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 A combination of closely associated positive and negative cis-acting promoter elements regulates transcription of the skeletal alpha-actin gene.

1990 ◽  
Vol 10 (2) ◽  
pp. 528-538 ◽  
Author(s):  
K L Chow ◽  
R J Schwartz
Keyword(s):  
Transcriptional Activity ◽  
Primary Cultures ◽  
Gene Promoter ◽  
Regulatory Sequence ◽  
Actin Gene ◽  
Cell Type ◽  
Specific Expression ◽  
Cis Acting ◽  
Cell Type Specific Expression ◽  
Alpha Actin

The chicken skeletal alpha-actin gene promoter region provides at least a 75-fold-greater transcriptional activity in muscle cells than in fibroblasts. The cis-acting sequences required for cell type-restricted expression within this 200-base-pair (bp) region were elucidated by chloramphenicol acetyltransferase assays of site-directed Bg/II linker-scanning mutations transiently transfected into primary cultures. Four positive cis-acting elements were identified and are required for efficient transcriptional activity in myogenic cells. These elements, conserved across vertebrate evolution, include the ATAAAA box (-24 bp), paired CCAAT-box-associated repeats (CBARs; at -83 bp and -127 bp), and the upstream T+A-rich regulatory sequence (at -176 bp). Basal transcriptional activity in fibroblasts was not as dependent on the upstream CBAR or regions of the upstream T+A-rich regulatory sequence. Transfection experiments provided evidence that positive regulatory factors required for alpha-actin expression in fibroblasts are limiting. In addition, negative cis-acting elements were detected and found closely associated with the G+C-rich sequences that surround the paired CBARs. Negative elements may have a role in restricting developmentally timed expression in myoblasts and appear to inhibit promoter activity in nonmyogenic cells. Cell type-specific expression of the skeletal alpha-actin gene promoter is regulated by combinatorial and possibly competitive interactions between multiple positive and negative cis-acting elements.

A combination of closely associated positive and negative cis-acting promoter elements regulates transcription of the skeletal alpha-actin gene

1990 ◽  
Vol 10 (2) ◽  
pp. 528-538
Author(s):  
K L Chow ◽  
R J Schwartz
Keyword(s):  
Transcriptional Activity ◽  
Primary Cultures ◽  
Gene Promoter ◽  
Regulatory Sequence ◽  
Actin Gene ◽  
Cell Type ◽  
Specific Expression ◽  
Cis Acting ◽  
Cell Type Specific Expression ◽  
Alpha Actin

The chicken skeletal alpha-actin gene promoter region provides at least a 75-fold-greater transcriptional activity in muscle cells than in fibroblasts. The cis-acting sequences required for cell type-restricted expression within this 200-base-pair (bp) region were elucidated by chloramphenicol acetyltransferase assays of site-directed Bg/II linker-scanning mutations transiently transfected into primary cultures. Four positive cis-acting elements were identified and are required for efficient transcriptional activity in myogenic cells. These elements, conserved across vertebrate evolution, include the ATAAAA box (-24 bp), paired CCAAT-box-associated repeats (CBARs; at -83 bp and -127 bp), and the upstream T+A-rich regulatory sequence (at -176 bp). Basal transcriptional activity in fibroblasts was not as dependent on the upstream CBAR or regions of the upstream T+A-rich regulatory sequence. Transfection experiments provided evidence that positive regulatory factors required for alpha-actin expression in fibroblasts are limiting. In addition, negative cis-acting elements were detected and found closely associated with the G+C-rich sequences that surround the paired CBARs. Negative elements may have a role in restricting developmentally timed expression in myoblasts and appear to inhibit promoter activity in nonmyogenic cells. Cell type-specific expression of the skeletal alpha-actin gene promoter is regulated by combinatorial and possibly competitive interactions between multiple positive and negative cis-acting elements.

scholarly journals Olfactory expression of trace amine-associated receptors requires cooperative cis-acting enhancers

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Ami Shah ◽  
Madison Ratkowski ◽  
Alessandro Rosa ◽  
Paul Feinstein ◽  
Thomas Bozza
Keyword(s):  
Gene Expression ◽  
Large Family ◽  
Sequence Motifs ◽  
Specific Expression ◽  
Cis Acting ◽  
Conserved Sequence ◽  
Trace Amine ◽  
Sequence Elements ◽  
Cell Type Specific Expression ◽  
Cell Type Specific

AbstractOlfactory sensory neurons express a large family of odorant receptors (ORs) and a small family of trace amine-associated receptors (TAARs). While both families are subject to so-called singular expression (expression of one allele of one gene), the mechanisms underlying TAAR gene choice remain obscure. Here, we report the identification of two conserved sequence elements in the mouse TAAR cluster (T-elements) that are required for TAAR gene expression. We observed that cell-type-specific expression of a TAAR-derived transgene required either T-element. Moreover, deleting either element reduced or abolished expression of a subset of TAAR genes, while deleting both elements abolished olfactory expression of all TAARs in cis with the mutation. The T-elements exhibit several features of known OR enhancers but also contain highly conserved, unique sequence motifs. Our data demonstrate that TAAR gene expression requires two cooperative cis-acting enhancers and suggest that ORs and TAARs share similar mechanisms of singular expression.

