Lens-specific promoter activity of a mouse gamma-crystallin gene.

Lok, Si; Breitman, Martin L.; Chepelinsky, Ana B.; Piatigorsky, Joram; Gold, R; Tsui, Lap‐Chee

doi:10.1128/mcb.5.9.2221

Cited by 70 publications

(43 citation statements)

References 59 publications

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“…In fact, wellcharacterized GC box-dependent genes have been either viral or housekeeping cellular genes (e.g., the simian virus 40, herpes simplex virus tk, and dehydrofolate reductase genes), and this is the first example of the involvement of a GC box in the expression of a tissue-specific gene. Among the crystallin genes which have been cloned and whose expression has been characterized (2,4,9,14,19), only the b-crystallin genes have GC boxes in their promoters. It is possible that interactions of a tissue-specific factor(s) with the B-crystallin promoter give rise to a potential for lensspecific b-crystallin expression, and binding of an Spl-like factor to the GC box may activate this expression.…”

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confidence: 99%

In vivo competition of delta-crystallin gene expression by DNA fragments containing a GC box.

Hayashi¹,

Kondoh²

1986

Mol. Cell. Biol.

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Expression of the chicken 8-crystallin gene 1 injected into the nuclei of mouse cells is lens specific. Coinjection of GC box-containing DNA fragments from 8-crystallin, simian virus 40 early, and herpes simplex virus type 1 tk promoters effectively suppressed 8-crystallin expression in the lens, but coinjection with DNA fragments not containing the GC box did not. This suppression was likely due to the competition of an Spl-like transcription factor(s) and indicates involvement of the apparently ubiquitous factor(s) in the tissue-specific expression of the 8-crystallin gene.B-Crystallin is the most abundantly produced lens-specific protein of avians. The cloned 8-crystallin gene 1 (17,18), the active entity of two nonallelic genes of chickens (2,16,23), is expressed with a high lens specificity after nuclear injection into mouse cells (9,11,13). By introducing a series of deletions and substitutions in the gene and its flanking sequences ( Fig. 1), we located the DNA region responsible for the lens-specific gene regulation within 136 base pairs (bp) encompassing the transcription initiation site (from nucleotide positions -81 to + 55) (9). Most remarkably, deletion of the segment from -81 to -65 reduced the level of expression 20-fold. This segment contains the GC box sequence ( Fig. 1) which is often part of viral and animal gene promoters as an essential cis-acting element for transcription initiation (5-7, 10, 15). To characterize regulatory interactions of putative trans-acting cellular factors with the cisacting DNA element of the 5-crystallin gene, we analyzed competition between the nuclear-injected gene and various coinjected DNA fragments.First, a 72-bp fragment of the promoter region (from positions -107 to -36) of the B-crystallin gene (fragment A, Fig. 2B) was tested for its effect on b-crystallin expression upon coinjection with the gene. Two test genes were used: one was the 5-crystallin gene on plasmid pSClB (12) and the other was a chimeric gene (Mov-8 gene) on p5Kp-Mov (11) in which the Moloney murine leukemia virus (MoMuLV) long terminal repeat (LTR) (22) was linked to the 5-crystallin gene at the second intron. The former directs the synthesis of a 50-kilodalton polypeptide (Fig. 3A, lane 1), and the latter directs the synthesis of 48 and 51-kilodalton polypeptides (11) (Fig. 3A, lane 3); therefore, their products could be separately analyzed on an immunoblot (Fig. 3A, lane 2). A mixture of both test genes (125 pLg of each per ml) and fragment A (500 ,ug/ml) was injected into the nuclei of mouse lens epithelial cells in primary culture (13). Assuming an average injection volume of 10 fl (1,3,8), we estimated the numbers of injected competitor fragments and each of the test genes to be 6 x 104 and 60 per nucleus, respectively. The injected cells were harvested 48 h later, and expression of the test genes was examined (Fig. 3A, lane 4) (Fig. 3B). It is difficult to estimate the actual number of regulatory-factor molecules which may bind to the fragment A region of the 5-crystallin promoter, but...

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In vivo competition of delta-crystallin gene expression by DNA fragments containing a GC box.

Hayashi¹,

Kondoh²

1986

Mol. Cell. Biol.

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“…Among other crystallins, a^-crystallin of mouse and chicken and 7-crystallin of mouse have been studied for expression of the cloned gene in cultured cells Okazaki et al 1985;Lok et al 1985) and in transgenic mice (Overbeek et al 1985;Goring et al 1987). In all of these cases, the 5'-flanking sequence of the genes directed lens-specific expression of linked genes, and it was found that the sequences upstream of -100 were essential for the lens specificity.…”

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confidence: 99%

Lens-specific enhancer in the third intron regulates expression of the chicken delta 1-crystallin gene.

