelium of the kidney, villar epithelium of the small intestine, epithelium of the colon, smooth muscle of the vagina and lungs, ductal epithelium of the sublingual gland, and white and brown adipocytes). Solution hybridization of hGH mRNA expressed from the transgenes indicated that white and brown fat-specific elements are located distally (-2088 to -888 bp) and that liver-, gut-, and kidney-specific elements are located proximally (-600 to +69 bp). However, elements outside of the region tested are necessary for the correct developmental pattern and level of PEPCK expression in kidney. Both the -2088 and -402 transgenes responded in a tissue-specific manner to dietary stimuli, and the -2088 transgene responded to glucocorticoid stimuli. Thus, different tissues utilize distinct cell-specific cis-acting elements to direct and regulate the PEPCK gene.

scholarly journals A 4.2 kb Upstream Region of the Human Corneodesmosin Gene Directs Site-Specific Expression in Hair Follicles and Hyperkeratotic Epidermis of Transgenic Mice

2004 ◽  
Vol 122 (3) ◽  
pp. 730-738 ◽  
Author(s):  
Hélène Gallinaro ◽  
Nathalie Jonca ◽  
Lutz Langbein ◽  
Christian Vincent ◽  
Michel Simon ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Hair Follicles ◽  
Upstream Region ◽  
Specific Expression ◽  
Site Specific

Regulation of Pax6 expression is conserved between mice and flies

Development ◽  
1999 ◽  
Vol 126 (2) ◽  
pp. 383-395 ◽  
Author(s):  
P.X. Xu ◽  
X. Zhang ◽  
S. Heaney ◽  
A. Yoon ◽  
A.M. Michelson ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
System Development ◽  
Regulatory Element ◽  
Amacrine Cells ◽  
Pax6 Expression ◽  
Specific Expression ◽  
Cis Acting ◽  
Evolutionarily Conserved ◽  
Visual System Development ◽  
Lens Placode

Pax6 plays a key role in visual system development throughout the metazoa and the function of Pax6 is evolutionarily conserved. However, the regulation of Pax6 expression during eye development is largely unknown. We have identified two physically distinct promoters in mouse Pax6, P0 and P1, that direct differential Pax6 expression in the developing eye. P0-initiated transcripts predominate in lens placode and corneal and conjunctival epithelia, whereas P1-initiated transcripts are expressed in lens placode, optic vesicle and CNS, and only weakly in corneal and conjunctival epithelia. To further investigate their tissue-specific expression, a series of constructs for each promoter were examined in transgenic mice. We identified three different regulatory regions which direct distinct domains of Pax6 expression in the eye. A regulatory element upstream of the Pax6 P0 promoter is required for expression in a subpopulation of retinal progenitors and in the developing pancreas, while a second regulatory element upstream of the Pax6 P1 promoter is sufficient to direct expression in a subset of post-mitotic, non-terminally differentiated photoreceptors. A third element in Pax6 intron 4, when combined with either the P0 or P1 promoter, accurately directs expression in amacrine cells, ciliary body and iris. These results indicate that the complex expression pattern of Pax6 is differentially regulated by two promoters acting in combination with multiple cis-acting elements. We have also tested whether the regulatory mechanisms that direct Pax6 ocular expression are conserved between mice and flies. Remarkably, when inserted upstream of either the mouse Pax6 P1 or P0 promoter, an eye-enhancer region of the Drosophila eyeless gene, a Pax6 homolog, directs eye- and CNS-specific expression in transgenic mice that accurately reproduces features of endogenous Pax6 expression. These results suggest that in addition to conservation of Pax6 function, the upstream regulation of Pax6 has also been conserved during evolution.

