Gene Networks Database

Strongylocentrotus purpuratus Genes in Development: Epidermal growth factor-related proteins

SpEGF I

Function

The sea urchin SpEGF I gene belongs to a growing family of developmentally important genes which encode proteins that contain repeated epidermal growth factor-like motifs (Bisgrove et al., 1991).

Protein

Polyclonal antisera produced from SpEGF I fusion proteins recognize two glycoproteins of 145 and 185 kDa, which were named fibropellins. They contain repeated epidermal growth factor-like motifs (12 and 20 correspondingly) separating an amino terminal domain similar to complement C1s and a carboxy terminal domain similar to avidin (Delgadillo-Reynoso et al., 1989; Hunt and Barker, 1989). Both proteins posess a hydrophobic leader but lack any apparent membrane spanning or anchoring domains (Bisgrove et al., 1991).
SWISS_PROT: P10079 (long variant)

Subcellular location

Immunofluorescence assay demonstrated that the fibropellins are stored in cytoplasmic vesicles in the oocyte. Fibropellins are released soon after fertilization in a distinct secretory event following the exocytosis of cortical granule contents.
During cleavage, these proteins are restricted to the apical surface of the blastomeres and are excluded from the contact zone between blastomeres. At the mesenchyme blastula stage fibropellins are present at the apices of cells of the blastoderm (Bisgrove et al., 1991).

Expression Pattern

SpEGF I encodes two abundant transcripts of 3 and 4 kb expressed both maternally and zygotically (Hursh et al., 1987). In late embryonic and larval stages, the transcripts are localized predominantly in ectodermal tissues (Grimwade et al., 1991).
Fibropelins are present in unfertilized oocytes as well as throughout embryogenesis and early larval development.
There is a noticable increase in the relative abundance of SpEGF I protein between early cleavage and mesenchyme blastula stages and a slight increase during subsequent development.
Following fertilization the fibropellins accumulate at the surface of the oocyte and are localized to the hyaline layer. At the mesenchyme blastula stage, the fibropellins are found on the surface of the cells of the blastoderm and are absent from the inside of the blastocoel and the ingressing primary mesenchyme cells. During gastrulation as the archenteron invaginates into the blastocoel, the fibropellin layer is carried in with the invaginating cells and lines the inner surface of the archenteron. At this stage fibropellins continue to be expressed at the surface of the embryo. The fibropellins disappear from the archenteron as development proceeds; by the time the gut has differentiated in the early pluteus larve, these proteins are restricted to the apical side of the epidermis.
Staining of whole embryos by indirect immunofluorescence revealed that the fibropellins form a matrix covering the surface of the embryo. The matrix is initially amorphous but increases in structural complexity during development. In early embryos removed from the fertilization envelope prior to fixation, the fibropellin matrix appears as irregular patches of fluorescence at the surface of the blastomeres.
After hatching, the matrix becomes organized into a tight meshwork of distinct fibers. At the mesenchyme blastula stage, the interstices of mesh are about 6mm and the fibers range from about 0.3 to 0.4 mm in diameter. The matrix surrounds the bases of the cilia.
At the gastrula stage, the fibers are longer and thicker, and the interstices of the meshwork are less uniform and are expanded, particularly toward the vegetal pole of the embryo. In prism stage embryos, the meshwork appears more loosely woven that in earlier stages except the apical plate where the matrix formes a dense mat perforated by the cilia of the apical tuft. The meshowrk is not altered at the blastopore where the matrix invaginates with the cells of the archenteron but is compressed over the developing stomadeum.
In the early pluteus larva, the matrix appears to be composed of at least two layers; the outer layer is a loose meshwork of large strands (about 1 mm in diameter) overlying a more basal layer comprised of finer strands (about 0.5 mm in diameter) separated by smaller interstices. The fibrous meshwork is somewhat tighter over the oral ectoderm, and the matrix is very dense over the ciliated cells of the circumoral ciliary band.
The molecular weights of the fibropellins and their pattern of embryonic localization indicate that these proteins form a component of the hyaline layer previously described as the apical lamina (Bisgrove et al., 1991).

Protein level

Temporal accumulation
Method 1: Western blot analysis
Reference: Bisgrove et al., 1991

Stage	Unfertilized egg	8 hr 16-cell embryos	27 hr mesenchyme blastula	44 hr gastrula	68 hr prism	96 hr pluteus
Level	+	+	+ +	+	+	+

Protein spatial localization
Method: Immunofluorescence assay
Reference: Bisgrove et al., 1991

Stage	Unfertilized egg	30 min after fertilization	16 cells	Mesenchyme blastula	Gastrula	Pluteus larva
Tissue	cytoplasmic vesicles	surface of the oocyte, form a descrete component of the ECM associated with the inner surface of the hyaline layer	restricted to the apical surface of the blastomeres, excluded from the contact zone between blastomeres	surface of the cells of the blastoderm, absent from the inside of the blastocoel and the ingressing primary mesenchyme cells	surface of the embryo and the inner surface of the archenteron	disappear from the gut, continue to be restricted to the apical side of the epidermis