Mouse testis Pdha-2 promoter upstream sequences confer tissue-and temporal-specific activity in transgenic mice

1994 ◽  
Vol 6 (5) ◽  
pp. 599 ◽  
Author(s):  
RC Iannello ◽  
JC Young ◽  
S Sumarsono ◽  
MJ Tymms ◽  
I Kola
Keyword(s):  
Transgenic Mice ◽  
Specific Activity ◽  
Pyruvate Dehydrogenase Complex ◽  
Somatic Tissue ◽  
Specific Expression ◽  
Complex Process ◽  
Specific Manner ◽  
Cat Gene ◽  
Testis Specific Expression

Spermatogenesis is a complex process requiring the coordinate expression of a number of testis-specific genes. One of these, Pdha-2, codes for the murine testis-specific isoform of the E1 alpha subunit of the pyruvate dehydrogenase complex. To elucidate the mechanisms regulating its expression in vivo, we have begun to investigate the Pdha-2 promoter in transgenic mice. In this paper, a construct containing 3.0 kb of promoter and upstream sequences is reported to be sufficient for directing the testis-specific expression of a CAT reporter gene in mice harbouring the transgene. Similarly to the endogenous Pdha-2, the CAT gene is expressed in testis in a stage-specific manner. However, the 3.0-kb Pdha-2 promoter is not active in somatic tissue suggesting that repressor elements may be present within these sequences.

scholarly journals The CD11b promoter directs high-level expression of reporter genes in macrophages in transgenic mice [published erratum appears in Blood 1995 Apr 1;85(7):1983]

Blood ◽  
1995 ◽  
Vol 85 (2) ◽  
pp. 319-329 ◽  
Author(s):  
S Dziennis ◽  
RA Van Etten ◽  
HL Pahl ◽  
DL Morris ◽  
TL Rothstein ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transgenic Mice ◽  
Transgene Expression ◽  
Heterologous Gene Expression ◽  
Myeloid Cells ◽  
Reporter Genes ◽  
Regulatory Elements ◽  
Specific Expression ◽  
High Level

Abstract CD11b is the alpha chain of the Mac-1 integrin and is preferentially expressed in myeloid cells (neutrophils, monocytes, and macrophages). We have previously shown that the CD11b promoter directs cell-type- specific expression in myeloid lines using transient transfection assays. To confirm that these promoter sequences contain the proper regulatory elements for correct myeloid expression of CD11b in vivo, we have used the -1.7-kb human CD11b promoter to direct reporter gene expression in transgenic mice. Stable founder lines were generated with two different reporter genes, a Thy 1.1 surface marker and the Escherichia coli lacZ (beta-galactosidase) gene. Analysis of founders generated with each reporter demonstrated that the CD11b promoter was capable of driving high levels of transgene expression in murine macrophages for the lifetime of the animals. Similar to the endogenous gene, transgene expression was preferentially found in mature monocytes, macrophages, and neutrophils and not in myeloid precursors. These experiments indicate that the -1.7 CD11b promoter contains the regulatory elements sufficient for high-level macrophage expression. This promoter should be useful for targeting heterologous gene expression to mature myeloid cells.

In vitro and in vivo analysis of murine lipoprotein lipase gene promoter: tissue-specific expression

1995 ◽  
Vol 268 (2) ◽  
pp. E213-E218 ◽  
Author(s):  
J. M. Gimble ◽  
X. Hua ◽  
F. Wanker ◽  
C. Morgan ◽  
C. Robinson ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Lipoprotein Lipase ◽  
Reporter Gene ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Specific Expression ◽  
Tissue Specific ◽  
Brown Adipose ◽  
Octamer Motif

Lipoprotein lipase, an enzyme of central importance to lipid metabolism, is most abundant in adipose tissues, cardiac and skeletal muscle, and portions of the brain. The current work examined the murine lipoprotein lipase promoter using transient transfection, gel-retention analyses, and transgenic mice. Maximum expression of the luciferase reporter gene in transfected cells was observed with -101 bp of the promoter. Nuclear extracts from tissues expressing lipoprotein lipase contained DNA binding proteins that recognize the CCAAT box (-64 bp) and an octamer motif (-46 bp); this combination of factors was absent in nonexpressing tissues. Transgenic mice from three of five founders prepared with -1,824-bp promoter constructs expressed the luciferase reporter gene at highest levels in brown adipose tissue and brain. These findings suggest that the -1,824-bp promoter region contains sequence elements responsible for the tissue-specific transcription of lipoprotein lipase in vivo.

