Highly identical cassettes of gene regulatory elements, genomically repetitive and present in RNA.

Abstract

A region in the first intron of a metallothionein-encoding gene of the sea urchin Strongylocentrotus purpuratus (SpMTA gene) regulates its 5' promoter activity. Within this region is a 290-bp cassette of six sequence motifs that are present in other genes in this species and posited to operate as regulatory elements. The cassette, present at high multiplicity in the genome, was used to screen genomic DNA clones. Of these, six diverse individuals were partially sequenced and found to have segments 94% identical to the 290-bp cassette in the SpMTA gene. Their next 80 bp diverged from the SpMTA sequence but were highly identical among the six non-SpMTA clones and contained an additional regulatory motif. These diverse clones thus contained 370-bp cassettes of an overall 94% sequence identity and an apparent content of seven regulatory elements. The regulatory cassettes were transposon-like, insofar as the termini of the highly identical regions consisted of 24-to 25-bp inverted repeats, bracketed by 6-to 9-bp direct repeats in the divergent regions. In addition to being in transcripts of the SpMTA intron, the cassette was found in other sea urchin embryo poly(A)+ RNAs, in eggs and embryos, and enriched in pluteus ectoderm. The cassette sequence was present in moderate abundance in transcripts in both sense and antisense orientation. We report the presence of a transposon-like cassette of regulatory elements that is also represented in RNA, which potentially could function differently from previously described transposons.A DNA segment appearing as an insert in the interior of the first intron of the MTA metallothionein (MT) gene (1) of the sea urchin Strongylocentrotus purpuratus (SpMTA gene) distinguishes it from the first intron of another, substantially homologous MT gene in this species, SpMTB1 (2). The interior of the SpMTA intron is largely responsible for the high degree of activity of this gene (3), in contrast to the low-level activity (4) of the SpMTB, gene. This specifically SpMTA DNA segment may be functionally similar to inserted DNA elements, which in various situations can elicit changes in gene regulation (5,6), become, themselves, transcriptionally regulated (7), or produce disruptive and deleterious effects (8,9). Through such an intronic insert, the SpMTA gene may have been recruited evolutionarily into a specifically regulated set, to which the other MT genes in this species do not belong. This insert is particularly interesting because it contains within a stretch of0.3 kb several sequence motifs highly identical to sites in several other genes (10) and specifically implicated as regulatory elements in CyIla (11,12), an actin-encoding gene that has the same aboral ectodermal specificity of expression (13) as the SpMTA gene (2).The premise that this 0.3-kb intronic region, enriched in putative regulatory sites, was indeed inserted into an evolutionary progenitor of the SpMTA gene has led us here to The publication costs of this article were defrayed in part by page charge payment. This a...

Structural and bioinformatic analysis of the Roman snail Cd‐Metallothionein gene uncovers molecular adaptation towards plasticity in coping with multifarious environmental stress

“…The transcription of the MT‐A gene of the sea urchin Strongylocentrotus purpuratus , for example, was significantly enhanced in fused reporter constructs carrying a regulatory sequence from its own intron that contained several repeats in a cassette‐like aggregation, compared to an intronless fusion construct (Bai et al . 1993; Nemer et al . 1993).…”

Modular cis-regulatory organization of developmentally expressed genes: two genes transcribed territorially in the sea urchin embryo, and additional examples.

“…The cis-regulatory regions of several of the Ca2+ binding Spec genes have been shown by Klein and colleagues (64,65) to contain what they term "RSR" modules, which in reporter constructs confer aboral ectoderm expression, where R is a repeated sequence and S is a sequence that contains transcription factor binding sites. In another case a "cassette" or module that includes many of the CyIIIa target sites has been identified in an intron of the sea urchin metallothionein gene, and has been shown to confer aboral ectoderm specific expression to this gene (66,67).…”

Synergistic Activation of theN-Methyl-d-aspartate Receptor Subunit 1 Promoter by Myocyte Enhancer Factor 2C and Sp1

“…DNA content in cell lysates was measured with Hoechst dye 33258 on a TKO 100 fluorometer following the manufacture's protocol (Hoefer Scientific Instrument). Copy numbers of integrated fusion genes in stable PC12 transfectants were measured using a slot blot assay as described previously (44). Briefly, extracted genomic DNA was quantified both by absorbency at 260 nm and by Hoechst dye 33258.…”