绵羊卵母细胞超低温冷冻对四跨膜蛋白CD9表达的影响

doi:10.16431/j.cnki.1671-7236.2016.08.024

Abstract

Abstract:

The experiment was aimed to explore the expression of CD9 in the follicles at different developmental stages, and investigate the effect of in vitro maturation and cryopreservation on CD9. The expression of CD9 protein in the follicle was detected by immunohistochemistry technique, the expression of CD9 mRNA was detected by Real-time quantitative PCR, CD9 protein was detected by Western blotting. The results showed that the CD9 fluorescence signal was detected by immunohistochemistry technique, and the fluorescence signal gradually increased with the maturation of the follicle, and fluorescence signal was the strongest in mature follicle;Real-time quantitative PCR showed that the expression of CD9 mRNA in fresh MⅡ oocytes was significantly increase (P<0.05), and was the least in frozen GV oocytes;The result of Western blotting indicated that CD9 protein was the most in fresh MⅡ oocytes and the expression of frozen GV oocytes was the least, consistent with the results of Real-time quantitative PCR. Cryopreservation of oocytes was more extensive than embryo cryopreservation. Tetraspanins CD9 played a very important role in sperm egg fusion. The above experimental results showed that the cryopreservation made damage to oocytes, decrease content of tetraspanins CD9 protein, and CD9 injury was one of the important reasons for the decline of fertilization rate.

Key words: CD9; ovary; oocytes; in vitro maturation; immunohistochemistry; Western blotting; Real-time quantitative PCR

CLC Number:

S814.8

Wuliyasu, Wuyahan, SHI Zhen-dan, Ailungaowa, Zhamuga, Wuyunsiqin, Enhemarite, Narenhua. Effect of Cryopreservation on the Expression of Tetraspanin CD9 in Sheep Oocytes[J]. , 2016, 43(8): 2104-2111.