Identification of DNA elements cooperatively activating proopiomelanocortin gene expression in the pituitary glands of transgenic mice

1992 ◽  
Vol 12 (9) ◽  
pp. 3978-3990
Author(s):  
B Liu ◽  
G D Hammer ◽  
M Rubinstein ◽  
M Mortrud ◽  
M J Low
Keyword(s):  
Gene Expression ◽  
Transgenic Mice ◽  
Reporter Gene ◽  
Transgene Expression ◽  
Intermediate Lobe ◽  
Mobility Shift ◽  
Specific Expression ◽  
Dna Elements ◽  
Pomc Gene

The proopiomelanocortin (POMC) gene is highly expressed in adult mouse pituitary anterior lobe corticotrophs and intermediate lobe melanotrophs. To identify the DNA elements important for this tissue-specific expression, we analyzed a series of POMC reporter genes in transgenic mice. A DNA fragment containing rat POMC 5'-flanking sequences from -323 to -34 recapitulated both basal pituitary cell-specific and hormonally stimulated expression in adult mice when fused to a heterologous thymidine kinase promoter. Developmental onset of the reporter gene expression lagged by 1 day but otherwise closely paralleled the normal ontogeny of murine POMC gene expression, including corticotroph activation at embryonic day 14.5 (E14.5) followed by melanotroph activation at E15.5 to E16.5. AtT20 corticotroph nuclear protein extracts interacted with three specific regions of the functional POMC promoter in DNase I protection assays. The positions of these protected sites were -107 to -160 (site 1), -182 to -218 (site 2), and -249 to -281 (site 3). Individual deletions of these footprinted sites did not alter transgene expression; however, the simultaneous deletion of sites 2 and 3 prevented transgene expression in both corticotrophs and melanotrophs. Electrophoretic mobility shift and Southwestern (DNA-protein) assays demonstrated that multiple AtT20 nuclear proteins bound to these footprinted sites. We conclude that the sequences between -323 and -34 of the rat POMC gene promoter are both necessary and sufficient for correct spatial, temporal, and hormonally regulated expression in the pituitary gland. Our data suggest that the three footprinted sites within the promoter are functionally interchangeable and act in combination with promoter elements between -114 and -34. The inability of any reporter gene construction to dissociate basal and hormonally stimulated expression suggests that these DNA elements are involved in both of these two characteristics of POMC gene expression in vivo.

Beta-MHC and SMLC1 transgene induction in overloaded skeletal muscle of transgenic mice

1996 ◽  
Vol 270 (4) ◽  
pp. C1111-C1121 ◽  
Author(s):  
J. L. Wiedenman ◽  
I. Rivera-Rivera ◽  
D. Vyas ◽  
G. Tsika ◽  
L. Gao ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Troponin C ◽  
Type I ◽  
Initial Attempt ◽  
Specific Expression ◽  
Slow Type ◽  
Transgenic Lines ◽  
Fast Twitch Muscle ◽  
Mechanical Overload

The hypertrophic responses of white fast-twitch muscle to mechanical overload has been investigated using transgenic mice. After 7 wk of overload, endogenous beta-myosin heavy chain (MHC) and slow myosin light chain 1 and 2 (SMLC1, SMLC2) protein were increased in the overloaded plantaris (OP) muscle compared with sham-operated control plantaris (CP)muscle. Concurrently, the levels of endogenous beta-MHC, SMLC1, SMLC2, and cardiac/slow troponin C (CTnC) mRNA transcripts were significantly increased in OP muscles, whereas skeletal troponin C (sTnC) mRNA transcript levels decreased. As an initial attempt to locate DNA sequence(s) that governs beta-MHC induction in response to mechanical overload, multiple independent transgenic lines harboring four different human beta-MHC transgenes (beta 1286, beta 988, beta 450, beta 141) were generated. Except for transgene beta 141, muscle-specific expression and induction (3- to 22-fold) in OP muscles were observed by measuring chloramphenicol acetyltransferase activity (CAT assay). Induction of a SMLC1 transgene (3920SMLC1) in OP muscles was also observed. Collectively, these in vivo data provide evidence that 1) a mechanical overload inducible element(s) is located between nucleotides -450 and +120 of the human beta-MHC transgene, 2) 3,900 bp of 5' sequence is sufficient to confer mechanical overload induction of a SMLC1 transgene, and 3) the increased expression of slow/type I isomyosin (beta-MHC, SMLC1, SMLC2) in response to mechanical overload is regulated, in part, transcriptionally.

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 Developmental and tissue-specific expression of U4 small nuclear RNA genes.

1988 ◽  
Vol 8 (12) ◽  
pp. 5566-5569 ◽  
Author(s):  
G M Korf ◽  
I W Botros ◽  
W E Stumph
Keyword(s):  
Domestic Chicken ◽  
Sequence Variant ◽  
Small Nuclear Rna ◽  
Specific Expression ◽  
Small Nuclear Rnas ◽  
Specific Manner ◽  
Nuclear Rna ◽  
U4 Rna ◽  
Rna Genes

U4 RNA is one of several small nuclear RNAs involved in the splicing of mRNA precursors. The domestic chicken has two genes per haploid genome that are capable of encoding U4 RNA. The U4X RNA gene (which encodes a sequence variant of U4 RNA that was unknown prior to the cloning of the gene) and the U4B RNA gene were both expressed in vivo in each of seven adult and three embryonic chicken tissues examined. However, the ratio of U4B RNA to U4X RNA can vary more than sevenfold in both a tissue- and stage-specific manner.