Hayashi

Goto²,

Okada³

et al. 1987

Genes Dev.

116

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We have previously shown that the tissue specificity determinant of the chicken ~l-crystallin gene lies 3' of position -100 (Hayashi et al. 1985). Since the promoter of the gene (81-crystallin promoter) did not show any tissue specificity, we examined various segments of the ~l-crystallin gene for a tissue-specific enhancer activity by placing each segment downstream of a heterologous transcriptional unit coding for chloramphenicol acetyltransferase (CAT) and by transfecting chicken tissues in primary culture. We found that a segment spanning the third intron bears a strong lens-specific enhancer activity. This "~l-crystallin enhancer" activates transcription from the 81-crystallin promoter 20-to 40-fold in lens cells and to various degrees with other promoters. Deletion analysis of the enhancer region indicated that it covered nearly 1 kb but did not indicate clear-cut boundaries. For its enhancer effect the core region of 120 bp and associations with certain adjoining regions were required. Removal of the enhancer from the gene totally abolished ~l-crystallin expression, and reinsertion of the enhancer in either upstream, internal, or downstream positions restored expression. We conclude that the ~l-crystallin enhancer is an essential and major determinant for lens-specificity of 81-crystallin expression.

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“…In the otA-crystallin gene, the 5' flanldng sequence of the mouse (8) and hamster (10) is highly conserved; however, much of this similarity is lost in the chicken (21). In the mouse, rat, and human y-crystallin gene family there are several sequences upstream from the TATA box that are highly conserved (19,(22)(23)(24)(25). Except for the 81-crystallin gene (26,27), no other crystallin gene contains the 5'CCAAT3' sequence in their 5' flanking region (5,8,10,19,(22)(23)(24)(25)(28)(29)(30).…”

Section: Introductionmentioning

confidence: 99%

“…It is particularly interesting that crystallin promoters can function in lens cells ofheterologous species, even when the species from which the lens cells were derived do not contain the crystallin gene being tested. For example, the chicken 51-crystallin promoter can function in mouse lens cells (17,18), which do not contain endogenous 8-crystallin genes, and the mouse -y2-crystallin promoter can function in embryonic chicken y2 lens cells (19), which do not contain endogenous -y-crystallin genes.…”

Section: Introductionmentioning

confidence: 99%

“…In the mouse, rat, and human y-crystallin gene family there are several sequences upstream from the TATA box that are highly conserved (19,(22)(23)(24)(25). Except for the 81-crystallin gene (26,27), no other crystallin gene contains the 5'CCAAT3' sequence in their 5' flanking region (5,8,10,19,(22)(23)(24)(25)(28)(29)(30). The CCAAT sequence has been found to interact with a specific trans-acting factor (31)(32)(33).…”

Section: Introductionmentioning

confidence: 99%

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Crystallin genes: lens specificity of the murine alpha A-crystallin gene.

Chepelinsky

Khillan

Mahon

et al. 1987

Environ Health Perspect

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The abundant soluble proteins of the eye lens, the crystallins, are encoded by several gene families which are developmentally regulated in the embryonic lens. We have studied the expression of the murine aA-crystallin gene. Transfection experiments using the pSVO-CAT vector and explanted lens epithelia from embryonic chickens demonstrated proximal (-88 to -60) and distal (-111 to -85) regulatory sequences which interact when the aA-crystallin promoter is activated in the lens cells. Transgenic mouse experiments showed that the sequence between positions -366 to + 46 of the aA-crystallin promoter can drive foreign genes selectively in the lens. A fusion gene consisting of this aA-crystallin promoter sequence and the T-antigen gene of SV40 produced a lens tumor in transgenic mice. Thus, crystallin promoters provide a useful model for tissue-specific gene expression and permit targeting the expression of foreign genes to a highly differentiated tissue during development.

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Lens-specific promoter activity of a mouse gamma-crystallin gene.

Cited by 70 publications

References 59 publications

In vivo competition of delta-crystallin gene expression by DNA fragments containing a GC box.

In vivo competition of delta-crystallin gene expression by DNA fragments containing a GC box.

Lens-specific enhancer in the third intron regulates expression of the chicken delta 1-crystallin gene.

Crystallin genes: lens specificity of the murine alpha A-crystallin gene.

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