The Effects of Polyadenylation Status on MPFs During In Vitro Porcine Oocyte Maturation

Cell Physiol Biochem. 2016;39(5):1735-1745. doi: 10.1159/000447874. Epub 2016 Oct 17.

Abstract

Aims: This study aims to clarify the effects of polyadenylation status on M-phase promoting factors (MPFs) during in vitro porcine oocyte maturation.

Methods: In this study, porcine follicular oocytes from large follicles (> 5 millimeter (mm)) and small follicles (< 3 mm) were examined at different follicular developmental stages. The polyadenylation of maternal mRNAs was inhibited by the addition of 3'-deoxyadenosine (3'-da) during the germinal vesicle (GV)(0 h), GV breakdown (GVBD)(18 h), metaphase I (MI)(28 h), and metaphase II (MII) (44 h) stages. In addition, the expression levels and poly-(A) tail lengths of the maternal mRNAs Cyclin B1 and cell division cycle 2 (Cdc2) were determined by real-time quantitative PCR. Immunofluorescence was used to assess spindle formation and chromosome alignment in the examined oocytes.

Results: In large-follicle oocytes, the effects of inhibiting polyadenylation caused the percentage of mature to be significantly lower for the treated group than for the untreated group (p < 0.01). 3'-da can significantly improve the rate of small oocyte maturation in vitro and inhibits Cdc2 polyadenylation. Cyclin B1 plays a significant role in promoting the maturation of large-follicle oocytes. Polyadenylation contributes to the formation of dominant follicles and facilitates the selection of dominant follicles. However, the inhibition of adenylation affected spindle formation-related propulsion and chromosome alignment in both large- and small-follicle oocytes. The first polar body could not be extruded in certain large follicles.

Conclusions: 3'-da can significantly improve the rate of small oocyte maturation in vitro, but it can also affect spindle formation-related propulsion and chromosome alignment.

MeSH terms

  • Animals
  • CDC2 Protein Kinase / genetics*
  • CDC2 Protein Kinase / metabolism
  • Chromosome Segregation / drug effects
  • Cyclin B1 / genetics*
  • Cyclin B1 / metabolism
  • Deoxyadenosines / pharmacology
  • Female
  • In Vitro Oocyte Maturation Techniques*
  • Metaphase
  • Oocytes / drug effects
  • Oocytes / immunology
  • Oocytes / metabolism*
  • Oocytes / ultrastructure
  • Ovarian Follicle / cytology
  • Ovarian Follicle / drug effects
  • Ovarian Follicle / metabolism
  • Polyadenylation / drug effects*
  • Primary Cell Culture
  • RNA, Messenger / genetics*
  • RNA, Messenger / metabolism
  • Spindle Apparatus / drug effects
  • Spindle Apparatus / genetics
  • Spindle Apparatus / metabolism
  • Spindle Apparatus / ultrastructure
  • Swine

Substances

  • Cyclin B1
  • Deoxyadenosines
  • RNA, Messenger
  • CDC2 Protein Kinase
  • cordycepin