Distal cis-acting elements restrict expression of the CyIIIb actin gene in the aboral ectoderm of the sea urchin embryo

Abstract

The distal region of the S. purpuratus actin CyIIIb gene, between -400 and -1400 nucleotides, contains at least three distinct cis-acting elements (C1R, C1L and E1) which are necessary for correct expression of fusion reporter genes in transgenic sea urchin embryos. The contribution of these elements in the temporal and spatial regulation of the gene was analyzed by single and double site-directed mutagenesis in fusion constructs which carry the bacterial chloramphenicol acetyl transferase (CAT) gene as a reporter. Following microinjection of the transgenes in sea urchin embryos, the activity of the mutants was compared to the wild type in time and space by measuring CAT activity at the blastula and pluteus embryonic stages and by in situ hybridization to the CAT mRNA at pluteus stage. Our results indicate that E1 is involved in the temporal regulation of CyIIIb and that all three elements are necessary and sufficient to confer aboral (dorsal) ectoderm specificity to the proximal promoter. This is achieved by suppressing the promoter's activity in all other tissues by the cooperative interaction of the cis-acting elements. The C1R element, binding site of the nuclear receptors SpCOUP-TF and SpSHR2, is by itself sufficient to restrict expression in the ectoderm, whereas the aboral ectoderm restricted expression requires in addition the presence of both C1L and E1. It is therefore evident, that the actin CyIIIb gene is exclusively expressed in the aboral ectoderm by a combinatorial repression in all other cell lineages of the developing embryo.

Embryonic and post-embryonic utilization and subcellular localization of the nuclear receptor SpSHR2 in the sea urchin

“…In the early embryo, the maternal SpSHR2 receptor, which is present in all nuclei, may function as a transcription factor, which alone or in co-operation with a number of other celltype specific transcription factors regulates, either by activation or repression, the expression pattern of target genes such as the CyIIIb actin (see also Xu et al, 1996). As shown in the present study, SpSHR2 receptor present in blastula nuclear extracts binds in vitro the C1R element located in the upstream promoter region of the CyIIIb actin gene.…”

“…The hormone response element C1R is involved in the spatial repression of CyIIIb in cell types other than the aboral ectoderm. In the aboral ectoderm of the late embryo, in co-operation with the upstream element E1, C1R seems to act as a positive regulatory site (Xu et al, 1996).…”

The developmental transcriptome for Lytechinus variegatus exhibits temporally punctuated gene expression changes

“…Lytechinus variegatus (Lv) is a well-studied Atlantic sea urchin species, and many important insights in signal transduction, patterning, and morphogenesis have been obtained from Lv (Armstrong et al, 1993;Ettensohn and Malinda, 1993;Ruffins and Ettensohn, 1996;Guss and Ettensohn, 1997;Logan et al, 1999;Sherwood and McClay, 1999;Sweet et al, 2002;Beane et al, 2006a;Bradham and McClay, 2006;Croce et al, 2006a;Ettensohn et al, 2007;Bradham et al, 2009;Walton et al, 2009;McIntyre et al, 2013;Saunders and McClay, 2014;Martik and McClay, 2015;Piacentino et al, 2015;Schatzberg et al, 2015;Piacentino et al, 2016a;Piacentino et al, 2016b). Investigations in Strongylocentrotus purpuratus (Sp), a well-studied Pacific sea urchin species, have been particularly important for GRN and cis-regulatory analyses; the latter in particular depend on interspecies comparisons, which have often been made between Sp and Lv (Wei et al, 1995;Xu et al, 1996;Yuh et al, 2001;Yuh et al, 2002;Oliveri et al, 2003;Revilla-i-Domingo et al, 2004;Yuh et al, 2004;Minokawa et al, 2005;Ransick and Davidson, 2006;Lee et al, 2007;Livi and Davidson, 2007;Nam et al, 2007;Ochiai et al, 2008;Sethi et al, 2009;Su et al, 2009; Ben-Tabou de-Leon and Davidson, 2010;Damle and Davidson, 2011;Li et al,...…”

The Developmental Transcriptome forLytechinus variegatusExhibits Temporally Punctuated Gene Expression Changes

“…Lytechinus variegatus (Lv) is a well-studied Atlantic sea urchin species, and many important insights in signal transduction, patterning, and morphogenesis have been obtained from Lv (Armstrong et al, 1993;Ettensohn and Malinda, 1993;Ruffins and Ettensohn, 1996;Guss and Ettensohn, 1997;Logan et al, 1999;Sherwood and McClay, 1999;Sweet et al, 2002;Beane et al, 2006a;Bradham and McClay, 2006;Croce et al, 2006a;Ettensohn et al, 2007;Bradham et al, 2009;Walton et al, 2009;McIntyre et al, 2013;Saunders and McClay, 2014;Martik and McClay, 2015;Piacentino et al, 2015;Schatzberg et al, 2015;Piacentino et al, 2016a;Piacentino et al, 2016b). Investigations in Strongylocentrotus purpuratus (Sp), a well-studied Pacific sea urchin species, have been particularly important for GRN and cis-regulatory analyses; the latter in particular depend on interspecies comparisons, which have often been made between Sp and Lv (Wei et al, 1995;Xu et al, 1996;Yuh et al, 2001;Yuh et al, 2002;Oliveri et al, 2003;Revilla-i-Domingo et al, 2004;Yuh et al, 2004;Minokawa et al, 2005;Ransick and Davidson, 2006;Lee et al, 2007;Livi and Davidson, 2007;Nam et al, 2007;Ochiai et al, 2008;Sethi et al, 2009;Su et al, 2009;Ben-Tabou de-Leon and Davidson, 2010;Damle and Davidson, 2011;…”