1Department of Biomedical Science and Technology, Institute of Biomedical Science and Technology, Konkuk University, Seoul 05029, Korea 2Department of Biomedical Science, CHA University, Seongnam 13884, Korea 3Department of Veterinary Medicine, Konkuk University, Seoul 05029, Korea
Received: October 12, 2016; Revised: December 2, 2016. Accepted: December 7, 2016. Published online December 17, 2016. Hyejin Shin and Dong-Won Seol contributed equally to this work.
Objective: The early growth response (Egr) family consists of four members (Egr1, Egr2, Egr3, and Egr4) that are zinc finger transcription factors. Among them, Egr3 is involved in transcriptional regulation of target genes during muscle spindle formation and neurite outgrowth. We previously showed that the immunoreactive Egr3 is localized on oocyte spindle and accumulate near the microtubule organizing center during meiosis I in mice. Egr3 was also shown to be localized on spermatocytes. We herein investigated if Egr3 is expressed in mouse gonads and if Egr3 blockade results in any defect in oocyte maturation. Method: Expression of Egr3 in mouse gonads was examined by reverse transcriptionpolymerase chain reaction. Fulllength Egr3 and truncated Egr3 (ΔEgr3) complementary RNAs (cRNAs) with Xpress tag at Nterminus and DsRed2 at Cterminus, and small interfering RNA (siRNA) targeting Egr3 were microinjected into mouse oocytes at germinal vesicle stage. Localization of microinjected Egr3 was examined by confocal live imaging and immunofluorescence staining. Results: Egr3 mRNA was detected in mouse ovaries and testes from 1 to 4 weekold mice. An uncharacterized longer transcript containing 5’untranslated region was also detected in 3 and 4 weekold gonads. Microinjected XpressEgr3DsRed2 or XpressΔEgr3DsRed2 localized to nuclei and chromosomes during meiotic progression. Microinjection of these cRNAs or Egr3 siRNA in oocytes did not affect meiotic maturation. Immunofluorescence staining of Egr3 in XpressΔEgr3DsRed2injected oocytes showed a positive signal only on meiotic spindle, suggesting that this antibody does not detect endogenous or exogenous Egr3 in mouse oocytes. Conclusion: The results show that Egr3 localizes to chromosomes during meiotic progression and that certain antibodies may not faithfully represent localization of target proteins in oocytes. Egr3 seems to be dispensable during oocyte maturation in mice.
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