The Functions Of GBF1 On Oocyte Maturation

In mammals,oocyte meiotic maturation differs from mitosis in that oocytes maturation is a unique process of cell division during which oocytes undergo the separation of genetic material and asymmetric cell division.High quality mature oocyte is the basis for successful subsequent fertilization and normal embryo development.Cytoplasmic maturation is an important part of mammalian oocyte maturation.The maturation quality of oocytes can be affected by external environmental factors or in vivo molecular regulation.The expression and accumulation of m RNA,protein,substrates and transcription factors in organelles and the recombination of organelles are indispensable parts of cytoplasmic maturation.The proper localization,morphology,and biochemical properties of organelles ensure that oocytes posses high developmental potential.Vesicle transport is used to maintain intracellular communication between organelles.and the first step in vesicle formation requires activation of small GTPase Arf family members.In addition,Arf also regulates the actin cytoskeleton,affecting Golgi apparatus integrity,chromosome segregation and cleavage furrow formation.Activation of Arf requires guanine nucleotide exchange factor(GEF)catalysis through the conserved Sec7 domain.As a member of guanine nucleotide exchange factor,GBF1 can regulates Arf alternating between inactive state(GDP-)and active state(GTP-).In eukaryotic cells,GBF1 locates in the medial compartment of cis Golgi apparatus and endoplasmic reticulum-Golgi apparatus and participates in endoplasmic reticulum-Golgi apparatus transport by assisting in the recruitment of coat protein COPI.GBF1 is necessary for the formation,differentiation and transport of pre-Golgi intermediates and for the maintenance of Golgi integrity.However,the role of GBF1 in meiosis has not been revealed.In this study,ICR female mice oocytes were used as the model and Golgicide A,the specific inhibitor of GBF1,was used to explore the role of GBF1 in the meiosis.Experiments were carried out in the following aspects: 1.The expression level and localization of GBF1 protein in mouse oocytes at different developmental stages were detected.2.The rate and the size of the first polar body after GBF1 inhibitor treatment were counted.3.Observe the distribution and functional protein GM130 changes of Golgi apparatus after inhibitor treatment.4.The distribution pattern of endoplasmic reticulum and endoplasmic reticulum stress response were observed after treatment with inhibitors.5.Observed the changes of mitochondrial distribution and membrane potential after inhibitor treatment.The final results are as follow: First,GBF1 protein was consistently expressed in all stages of mouse oocytes during meiosis.From MI phase to MII phase during meiosis,GBF1 was mainly distributed around the spindle,and its localization pattern was similar to that of Golgi apparatus.Compared with control group,inhibition of GBF1 activity had no effect on polar body extrusion but resulted in the production of large polar bodies.Abnormal accumulation of Golgi apparatus around the spindle in oocytes treated with GBF1 inhibitors might caused by GBF1 affecting GM130 localization.In addition,the loss of GBF1 activity affects the distribution of ER and causes ER stress.Mitochondrial localization and function were also affected by inhibition of GBF1 activity,leading to a decrease in mitochondrial membrane potential level.In summary,this study shows that GBF1 participates the function and distribution of Golgi apparatus,endoplasmic reticulum and mitochondria in the cytoplasm during mouse oocytes meiosis.