| Project/Area Number |
10470011
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (B).
|
| Allocation Type | Single-year Grants |
|---|
| Section | 一般 |
|---|
| Research Field |
General physiology
|
|---|
| Research Institution | Tokyo Women's Medical University |
|---|
Principal Investigator |
MIYAZAKI Shunichi Tokyo Women's Medical Univ, Dept of Physiology, Professor, 医学部, 教授 (80010081)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
KOHCHI Zen Tokyo Women's Medical Univ, Dept of Physiology, Research Assistant, 医学部, 助手 (70322485)
AWAJI Takeo Tokyo Women's Medical Univ, Dept of Physiology, Research Assistant, 医学部, 助手 (60297546)
ODA Shoji Tokyo Women's Medical Univ, Dept of Physiology, Research Assistant, 医学部, 助手 (50266714)
MOHRI Tatsuma National Institute for Physiologocal Sciences Laboratory of Intracellular Metabolism, Research Assistant, 細胞内代謝部門, 助手 (60290912)
|
|---|
| Project Period (FY) |
1999 – 2000
|
| Project Status |
Completed (Fiscal Year 2000)
|
| Budget Amount *help |
¥11,600,000 (Direct Cost: ¥11,600,000)
Fiscal Year 2000: ¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 1999: ¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 1998: ¥5,600,000 (Direct Cost: ¥5,600,000)
|
|---|
| Keywords | Fertilization / Mammalian Egg / Intracellular Ca^<2+> / Ca^<2+> Oscillation / Egg Activation / Sperm Factor / 細胞内カルシウムイオン / カルシウムオシレーション / 精子蛋白質 |
|---|
| Research Abstract |
Cytoplasmic Ca^<2+> is a key factor that regulates important cellular functions. Repetitive rises in intracellular Ca^<2+> concentration ([Ca^<2+>]_i) called "Ca^<2+> oscillations" occur upon stimulation of various cells. In eggs, a dramatic [Ca^<2+>]_i rise occurs at fertilization and triggers egg activation, commom to all species. We have found that fertilized mammalian eggs show ca^<2+> oscillations. This research aimed to elucidate the mechanism of the action of a Ca^<2+> oscillations-inducing protein (COIP) and identify COIP which is thought be derived from the sperm. We obtained the following results.
1) We performrd spatiotemporal analysis of [Ca^<2+>]_i rises in mouse eggs at fertilization (published in Dev. Biol.), upon injection of a spermatozoon (Cell Calcium) or sperm extract (Dev. Biol.) into eggs, and found that COIP exists in the sperm cytoplasm and produces Ca^<2+> oscillations similar to those seen at fertilization. The egg cortex is sensitive to COIP which causes Ca^<2
… More
+> release from the endoplasmic reticulum (ER) via IP_3 receptors. Ca^<2+> entry from outdide the egg is necessary to refill the ER (Cell Calcium). We found that COIP also functions in the ascidia (Development).
2) We found that progesterone and cyclic GMP/AMP cause a [Ca^<2+>]_i, rise in the mouse sperm, leading to the acrosome reaction which is prerequisite for sperm-egg fusion (Biol. Reprod.). In the mice which lack surface protein CD9, sperm-egg fusion was defective, and [Ca^<2+>]_i rise was lacking, suggesting that COIP is introduced through sperm-egg fusion (Nature Genetics).
3) Round spermatids (immature sperm) lack COIP.We injected a spermatid together with sperm extract into a mouse egg and succeeded in fertilization, enbryonic development, and offspring (Mol. Human Reprod.). Thus, COIP is available for practical use.
4) We purified COIP from hamster sperm extract by treating with ammonium sulfated and passing various chromatography. After determinating amino-acid sequence in the terminus, we cloned cDNA of a COIP candidate, and synthesized it in E.Coli. However, this protein alone did not exhibit the COIP activity. Multiple factors may be necessary for function of COIP.We have been continuing to purifiy COIP from pig testis which provides much more samples. Less
|