Tissue-specific expression of the human CD19 gene in transgenic mice inhibits antigen-independent B-lymphocyte development

1994 ◽  
Vol 14 (6) ◽  
pp. 3884-3894
Author(s):  
L J Zhou ◽  
H M Smith ◽  
T J Waldschmidt ◽  
R Schwarting ◽  
J Daley ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Cell Surface ◽  
Transgenic Mice ◽  
B Cell ◽  
B Lymphocyte ◽  
Cell Development ◽  
B Cell Development ◽  
Specific Expression ◽  
B Lineage

CD19 is a B-cell-specific member of the immunoglobulin superfamily expressed from early pre-B-cell development until plasma cell differentiation. In vitro studies demonstrate that the CD19 signal transduction molecule can serve as a costimulatory molecule for activation through other B-lymphocyte cell surface molecules. However, much remains to be known regarding how CD19 functions in vivo and whether CD19 has different roles at particular stages of B-cell differentiation. Therefore, transgenic mice overexpressing the human CD19 (hCD19) gene were generated to determine whether this transgene would be expressed in a B-lineage-specific fashion and to dissect the in vivo role of CD19 in B-cell development and activation. Expression of the human transgene product was specifically restricted to all B-lineage cells and appeared early in development as occurs with hCD19. In addition, expression of hCD19 severely impaired the development of immature B cells in the bone marrow, with dramatically fewer B cells found in the spleen, peripheral circulation, and peritoneal cavity. The level of hCD19 expressed on the cell surface correlated directly with the severity of the defect in different transgenic lines. These results demonstrate that the hCD19 gene is expressed in a lineage-specific fashion in mice, indicating that the hCD19 gene may be useful for mediating B-lineage-specific expression of other transgene products. In addition, these results indicate an important role for the lineage-specific CD19 molecule during early B-cell development before antigen-dependent activation.

The Vascular Endothelial-Cadherin Promoter Directs Endothelial-Specific Expression in Transgenic Mice

Blood ◽  
1999 ◽  
Vol 93 (1) ◽  
pp. 184-192 ◽  
Author(s):  
S. Gory ◽  
M. Vernet ◽  
M. Laurent ◽  
E. Dejana ◽  
J. Dalmon ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Transgenic Mice ◽  
Dna Sequences ◽  
Vascular System ◽  
Proximal Region ◽  
Vascular Endothelial ◽  
Specific Expression ◽  
Functional Regions ◽  
Vascular Endothelial Cadherin

Vascular endothelial-cadherin (VE-cadherin) is a calcium-dependent adhesive molecule, exclusively and constitutively expressed in endothelial cells. Analysis of the VE-cadherin promoter fused to a reporter gene in bovine aortic endothelial cells showed three major functional regions. The proximal region alone (−139, +24) promoted nonspecific transcription; the addition of the (−289, −140) and (−2226, −1190) domains abolished transcription in fibroblasts while expression in endothelial cells remained unchanged, suggesting that fragments (−2226, +24) and longer contain the full endogenous promoter activity. To study the transcriptional specificity of the promoter region in vivo, we generated transgenic mice carrying the chimeric construct containing the (−2486, +24) region. The promoter directed reporter expression in all examined organs of adult transgenic mice. During embryonic development, transgene expression was detected at the early steps of vasculogenesis. Later, the expression persisted during development of the vascular system and was restricted to the endothelial layer of the vessels. Together, these data provide evidence for specific regulatory regions within the VE-cadherinpromoter. Furthermore, the identification of DNA sequences restricting gene expression to the endothelium has many potential applications for the development of animal models of cardiovascular or angiogenic diseases or for the delivery of therapeutic molecules.

scholarly journals Differential regulation of the N-myc gene in transfected cells and transgenic mice.

1990 ◽  
Vol 10 (5) ◽  
pp. 2096-2103 ◽  
Author(s):  
K Zimmerman ◽  
E Legouy ◽  
V Stewart ◽  
R Depinho ◽  
F W Alt
Keyword(s):  
Transgenic Mice ◽  
Developmental Stages ◽  
Specific Expression ◽  
Endogenous Gene ◽  
Numerous Cell ◽  
Transgenic Lines ◽  
Myc Gene ◽  
Early Developmental

The N-myc gene is expressed specifically in the early developmental stages of numerous cell lineages. To assay for sequences that could potentially regulate N-myc expression, we transfected constructs that contained murine N-myc genomic sequences linked to a reporter gene and genomic clones that contained the complete human or murine N-myc genes into cell lines that either express or do not express the endogenous N-myc gene. Following either transient or stable transfection, the introduced N-myc sequences were expressed regardless of the expression status of the endogenous gene. In contrast, when the clones containing the complete human N-myc gene were introduced into the germline of transgenic mice, expression in some transgenic lines paralleled the tissue- and stage-specific expression of the endogenous murine gene. These findings demonstrate differences in the regulation of N-myc genes in recipient cells following in vitro versus in vivo introduction, suggesting that early developmental events may play a role in the regulation of N-myc expression.

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