Identification of cis sequences controlling efficient position-independent tissue-specific expression of human major histocompatibility complex class I genes in transgenic mice

1991 ◽  
Vol 11 (7) ◽  
pp. 3564-3572 ◽  
Author(s):  
J W Chamberlain ◽  
H A Vasavada ◽  
S Ganguly ◽  
S M Weissman
Keyword(s):  
Major Histocompatibility Complex ◽  
Transgenic Mice ◽  
Major Histocompatibility Complex Class ◽  
Class I ◽  
Complex Class ◽  
Specific Expression ◽  
Tissue Specific ◽  
Cis Acting ◽  
Tissue Specific Expression ◽  
Histocompatibility Complex

We previously reported that genomic major histocompatibility complex class I human leukocyte antigen (HLA)-B7 gene constructs with as little as 0.66 kb of 5'- and 2.0 kb of 3'-flanking DNA were expressed efficiently and appropriately in transgenic mice. To identify and characterize the relevant cis-acting regulatory elements in more detail, we have generated and analyzed a series of transgenic mice carrying native HLA-B7 genes with further 5' truncations or intronic deletions and hybrid constructs linking the 5'-flanking region of B7 to a reporter gene. We were unable to detect a specific requirement for sequence information within introns 2 to 7 for either appropriate constitutive or inducible class I expression in adult animals. The results revealed the presence of cis-acting regulatory sequences between -0.075 kb and -0.66 kb involved in driving efficient copy number-dependent constitutive and gamma interferon-enhanced tissue-specific expression. The region from -0.11 to -0.66 kb is also sufficient to prevent integration site-specific "position effects," because in its absence HLA-B7 expression is frequently detected at significant levels at inappropriate sites. Conserved sequence elements homologous to the H-2 class I regulatory element, or enhancer A, and the interferon response sequence are located between about -151 and -228 bp of the B7 gene. Our results also indicate the existence of sequences downstream of -0.11 kb which can influence the pattern of tissue-specific expression of the HLA-B7 gene and the ability of this gene to respond to gamma interferon.

Tissue-specific and hormonally regulated expression of a rat alpha 2u globulin gene in transgenic mice

1987 ◽  
Vol 7 (10) ◽  
pp. 3749-3758
Author(s):  
V da C Soares ◽  
R M Gubits ◽  
P Feigelson ◽  
F Costantini
Keyword(s):  
Transgenic Mice ◽  
Gene Family ◽  
Hormonal Regulation ◽  
Germ Line ◽  
Regulatory Elements ◽  
Preputial Gland ◽  
Specific Expression ◽  
Tissue Specific ◽  
Cis Acting ◽  
Globulin Gene

To investigate the tissue-specific and hormonal regulation of the rat alpha 2u globulin gene family, we introduced one cloned member of the gene family into the mouse germ line and studied its expression in the resulting transgenic mice. Alpha 2u globulingene 207 was microinjected on a 7-kilobase DNA fragment, and four transgenic lines were analyzed. The transgene was expressed at very high levels, specifically in the liver and the preputial gland of adult male mice. The expression in male liver was first detected at puberty, and no expression was detected in female transgenic mice. This pattern of expression is similar to the expression of endogenous alpha 2u globulin genes in the rat but differs from the expression of the homologous mouse major urinary protein (MUP) gene family in that MUPs are synthesized in female liver and not in the male preputial gland. We conclude that these differences between rat alpha 2u globulin and mouse MUP gene expression are due to evolutionary differences in cis-acting regulatory elements. The expression of the alpha 2u globulin transgene in the liver was abolished by castration and fully restored after testosterone replacement. The expression could also be induced in the livers of female mice by treatment with either testosterone or dexamethasone, following ovariectomy and adrenalectomy. Therefore, the cis-acting elements responsible for regulation by these two hormones, as well as those responsible for tissue-specific expression, are closely linked to the alpha 2u globulin gene.

scholarly journals Separable cis-regulatory elements that contribute to tissue- and site-specific alpha 2(XI) collagen gene expression in the embryonic mouse cartilage.

