Direct molecular interaction of a conserved yolk granule protein in sea urchins.
The regulation of yolk storage in oocytes and subsequent utilization in embryos is critical for embryogenesis. In sea urchins, the major yolk protein is made in the intestines, transported to the ovaries and accumulated in developing oocytes within membrane-bound vesicles comprising approximately 10% of the mass of an egg. Here, a non-yolk protein that accumulates specifically in yolk granules is reported. This protein was identified by cDNA cloning and, by use of antibodies to the recombinant protein, it was shown that this molecule is stored selectively in yolk granules of oocytes and embryos. No accumulation was seen in the accessory cells, testis, or intestines. In situ ribonucleic acid (RNA) hybridizations showed that the transcript accumulated only in oocytes, and was more highly concentrated in young oocytes. However, later in oogenesis, the messenger ribonucleic acid (mRNA) levels decreased significantly so that no signal was detectable in mature haploid eggs or at any later stage in development. However, by immunofluorescence and western blot analysis, the 30 kDa band was present throughout development. The predicted sequence of this protein shows that it is a member of the bep, HLC-32, EBP family of sea urchin proteins, but as it does not accumulate at the cell surface, nor in the hyaline layer in the two species studied here, as do other members of the family, it has been referred to as YP30 (30 kDa protein of the yolk platelet). To address its potential function, yeast two-hybrid analysis was performed to screen for proteins that potentially interact with YP30. It was found that it binds itself, and forms strongly interacting dimers. It is hypothesized that YP30 participates in the packaging and storage of major yolk protein during oogenesis, or in the utilization of the major yolk protein in development.
Wessel
G M
GM
Department of Molecular and Cell Biology and Biochemistry, Brown University, Providence, RI 02912, USA. rhet@brown.edu
Zaydfudim
V
V
Hsu
Y J
YJ
Laidlaw
M
M
Brooks
J M
JM
D000595:Amino Acid Sequence; D000818:Animals; D018076:DNA, Complementary; D004527:Egg Proteins / Q000235:genetics / Q000378:metabolism*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D010063:Ovum / Q000378:metabolism / Q000648:ultrastructure; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D012617:Sea Urchins / Q000196:embryology*; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques
D018076:DNA, Complementary; D004527:Egg Proteins; D012333:RNA, Messenger; C419312:yolk protein, 30-kDa