References

[1] Jahn R,Grubmüller H.Membrane fusion[J].Cell,2003,112(4):488-495.
[2] Fradkin L G,Kamphorst J T,Aaron D A,et al.Genomewide analysis of the Drosophila tetraspanins reveals a subset with similar function in the formation of the embryonic synapse[J].Proceedings of the National Academy of Sciences of the United States of America,2002,99(21):13663-13668.
[3] Gourgues M,Clergeot P H,Veneault C,et al.A new class of tetraspanins in fungi[J].Biochemical & Biophysical Research Communications,2002,297(5):1197-1204.
[4] Maecker H T,Todd S C,Levy S.The tetraspanin superfamily:Molecular facilitators[J].Faseb Journal Official Publication of the Federation of American Societies for Experimental Biology,1997,11(6):428-442.
[5] Hemler M E.Integrin associated proteins[J].Current Opinion in Cell Biology,1998,10(5):578-585.
[6] Funakoshi T,Tachibana I Y,Kimura H,et al.Expression of tetraspanins in human lung cancer cells:Frequent downregulation of CD9 and its contribution to cell motility in small cell lung cancer[J]. Oncogene,2003,22(5):674-687.
[7] Boucheix C,Rubinstein E.Tetraspanins[J].Cellular & Molecular Life Sciences Cmls,2001,58(9):1189-1205.
[8] Hemler M E.Specific tetraspanin functions[J].Journal of Cell Biology,2001,155(7):1103-1107.
[9] Miyake M,Koyama M,Seno M,et al.Identification of the motility-related protein (MRP-1),recognized by monoclonal antibody M31-15,which inhibits cell motility[J].Journal of Experimental Medicine,1992,174(6):1347-1354.
[10] Miyake M,Nakano K,Ieki Y,et al.Motility related protein 1(MRP-1/CD9) expression:Inverse correlation with metastases in breast cancer[J]. Cancer Research,1995,55(18):4127-4131.
[11] Huang C L,Ueno M D,Masuya D,et al.MRP-1/CD9 gene transduction regulates the actin cytoskeleton through the downregulation of WAVE2[J]. Oncogene,2006,25(49):6480-6488.
[12] Shaw A R,Domanska A,Mak A,et al.Ectopic expression of human and feline CD9 in a human B cell line confers beta 1 integrin-dependent motility on fibronectin and laminin substrates and enhanced tyrosine phosphorylation[J].Journal of Biological Chemistry,1995,270(41):24092-24099.
[13] Lagaudrière-Gesbert C,Le Naour F,Lebel-Binay S,et al.Functional analysis of four tetraspans,CD9,CD53,CD81,and CD82,suggests a common role in costimulation,cell adhesion,and migration:Only CD9 upregulates HB-EGF activity[J].Cellular Immunology,1997,182(2):105-112.
[14] L ffler S,Lottspeich F,Lanza F,et al.CD9,a tetraspan transmembrane protein,renders cells susceptible to canine distemper virus[J].Journal of Virology,1997,71(1):42-49.
[15] 桂朗,王兵,李富花,等.四跨膜蛋白超家族tetraspanins的免疫功能研究进展[J].生物化学与生物物理进展,2008,11:1231-1238.
[16] Naour F L,Rubinstein E,Jasmin C,et al.Severely reduced female fertility in CD9-deficient mice[J]. Science,2000,287(5451):319-321.
[17] Miyado K,Yamada G,Yamada S,et al.Requirement of CD9 on the egg plasma membrane for fertilization[J].Science,2000,287(5451):321-324.
[18] Kaji K,Oda S,Shikano T,et al.The gamete fusion process is defective in eggs of CD9-deficient mice[J].Nature Genetics,2000,24(3):279-282.
[19] Antalíková J,Jankovi?ová J,Simon M,et al.Localization of CD9 molecule on bull spermatozoa:Its involvement in the sperm-egg interaction[J].Reproduction in Domestic Animals,2015,50(3):423-430.
[20] Miyado K,Yoshida K,Yamagata K,et al.The fusing ability of sperm is bestowed by CD9-containing vesicles released from eggs in mice[J].Proc Natl Acad Sci USA,2008,105(35):12921-12926.
[21] Jankovi?ová J,Simon M,Antalíková J,et al.Role of tetraspanin CD9 molecule in fertilization of mammals[J].Physiological Research,2015,64(3):279-293.
[22] 闵江涛,杜卫华,赵学明,等.绵羊卵母细胞采集、体外成熟和胚胎体外培养对体外受精技术效率的影响[J].中国畜牧兽医,2014,41(5):202-207.
[23] Karolina Klinovska D H.Sperm-egg fusion:A molecular enigma of mammalian reproduction[J].International Journal of Molecular Sciences,2014,15(6):10652-10668.
[24] Satouh Y,Inoue N,Ikawa M,et al.Visualization of the moment of mouse sperm-egg fusion and dynamic localization of IZUMO1[J].Journal of Cell Science,2012,125(21):4985-4990.
[25] Masahito I,Naokazu I,Benham A M,et al.Fertilization:A sperm's journey to and interaction with the oocyte[J]. Journal of Clinical Investigation,2010,120(4):984-994.
[26] Inoue N,Ikawa M,Isotani A,et al.The immunoglobulin superfamily protein Izumo is required for sperm to fuse with eggs[J].Nature,2005,434(7030):234-238.
[27] Satouh Y,Inoue N,Ikawa M,et al.Visualization of the moment of mouse sperm-egg fusion and dynamic localization of IZUMO1[J].Journal of Cell Science,2012,125(21):4985-4990.
[28] Coonrod S A,Naaby S.Treatment of mouse oocytes with PI-PLC releases 70-kDa (pI 5) and 35-to 45-kDa (pI 5.5) protein clusters from the egg surface and inhibits sperm-oolemma binding and fusion[J].Developmental Biology,1999,207(2):334-349.
[29] Alfieri J A,Martin A D,Junji T,et al.Infertility in female mice with an oocyte-specific knockout of GPI-anchored proteins[J].Journal of Cell Science,2003,116(Pt11):2149-2155.
[30] Enrica B,Brendan D,David G,et al.Juno is the egg Izumo receptor and is essential for mammalian fertilization[J]. Nature,2014,508(7497):483-487.
[31] Enrica B,Wright G J.Izumo meets Juno:Preventing polyspermy in fertilization[J]. Cell Cycle,2014,13(13):2019-2020.
[32] He Z Y,Brakebusch C,Fassler R,et al.None of the integrins known to be present on the mouse egg or to be ADAM receptors are essential for sperm-egg binding and fusion[J].Developmental Biology,2003,254(2):226-237.
[33] Chen M S,Tung K S,Coonrod S A,et al.Role of the integrin-associated protein CD9 in binding between sperm ADAM 2 and the egg integrin alpha 6beta 1 implications for murine fertilization[J]. Proceedings of the National Academy of Sciences,1999,961(21):11830-11835.
[34] Kaji K,Oda S,Miyazaki S,et al.Infertility of CD9-deficient mouse eggs is reversed by mouse CD9,human CD9,or mouse CD81;polyadenylated mRNA injection developed for molecular analysis of sperm-egg fusion[J]. Developmental Biology,2002,247(2):327-334.
[35] 周光斌,曾艳,孟庆勇,等.超低温冷冻对牛卵母细胞组蛋白乙酰化和膜蛋白CD9表达的影响[J].中国兽医学报,2014,6:999-1004.
[36] Myriam C,Virginie B L,Benjamin R,et al.Binding of sperm protein Izumo1 and its egg receptor Juno drives Cd9 accumulation in the intercellular contact area prior to fusion during mammalian fertilization[J]. Development,2014,141(19):3732-3739.

Effect of Cryopreservation on the Expression of Tetraspanin CD9 in Sheep Oocytes

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