1996 ◽  
Vol 134 (6) ◽  
pp. 1573-1582 ◽  
Author(s):  
N Tsumaki ◽  
T Kimura ◽  
Y Matsui ◽  
K Nakata ◽  
T Ochi
Keyword(s):  
Promoter Region ◽  
Regulatory Elements ◽  
Collagen Gene ◽  
Lacz Expression ◽  
Specific Expression ◽  
Site Specific ◽  
Cis Acting ◽  
First Intron ◽  
Vertebral Bodies ◽  
Type Xi Collagen

Type XI collagen is a structural component of the cartilage extracellular matrix and plays an important role in skeletal morphogenesis. As a step toward defining the molecular mechanisms responsible for the regulation of type XI collagen expression, we characterized the promoter region of the mouse alpha 2(XI) collagen gene (Coll1a2). We also generated transgenic mice harboring various fragments of the promoter and the first intron of Coll1a2 linked to the Escherichia coli beta-galactosidase gene to identify the cis-acting elements responsible for tissue- and site-specific expression during development. Cloning and sequence analysis of the 5' flanking region of Coll1a2 showed that the putative 3' end of the retinoid X receptor beta gene was located 742 bp upstream of the Coll1a2 start site. This suggested that the promoter region of Coll1a2 was localized within this 742-bp sequence, which contained multiple consensus regulatory elements. Examination of the transgenic mice revealed that the longest DNA construct (containing the entire promoter and first intron sequences) directed lacZ expression in the notochord as well as in the primordial cartilage throughout the body, with the pattern of expression mimicking that of endogenous Coll1a2 transcripts. On the other hand, deletion of the upstream approximately 290 bp resulted in the elimination of lacZ expression in the primordial cartilage of the carpals, tarsals, and vertebral bodies, whereas lacZ expression in the notochord and in the other primordial cartilage elsewhere was not affected. Deletion of the first intron sequence also resulted in the loss of lacZ expression in the primordial cartilage of the carpals, tarsals, and vertebral bodies, as well as in the notochord. These results demonstrate that the upstream 742-bp and first intron segments of the mouse Coll1a2 gene contain the necessary information to confer high level tissue-specific expression in mouse embryos. In addition, our observations suggest the presence of site-specific cis-acting elements that control Coll11a2 gene expression in different cartilaginous components of the skeleton.

scholarly journals Tissue-specific and hormonally regulated expression of a rat alpha 2u globulin gene in transgenic mice.

1987 ◽  
Vol 7 (10) ◽  
pp. 3749-3758 ◽  
Author(s):  
V da C Soares ◽  
R M Gubits ◽  
P Feigelson ◽  
F Costantini
Keyword(s):  
Transgenic Mice ◽  
Gene Family ◽  
Hormonal Regulation ◽  
Germ Line ◽  
Regulatory Elements ◽  
Preputial Gland ◽  
Specific Expression ◽  
Tissue Specific ◽  
Cis Acting ◽  
Globulin Gene

To investigate the tissue-specific and hormonal regulation of the rat alpha 2u globulin gene family, we introduced one cloned member of the gene family into the mouse germ line and studied its expression in the resulting transgenic mice. Alpha 2u globulingene 207 was microinjected on a 7-kilobase DNA fragment, and four transgenic lines were analyzed. The transgene was expressed at very high levels, specifically in the liver and the preputial gland of adult male mice. The expression in male liver was first detected at puberty, and no expression was detected in female transgenic mice. This pattern of expression is similar to the expression of endogenous alpha 2u globulin genes in the rat but differs from the expression of the homologous mouse major urinary protein (MUP) gene family in that MUPs are synthesized in female liver and not in the male preputial gland. We conclude that these differences between rat alpha 2u globulin and mouse MUP gene expression are due to evolutionary differences in cis-acting regulatory elements. The expression of the alpha 2u globulin transgene in the liver was abolished by castration and fully restored after testosterone replacement. The expression could also be induced in the livers of female mice by treatment with either testosterone or dexamethasone, following ovariectomy and adrenalectomy. Therefore, the cis-acting elements responsible for regulation by these two hormones, as well as those responsible for tissue-specific expression, are closely linked to the alpha 2u globulin gene.

