Molecular characterization of egg envelope glycoprotein ZPD in the ovary of Japanese quail (Coturnix japonica).
The egg envelope surrounding avian oocytes exhibits a three-dimensional network of coarse fibers between the granulosa cells and the oocyte. Our previous studies have demonstrated that one of the matrix's components, ZP3, is synthesized in the ovarian granulosa cells. Another component, ZP1, which is critically involved in triggering the sperm acrosome reaction, is synthesized in the liver. We have previously isolated cDNAs encoding quail ZP3 and ZP1, and we now report the isolation of cDNA encoding quail ZPD. By RNase protection assay and in situ hybridization, we have demonstrated that ZPD transcripts are restricted to the granulosa cells of preovulatory follicles. The expression level of ZPD increased progressively during follicular development, and the highest expression was observed in the largest follicles. Western blot analyses using the specific antibody against ZPD indicate that the 40 kDa protein is the authentic ZPD, and the contents of ZPD protein also increased during follicular development. Moreover, we found that the addition of FSH to the culture media enhances the ZPD secretion in the cultured granulosa cells. Two-dimensional gel electrophoresis revealed the presence of several ZPD isoforms with different pI values ranging from 5.5 to 7. Immunohistochemical analyses indicate that the materials recognized with anti-quail ZPD antibody were accumulated in the egg envelope of large yellow follicles. These results demonstrate the presence of ZPD protein in the egg envelope, and that the amount of ZPD in the egg envelope as well as the mRNA in the cells increases at the latter stages of folliculogenesis.
Sato
Tsukasa
T
Department of Applied Biological Chemistry, Faculty of Agriculture, Shizuoka University, Shizuoka, Japan.
Kinoshita
Mihoko
M
Kansaku
Norio
N
Tahara
Kenichi
K
Tsukada
Akira
A
Ono
Hiroko
H
Yoshimura
Takashi
T
Dohra
Hideo
H
Sasanami
Tomohiro
T
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002478:Cells, Cultured; D003370:Coturnix / Q000378:metabolism*; D004527:Egg Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D005260:Female; D005640:Follicle Stimulating Hormone / Q000494:pharmacology; D006023:Glycoproteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D006107:Granulosa Cells / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D008562:Membrane Glycoproteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D006080:Ovarian Follicle / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis*; D011956:Receptors, Cell Surface / Q000032:analysis / Q000235:genetics*; D013268:Stimulation, Chemical; D014817:Vitelline Membrane / Q000737:chemistry*; D000072216:Zona Pellucida Glycoproteins
D004527:Egg Proteins; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D000072216:Zona Pellucida Glycoproteins; D005640:Follicle Stimulating Hormone