Functional cooperation of lens-specific and nonspecific elements in the delta 1-crystallin enhancer.

Abstract

The expression of the chicken 81-crystallin gene is primarily regulated by the action of a lens-specific enhancer 1 kilobase long and located in the third intron of the gene (S. Hayashi, K. Goto, T. S. Okada, and H. Kondoh, Genes Dev. 1: [818][819][820][821][822][823][824][825][826][827][828] 1987). The 120-base-long core segment is required for the activity of the 61-crystallin enhancer but by itself shows no activity. We analyzed the action of the core and adjoining segments of the 81-crystallin enhancer by two different approaches: (i) multiplication of the segments to express any cryptic effect and (ii) competition among enhancers for nuclear factors involved in enhancer action. We found that (i) the core defines a strictly lens-specific element, (ii) an adjoining segment defines an element with a broad specificity with regard to cell type, (iii) these elements cooperate in cis within the 81-crystallin enhancer, (iv) the multimers of these elements compete with each other and with 81-crystallin and simian virus 40 enhancers in trans apparently without sequence specificity but in a fashion reflecting the strength of the enhancers, and (v) the enhancers in trans do not affect the expression of enhancer-free genes, thereby ruling out the possibility of competition for general transcription factors. The last two observations raise the possibility that the enhancer segments interacting with different sequence-specific factors also interact with one other component involved in enhancer action.Tissue-specific regulation of animal genes is in large part ascribed to transcriptional regulatory elements which function in a particular set of cell types. Such elements are often assigned to enhancers which are found in the upstream, intragenic, or downstream regions of a gene. The mechanisms by which the enhancers modulate (mostly activate) the expression of a nearby gene are yet to be clarified. Recent studies on the DNA sequences of enhancers have shown that there are specific binding sites for nuclear proteins, some of which are ubiquitous and some of which are present only in certain groups of cell types (see references cited by Ondek et al. [12] and Fromental et al. [2]). It is a widely accepted hypothesis that the binding of these nuclear protein factors is directly or indirectly involved in enhancer action and/or in generating the tissue specificity of gene expression. However, an essential part of the hypothesis, namely, how such interactions result in the modulation of transcription, has remained obscure.We discovered a highly tissue-specific enhancer element in the third intron of the 81-crystallin gene of chickens (5). This enhancer not only activates reporter genes such as tkCAT (Fig. 1A) in a lens-specific manner (5, 10) but also is essential for the expression of the 5-crystallin gene itself (5). The 51-crystallin enhancer spans a region of 1 kilobase (Fig. 1B) and can be shortened to some extent without loss of the activity if it contains the core region, but further dissection into fragments of a few hun...

Xenopus gamma-crystallin gene expression: evidence that the gamma-crystallin gene family is transcribed in lens and nonlens tissues.

“…The a-crystallins share sequence similarity with small heat shock proteins, and the oaB promoter is able to confer heat inducibility on a reporter gene in transient-transfection assays of NIH 3T3 cells (23). In addition, aB has been shown to accumulate in nonlens tissues under certain disease conditions, suggesting that it is subject to stress regulation (9,20), and was recently induced under conditions of hypertonic stress in primary cultures of lens and kidney cells (5 Although enhancer core sequences promote lens expression, expression from a heterologous promoter can be induced in several different cell types by using sequences from the entire 81 enhancer (12,15,38). In contrast to differential expression being specified by enhancer sequences, there is also at least one example of alternative tissue-preferred promoters directing crystallin expression in lens versus nonlens tissue (11); however, this is not a generalized crystallin control mechanism (18).…”

Puzzle of crystallin diversity in eye lenses

“…Transfection (Hayashi et al, 1987;Thomas et al, 1990;Goto et al, 1990) and transgenic mouse (Kondoh et al, 1987) experiments have established the importance of transcriptional events for controlling 6-crystallin gene expression in lens and non-lens tissues. For example, transgenic mice expressed a fusion gene composed of a 62-crystallin promoter fragmenthacterial chloramphenicol acetyltransferase reporter genel62-crystallin enhancer (from intron 3) in the lens, neuroretina and brain (Purkinje cells of the cerebellum), reminescent of the expression pattern of the ASLl62-crystallin gene in the chicken embryo (Li et al, 1993a(Li et al, , 1993b; see cover immunofluorescence photomicrograph of this issue).…”

Interplay of Pax6 and SOX2 in lens development as a paradigm of genetic switch mechanisms for cell differentiation

“…The DC5 sequence by itself shows lens-specific enhancer activity, although it is much weaker than that of the full-length enhancer, and the lens-specific enhancer activity is clearly demonstrable using its multimeric forms (Goto, et al, 1990;Funahashi, et al, 1991;Kamachi and Kondoh, 1993). The first nuclear factor cloned based on its specific binding to the DC5 sequence was δEF1 (also called ZEB (Genetta, et al, 1994)), which has two zinc finger clusters and a homeodomain, and which became the vertebrate founder of ZFHX1 family transcription factors (Funahashi, et al, 1991;Funahashi, et al, 1993).…”

“…The DC5 sequence by itself shows lens-specific enhancer activity, although it is much weaker than that of the full-length enhancer, and the lens-specific enhancer activity is clearly demonstrable using its multimeric forms (Goto, et al, 1990;Funahashi, et al, 1991;Kamachi and Kondoh, 1993). (Kondoh, et al, 1983).…”