Sequences 5′ of the homeobox of the Hox-1.4 gene direct tissue-specific expression of lacZ during mouse development

Development ◽  
1993 ◽  
Vol 117 (3) ◽  
pp. 823-833 ◽  
Author(s):  
R.R. Behringer ◽  
D.A. Crotty ◽  
V.M. Tennyson ◽  
R.L. Brinster ◽  
R.D. Palmiter ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Fusion Protein ◽  
Germ Cells ◽  
Spinal Ganglia ◽  
Control Element ◽  
Lacz Gene ◽  
Mouse Development ◽  
Specific Expression ◽  
Male Germ Cells ◽  
Cis Acting

The murine homeobox-containing gene Hox-1.4 is expressed in restricted patterns during embryogenesis and in male germ cells. To begin identification of the cis-acting elements regulating this expression, transgenic mice were generated carrying a chimeric construct that contained approx. 4 kb of 5′ flanking sequence and approx. 1 kb of structural gene, fused in frame to the E. coli lacZ gene. This construct directed expression of the resulting Hox-1.4, beta-galactosidase fusion protein in a pattern that reproduced virtually the complete embryonic and adult sites of expression of the endogenous gene. Embryonic expression of the fusion protein was first detected in mesoderm at day 8.0 of gestation (E 8.0). Between gestational ages E 8.5 to E 12.5, beta-gal expression was observed in the somites, the lateral walls of the posterior myelencephalon, the dorsal region and ventral wall of the spinal cord, spinal ganglia and prevertebrae and their surrounding mesenchyme, between presumptive ribs, as well as in mesenchymal layers in the lung, kidney and portions of the gut. Expression was also noted in the pancreas and in the supporting cells and sheath around subsets of peripheral nerves, sites that had not been detected previously. Adult expression was observed in testes, specifically in meiotic and post-meiotic male germ cells. In contrast, transgenic mice carrying 5′ deletions of the construct which leave approx. 1.2 kb or approx. 2.0 kb of Hox-1.4 sequence 5′ to the embryonic promoter, did not exhibit beta-gal staining. These deletion experiments defined at least one cis-acting control element necessary for the expression of the Hox-1.4 gene to a 2 kb region located 2 to 4 kb 5′ of the embryonic transcription start site.

scholarly journals Myeloid expression of the human c-fps/fes proto-oncogene in transgenic mice.

1990 ◽  
Vol 10 (6) ◽  
pp. 2521-2527 ◽  
Author(s):  
P Greer ◽  
V Maltby ◽  
J Rossant ◽  
A Bernstein ◽  
T Pawson
Keyword(s):  
Bone Marrow ◽  
Transgenic Mice ◽  
Tyrosine Kinase ◽  
Protein Tyrosine Kinase ◽  
Specific Expression ◽  
Tissue Specific ◽  
Protein Tyrosine ◽  
Cis Acting ◽  
Tissue Specific Expression ◽  
Pair Fragment

The mammalian c-fps/fes proto-oncogene encodes a 92-kilodalton cytoplasmic protein-tyrosine kinase (p92c-fes), which is expressed in immature and differentiated hematopoietic cells of the myeloid lineage. To determine the limits of the c-fps/fes locus and to investigate the cis-acting sequences required to direct appropriate tissue-specific expression, a 13-kilobase-pair fragment of human genomic DNA containing the entire c-fps/fes coding sequence was introduced into the mouse germ line. Transcription of the human c-fps/fes transgene was highest in bone marrow and showed a tissue distribution identical to that of the endogenous mouse gene. Macrophages cultured from transgenic mouse bone marrow contained particularly high levels of human and murine c-fps/fes RNA. Furthermore, expression of human c-fps/fes RNA induced a proportionate increase in the level of the p92c-fes protein-tyrosine kinase in bone marrow, bone marrow-derived macrophages, and spleen. Elevated levels of normal human p92c-fes had no obvious effect on mouse development or hematopoiesis. Remarkably, given the short 5'- and 3'-flanking sequences, expression of the human proto-oncogene in bone marrow was independent of integration site, was proportional to the transgene copy number, and was of comparable efficiency to that of the endogenous mouse c-fps/fes gene. The 13-kilobase-pair fragment therefore defines a genetic locus sufficient for the appropriate tissue-specific expression of the fps/fes protein-tyrosine kinase and includes a dominant cis-acting element that directs integration-independent myeloid expression in transgenic mice.

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