SpRunt-1, a New Member of the Runt Domain Family of Transcription Factors, Is a Positive Regulator of the Aboral Ectoderm-SpecificCyIIIAGene in Sea Urchin Embryos

Abstract

In this paper we present a structural and functional characterization of a new sea urchin embryo transcription factor, SpRunt-1. This factor was isolated by means of its specific interaction with a cis-regulatory target site of the CyIIIa gene. Here we show that this target site, the P7I site, is required for normal embryonic activation of CyIIIa x CAT reporter gene constructs. An oligonucleotide affinity column bearing the P7I target site purifies a 21-kDa polypeptide from blastula-stage nuclear extracts, and the amino acid sequence obtained from this polypeptide was used to generate a nucleic acid probe with which the corresponding cDNA was cloned. The cDNA encodes an approximately 60-kDa protein, SpRunt-1, which includes a "runt domain" that is closely homologous to those of Drosophila and mammalian runt domain transcription factors. RNA and genomic blots show that SpRunt-1 is represented by a single embryonic transcript, encoded by one of possibly two runt-domain-containing genes. By RNA probe protection we found that transcripts of SpRunt-1 increase in concentration dramatically after the blastula stage of development, suggesting that the up-regulation of CyIIIa that occurs after blastula stage is a function of zygotically transcribed SpRunt-1. These results are discussed with reference to known features of the runt domain family of transcription factors.

Molecular basis of tissue‐specific gene expression mediated by the Runt domain transcription factor PEBP2/CBF

“…In this regard, it is worth noting that the amino acid sequence within NRDBc is highly conserved in an evolutionary sense (Fig. 5): even sea urchin Runt (Coffman et al 1996) shows a 49% sequence homology with its mouse and human counterpart. Furthermore, the exonVII-like autoinhibitory Transcriptional regulation by PEBP2/CBF ᭧ Blackwell Science Limited Genes to Cells (1999) 4, 685-696 691 Figure 4 Cooperation between ␣B 1 and Ets-1 for DNA binding .…”

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

“…Here the major positive input is provided by the SpRuntl factor. SpRuntl mRNA rises rapidly in the embryo after gastrulation as the middle module becomes the dominant CyIIIa regulatory element (20), and the P1 sites are no longer required for function. Two different negatively acting factors, SpZ12-1 and SpP7II (Fig.…”

“…The SpRunt 1 interaction in the middle module boosts the output of the proximal module early in development (16,20). In addition, the distal region of the CyIIIa cis-regulatory system increases the output of either or both of the two modules if included in expression constructs (16,22,23).…”

“…Seven of the nine transcription factors have been cloned (the eighth is known to be a CCAAT-binding factor). An extensive series of gene transfer experiments, revealing the functions of a different subregion of the regulatory system by means of deletions and mutations, has been carried out by Kirchhamer and Davidson (16), and additional key observations are described in other recent studies (20,21). As discussed in detail in ref.…”

Conservation ofEndo16expression in sea urchins despite evolutionary divergence in both cis and trans-acting components of transcriptional regulation

“…Eggs can be obtained in large quantities and develop synchronously upon fertilization, facilitating the collection of material for biochemical analyses. This has enabled researchers to characterize several promoter sequences in exceptional detail including CyIIIa (Calzone et al, 1988;Theze et al, 1990;Wang et al, 1995;Calzone et al, 1997;Coffman et al, 1996;Coffman et al, 1997) and Endo16 (Yuh et al, 1994;Yuh et al, 1998;Yuh et al, 2001a). Transient expression assays have proven remarkably successful for functional analysis of these promoter sequences in multiple species (reviewed by .…”