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Protein phosphatase 2A regulatory subunit B55? functions in mouse oocyte maturation and early embryonic development.

ABSTRACT: Protein phosphatase 2A regulatory subunit B55? (PP2A-B55?) has been studied in mitosis. However, its functions in mammalian meiosis and early embryonic development remain unknown. Here, we report that PP2A-B55? is critical for mouse oocyte meiosis and preimplantation embryo development. Knockdown of PP2A-B55? in oocytes led to abnormal asymmetric division, disordered spindle dynamics, defects in chromosome congression, an increase in aneuploidy, and induction of the DNA damage response. Moreover, knockdown of PP2A-B55? in fertilized mouse zygotes impaired development to the blastocyst stage. The impairment of embryonic development might have been due to induction of sustained DNA damage in embryos, which caused apoptosis and inhibited cell proliferation and outgrowth potential at the blastocyst stage. Overall, these results provide a novel insight into the role of PP2A-B55? as a novel meiotic and embryonic competence factor at the onset of life.

SUBMITTER: Liang S

PROVIDER: S-EPMC5432312 | biostudies-literature | 2017 Apr

REPOSITORIES: biostudies-literature

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Protein phosphatase 2A regulatory subunit B55α functions in mouse oocyte maturation and early embryonic development.

Liang Shuang S Guo Jing J Choi Jeong-Woo JW Shin Kyung-Tae KT Wang Hai-Yang HY Jo Yu-Jin YJ Kim Nam-Hyung NH Cui Xiang-Shun XS

Oncotarget 20170401 16

Protein phosphatase 2A regulatory subunit B55α (PP2A-B55α) has been studied in mitosis. However, its functions in mammalian meiosis and early embryonic development remain unknown. Here, we report that PP2A-B55α is critical for mouse oocyte meiosis and preimplantation embryo development. Knockdown of PP2A-B55α in oocytes led to abnormal asymmetric division, disordered spindle dynamics, defects in chromosome congression, an increase in aneuploidy, and induction of the DNA damage response. Moreover ...[more]

PMID: 28439046

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Project description:The serine/threonine protein phosphatase 2A (PP2A) is a master regulator of the complex cellular signaling that occurs during all stages of mammalian development. PP2A is composed of a catalytic, a structural, and regulatory subunit, for which there are multiple isoforms. The association of specific regulatory subunits determines substrate specificity and localization of phosphatase activity, however, the precise role of each regulatory subunit in development is not known. Here we report the generation of the first knockout mouse for the Ppp2r2a gene, encoding the PP2A-B55α regulatory subunit, using CRISPR/Cas9. Heterozygous animals developed and grew as normal, however, homozygous knockout mice were not viable. Analysis of embryos at different developmental stages found a normal Mendelian ratio of Ppp2r2a-/- embryos at embryonic day (E) 10.5 (25%), but reduced Ppp2r2a-/- embryos at E14.5 (18%), and further reduced at E18.5 (10%). No live Ppp2r2a-/- pups were observed at birth. Ppp2r2a-/- embryos were significantly smaller than wild-type or heterozygous littermates and displayed a variety of neural defects such as exencephaly, spina bifida, and cranial vault collapse, as well as syndactyly and severe epidermal defects; all processes driven by growth and differentiation of the ectoderm. Ppp2r2a-/- embryos had incomplete epidermal barrier acquisition, associated with thin, poorly differentiated stratified epithelium with weak attachment to the underlying dermis. The basal keratinocytes in Ppp2r2a-/- embryos were highly disorganized, with reduced immunolabeling of integrins and basement membrane proteins, suggesting impaired focal adhesion and hemidesmosome assembly. The spinous and granular layers were thinner in the Ppp2r2a-/- embryos, with aberrant expression of adherens and tight junction associated proteins. The overlying stratum corneum was either absent or incomplete. Thus PP2A-B55α is an essential regulator of epidermal stratification, and is essential for ectodermal development during embryogenesis.

Project description:BackgroundThe adenovirus E4orf4 protein must bind protein phosphatase 2A (PP2A) for its functions.ResultsThe E4orf4 binding site in PP2A was mapped to the α1,α2 helices of the B55α subunit.ConclusionThe E4orf4 binding site in PP2A-B55α lies above the substrate binding site and does not overlap it.SignificanceA novel functional significance was assigned to the α1,α2 helices of the PP2A-B55α subunit. The adenovirus E4orf4 protein regulates the progression of viral infection and when expressed outside the context of the virus it induces nonclassical, cancer cell-specific apoptosis. All E4orf4 functions known to date require an interaction between E4orf4 and protein phosphatase 2A (PP2A), which is mediated through PP2A regulatory B subunits. Specifically, an interaction with the B55α subunit is required for induction of cell death by E4orf4. To gain a better insight into the E4orf4-PP2A interaction, mapping of the E4orf4 interaction site in PP2A-B55α has been undertaken. To this end we used a combination of bioinformatics analyses of PP2A-B55α and of E4orf4, which led to the prediction of E4orf4 binding sites on the surface of PP2A-B55α. Mutation analysis, immunoprecipitation, and GST pulldown assays based on the theoretical predictions revealed that the E4orf4 binding site included the α1 and α2 helices described in the B55α structure and involved at least three residues located in these helices facing each other. Loss of E4orf4 binding was accompanied by reduced contribution of the B55α mutants to E4orf4-induced cell death. The identified E4orf4 binding domain lies above the previously described substrate binding site and does not overlap it, although its location could be consistent with direct or indirect effects on substrate binding. This work assigns for the first time a functional significance to the α1,α2 helices of B55α, and we suggest that the binding site defined by these helices could also contribute to interactions between PP2A and some of its cellular regulators.

Dataset Information

Protein phosphatase 2A regulatory subunit B55? functions in mouse oocyte maturation and early embryonic development.

Publications

Protein phosphatase 2A regulatory subunit B55α functions in mouse oocyte maturation and early embryonic development.

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