Cooperativity of sequence elements mediates tissue specificity of the rat insulin II gene

1990 ◽  
Vol 10 (4) ◽  
pp. 1784-1788
Author(s):  
Y P Hwung ◽  
Y Z Gu ◽  
M J Tsai
Keyword(s):  
Beta Cell ◽  
Tissue Specificity ◽  
Promoter Element ◽  
Heterologous Promoter ◽  
Specific Expression ◽  
Tissue Specific ◽  
Cis Acting ◽  
Insulin Promoter ◽  
Sequence Elements ◽  
Flanking Region

The 5'-flanking region of the rat insulin II gene (-448 to +50) is sufficient for tissue-specific expression. To further determine the tissue-specific cis-acting element(s), important sequences defined by linker-scanning mutagenesis were placed upstream of a heterologous promoter and transfected into insulin-producing and -nonproducing cells. Rat insulin promoter element 3 (RIPE3), which spans from -125 to -86, was shown to confer beta-cell-specific expression in either orientation. However, two subregions of RIPE3, RIPE3a and RIPE3b (defined by linker-scanning mutations), displayed only marginal activities. These results suggest that the two subregions cooperate to confer tissue specificity, presumably via their cognate binding factors.

Tissue-specific transcription of the mouse alpha-fetoprotein gene promoter is dependent on HNF-1

1989 ◽  
Vol 9 (10) ◽  
pp. 4204-4212
Author(s):  
M H Feuerman ◽  
R Godbout ◽  
R S Ingram ◽  
S M Tilghman
Keyword(s):  
Specific Binding ◽  
Gene Promoter ◽  
Binding Activity ◽  
Alpha Fetoprotein ◽  
Transient Expression Assay ◽  
Band Shift ◽  
Specific Expression ◽  
Tissue Specific ◽  
Cis Acting ◽  
Dnase I Footprinting

Previous work identified four upstream cis-acting elements required for tissue-specific expression of the alpha-fetoprotein (AFP) gene: three distal enhancers and a promoter. To further define the role of the promoter in regulating AFP gene expression, segments of the region were tested for the ability to direct transcription of a reporter gene in transient expression assay. Experiments showed that the region within 250 base pairs of the start of transcription was sufficient to confer liver-specific transcription. DNase I footprinting and band shift assays indicated that the region between -130 and -100 was recognized by two factors, one of which was highly sequence specific and found only in hepatoma cells. Competition assays suggested that the liver-specific binding activity was HNF-1, previously identified by its binding to other liver-specific promoters. Mutation of the HNF-1 recognition site at -120 resulted in a significant reduction in transcription in transfection assays, suggesting a biological role for HNF-1 in the regulation of AFP expression.

GATA and Ets cis-acting sequences mediate megakaryocyte-specific expression

1993 ◽  
Vol 13 (1) ◽  
pp. 668-676
Author(s):  
V Lemarchandel ◽  
J Ghysdael ◽  
V Mignotte ◽  
C Rahuel ◽  
P H Roméo
Keyword(s):  
Binding Site ◽  
Consensus Sequence ◽  
Mrna Level ◽  
Point Mutations ◽  
Specific Gene ◽  
Cell Type Specificity ◽  
Specific Expression ◽  
Platelet Factor ◽  
Cis Acting ◽  
Dna Fragment

The human glycoprotein IIB (GPIIB) gene is expressed only in megakaryocytes, and its promoter displays cell type specificity. We show that this specificity involved two cis-acting sequences. The first one, located at -55, contains a GATA binding site. Point mutations that abolish protein binding on this site decrease the activity of the GPIIB promoter but do not affect its tissue specificity. The second one, located at -40, contains an Ets consensus sequence, and we show that Ets-1 or Ets-2 protein can interact with this -40 GPIIB sequence. Point mutations that impair Ets binding decrease the activity of the GPIIB promoter to the same extent as do mutations that abolish GATA binding. A GPIIB 40-bp DNA fragment containing the GATA and Ets binding sites can confer activity to a heterologous promoter in megakaryocytic cells. This activity is independent of the GPIIB DNA fragment orientation, and mutations on each binding site result in decreased activity. Using cotransfection assays, we show that c-Ets-1 and human GATA1 can transactive the GPIIB promoter in HeLa cells and can act additively. Northern (RNA) blot analysis indicates that the ets-1 mRNA level is increased during megakaryocyte-induced differentiation of erythrocytic/megakaryocytic cell lines. Gel retardation assays show that the same GATA-Ets association is found in the human GPIIB enhancer and the rat platelet factor 4 promoter, the other two characterized regulatory regions of megakaryocyte-specific genes. These results indicate that GATA and Ets cis-acting sequences are an important determinant of megakaryocytic specific gene expression.

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