Mitofusin-2 (Mfn2) is essential for embryonic development, anti-apoptotic events, protection against free radical-induced lesions, and mitochondrial fusion in many cells. However, little is known about its mechanism and function during oocyte maturation. In this study, we found that Mfn2 was expressed in the cytoplasm during different stages of mouse oocyte maturation. Mfn2 was mainly associated with α-tubulin during oocyte maturation. Knockdown of Mfn2 by specific siRNA injection into oocytes caused the mitochondrial morphology and quantity to change, resulting in severely defective spindles and misaligned chromosomes. This led to metaphase I arrest and the failure of first polar body extrusion. Furthermore, Mfn2 depletion from GV stage oocytes caused the redistribution of p38 MAPK in oocyte cytoplasm. These findings provide insights into potential mechanisms of Mfn2-mediated cellular alterations, which may have significant implications for oocyte maturation.