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. 2008 Aug;135(15):2627-36.
doi: 10.1242/dev.016329. Epub 2008 Jul 3.

Role for PADI6 and the cytoplasmic lattices in ribosomal storage in oocytes and translational control in the early mouse embryo

Piraye Yurttas  1 , Alejandra M Vitale, Robert J Fitzhenry, Leona Cohen-Gould, Wenzhu Wu, Jan A Gossen, Scott A Coonrod
Affiliations

Role for PADI6 and the cytoplasmic lattices in ribosomal storage in oocytes and translational control in the early mouse embryo

Piraye Yurttas et al. Development. 2008 Aug.

Abstract

The mechanisms that mediate the establishment of totipotency during the egg-to-embryo transition in mammals remain poorly understood. However, it is clear that unique factors stored in the oocyte cytoplasm are crucial for orchestrating this complex cellular transition. The oocyte cytoplasmic lattices (CPLs) have long been predicted to function as a storage form for the maternal contribution of ribosomes to the early embryo. We recently demonstrated that the CPLs cannot be visualized in Padi6-/- oocytes and that Padi6-/- embryos arrest at the two-cell stage. Here, we present evidence further supporting the association of ribosomes with the CPLs by demonstrating that the sedimentation properties of the small ribosomal subunit protein, S6, are dramatically altered in Padi6-/- oocytes. We also show that the abundance and localization of ribosomal components is dramatically affected in Padi6-/- two-cell embryos and that de novo protein synthesis is also dysregulated in these embryos. Finally, we demonstrate that embryonic genome activation (EGA) is defective in Padi6-/- two-cell embryos. These results suggest that, in mammals, ribosomal components are stored in the oocyte CPLs and are required for protein translation during early development.

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Figures

Figure 1
Figure 1. PADI6 is required for cytoplasmic lattice formation in oocytes and two-cell embryos
(A) PADI6+/+ primordial follicle oocyte. N – nucleus. C – cytoplasm. (B) PADI6-/- primordial follicle oocyte. (C) CPLs are first observed in PADI6+/+ growing oocytes. Rb – ribosome. IS – intermediate structure. CPL – cytoplasmic lattice. (D) CPLs do not form in PADI6-/- growing oocytes. (E) CPLs are also observed in PADI6+/+ fully grown germinal vesicle stage oocytes. (F) CPLs are not observed in PADI6-/- fully grown germinal vesicle stage oocytes. (G) Arrows highlight CPLs in PADI6+/+ two-cell embryos. (H) CPLs are not observed in PADI6-/- two-cell embryos. Scale bar equals 200 nm.
Figure 2
Figure 2. PADI6 is required for incorporation of ribosomal components into the oocyte cytoplasmic lattices
(A) RT-PCR analysis using 18S rRNA primers and western blot analysis using anti-S6 antibodies indicates that levels of ribosomal components are similar between PADI6+/+ and PADI6-/- oocytes. (B) In contrast with PADI6+/+ oocytes, the majority of ribosomal protein S6 partitions in the supernatant of ruptured PADI6-/- oocytes. Oocytes were ruptured in hypotonic buffer, serially centrifuged at 650 and 9,000g for 5 min, and the partitioning of ribosomal components into the supernatant (Sup) and pellet fractions was evaluated by immuno-slot-blot analysis using anti-S6 antibodies. (C) Transmission electron microscopic analysis indicates that while putative CPLs (arrows) are observed in the PADI6+/+ 9,000g oocyte pellet, lattices are not observed in the PADI6-/- oocyte pellet. Arrowheads highlight ribosome-like particles associated with CPLs. Scale bar equals 100 nm.
Figure 3
Figure 3. Loss of PADI6 alters the levels and localization of ribosomal protein S6 and affects protein synthesis in two-cell embryos
(A) PADI6 (P6) colocalizes with ribosomal protein S6 in Triton X-100 extracted GV stage PADI6+/+ oocytes and S6 localization is altered in PADI6-/- oocytes. PADI6 and S6 co-localization in PADI6+/+ oocytes is highlighted in merged image (M). The degree of co-localization (oval) is shown in the scatter plot. (B) Fluorographic analysis of [35S]methionine labeled proteins from PADI6+/+ and PADI6-/- GV stage oocytes. Arrow indicates a protein that, based on its molecular weight (∼72 kDa) and absence from PADI6-/- oocytes, is likely PADI6. (C) PADI6 (P6) colocalizes with S6 at the non-apposed cortical regions of PADI6+/+ two-cell embryo blastomeres while S6 is absent from the embryonic cortex in PADI6-/- two-cell embryos. (D) Western blot analysis of protein extracts indicates that S6 levels are reduced in PADI6-/- two-cell embryos. (E) Fluorographic analysis of [35S]methionine labeled proteins indicates that specific mRNAs are translated at different efficiencies in PADI6-/- two-cell embryos. Asterisk indicates putative spindlin protein. (F) Confocal immunofluorescence analysis confirms that spindlin expression is upregulated in PADI6-/- two-cell embryos.
Figure 4
Figure 4. Embryonic genome activation is defective in PADI6-/- two-cell embryos
(A) Levels of RNA polymerase II (RNA Pol II), phosphorylated RNA Pol II (P-Ser2-CTD), and acetylated histone H4 (H4K5 acetyl) are significantly reduced in PADI6-/- two-cell embryos. Representative nuclei were chosen based on their average signal intensity matching the respective overall average intensity for PADI6+/+ and PADI6-/- embryo nuclei. Arrow indicates accumulation of RNA Pol II outside of the nuclear envelope in PADI6-/- two-cell embryos (B) BrUTP incorporation into the nucleus of PADI6-/- two-cell embryos is significantly reduced. Representative nuclei are shown above bars in histogram. (C) Fluorographic analysis of [35S]methionine labeled and Triton X-100 extracted two-cell embryos indicates that synthesis of Transcription Requiring Complex (TRC) proteins is reduced in PADI6-/- embryos.
Figure 4
Figure 4. Embryonic genome activation is defective in PADI6-/- two-cell embryos
(A) Levels of RNA polymerase II (RNA Pol II), phosphorylated RNA Pol II (P-Ser2-CTD), and acetylated histone H4 (H4K5 acetyl) are significantly reduced in PADI6-/- two-cell embryos. Representative nuclei were chosen based on their average signal intensity matching the respective overall average intensity for PADI6+/+ and PADI6-/- embryo nuclei. Arrow indicates accumulation of RNA Pol II outside of the nuclear envelope in PADI6-/- two-cell embryos (B) BrUTP incorporation into the nucleus of PADI6-/- two-cell embryos is significantly reduced. Representative nuclei are shown above bars in histogram. (C) Fluorographic analysis of [35S]methionine labeled and Triton X-100 extracted two-cell embryos indicates that synthesis of Transcription Requiring Complex (TRC) proteins is reduced in PADI6-/- embryos.
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References

    1. D. E.H. (Academic Press, New York City, 1986).

    1. Schultz RM. Hum Reprod Update. 2002 Jul-Aug;8:323. - PubMed
    1. Bachvarova R, De Leon V. Dev Biol. 1977 Jul 15;58:248. - PubMed
    1. Garcia RB, Pereyra-Alfonso S, Sotelo JR. Differentiation. 1979;14:101. - PubMed
    1. Wassarman PM, Josefowicz WJ. J Morphol. 1978 May;156:209. - PubMed

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