Effects of mitofusin 2 excess during oocyte growth in vitro

• Grant number: 18/23734-3
• Support type: Scholarships abroad - Research Internship - Scientific Initiation
• Effective date (Start): February 25, 2019
• Effective date (End): June 24, 2019
• Field of knowledge: Biological Sciences - Genetics
• Principal Investigator: Marcos Roberto Chiaratti
• Grantee: Fabrícia Heloísa Cavicchioli Sugiyama
• Supervisor abroad: Hugh Clarke
• Home Institution: Centro de Ciências Biológicas e da Saúde (CCBS). Universidade Federal de São Carlos (UFSCAR). São Carlos , SP, Brazil
• Local de pesquisa : McGill University, Montreal, Canada
• Associated to the scholarship: 18/06119-3 - Mfn1 knockout in oocytes arrests folliculogenesis in mice through inhibiting the PI3K-Akt pathway, BP.IC

Abstract

The determining role of mitochondria in regulating follicular development and oocyte growth is emerging from basic research on model species and clinical studies on female infertility. This new branch of research has been dedicated to deciphering the relationship between oocyte developmental competence and bioenergetics. Mitochondrial activity depends, among other things, on its organelle dynamics, which in turn is dependent on events of fusion and fission. Mitochondrial fusion relies on the activity of Mitofusins 1 and 2 (MFN1 and MFN2) and on the Protein of Optical Atrophy 1 (OPA1). Recent studies of our group have determined that the conditional knock out (cKO) of Mfn1 in the oocyte affects folliculogenesis, resulting in the blockage of oocyte growth and development leading to lower ATP levels. On the other hand, the cKO of Mfn2 alone or of both Mfn1 and Mfn2 did not affect oocyte growth or its ATP content at the germinal vesicle (GV) stage. This suggested an important role for the relative quantities of MFN1 and MFN2 during oocyte growth, in order to promote proper energy production and development. To test this hypothesis, we cultured Granulosa cells-Oocyte Complexes (GOC) overexpressing MFN2 and MFN1+MFN2. As a result, the oocytes overexpressing MFN2 did not grow, while the ones with higher contents of MFN1+MFN2 grew normally. These results hinted that the excess of MFN2 alone might be responsible for the impairment on oocyte growth. Therefore, we aim with this work to culture Granulosa cells-Oocyte Complexes (GOC) overexpressing MFN1 to confirm that the excess of MFN2 during oocyte growth is the situation resulting in detrimental growth. Specifically, we will use the DNA constructs previously built to overexpress MFN1 during GOC culture to evaluate its effect on oocyte development. If this experiment gives evidence that the relative excess of MFN1 oligomers does not prevent oocyte growth nor changes the oocyte ATP content (similarly to what was seen in the case of Mfn2 cKO) we will then overexpress MFN2 to evaluate whether the excess of MFN2 alters the oocyte ATP levels (similarly to what was seen in the case of Mfn1 cKO). This model is innovative and will enable us to address our hypothesis in a very elegant and thorough way. Thus, we expect with this work to deeper understand the role played by mitofusins and mitochondria during oocyte development. In addition, our group will certainly benefit from the ongoing collaboration and from the training proposed for the student during the internship at McGill University.