New Insights into Endogenous Retrovirus-K Transcripts in Amyotrophic Lateral Sclerosis.

Retroviral reverse transcriptase activity and the increased expression of human endogenous retroviruses (HERVs) are associated with amyotrophic lateral sclerosis (ALS). We were interested in confirming HERVK overexpression in the ALS brain, its use as an accessory diagnostic marker for ALS, and its potential interplay with neuroinflammation. Using qPCR to analyze HERVK expression in peripheral blood mononuclear cells (PBMCs) and in postmortem brain samples from ALS patients, no significant differences were observed between patients and control subjects. By contrast, we report alterations in the expression patterns of specific HERVK copies, especially in the brainstem. Out of 27 HERVK copies sampled, the relative expression of 17 loci was >1.2-fold changed in samples from ALS patients. In particular, the relative expression of two HERVK copies (Chr3-3 and Chr3-5) was significantly different in brainstem samples from ALS patients compared with controls. Further qPCR analysis of inflammation markers in brain samples revealed a significant increase in NLRP3 levels, while TNFA, IL6, and GZMB showed slight decreases. We cannot confirm global HERVK overexpression in ALS, but we can report the ALS-specific overexpression of selected HERVK copies in the ALS brain. Our data are compatible with the requirement for better patient stratification and support the potential importance of particular HERVK copies in ALS.

YY2 in Mouse Preimplantation Embryos and in Embryonic Stem Cells.

Yin Yang 2 encodes a mammalian-specific transcription factor (YY2) that shares high homology in the zinc finger region with both YY1 and REX1/ZFP42, encoded by the Yin Yang 1 and Reduced Expression Protein 1/Zinc Finger Protein 42 gene, respectively. In contrast to the well-established roles of the latter two in gene regulation, X chromosome inactivation and binding to specific transposable elements (TEs), much less is known about YY2, and its presence during mouse preimplantation development has not been described. As it has been reported that mouse embryonic stem cells (mESC) cannot be propagated in the absence of Yy2, the mechanistic understanding of how Yy2 contributes to mESC maintenance remains only very partially characterized. We describe Yy2 expression studies using RT-PCR and staining with a high-affinity polyclonal serum in mouse embryos and mESC. Although YY2 is expressed during preimplantation development, its presence appears dispensable for developmental progress in vitro until formation of the blastocyst. Attenuation of Yy2 levels failed to alter either Zscan4 levels in two-cell embryos or IAP and MERVL levels at later preimplantation stages. In contrast to previous claims that constitutively expressed shRNA against Yy2 in mESC prohibited the propagation of mESC in culture, we obtained colonies generated from mESC with attenuated Yy2 levels. Concomitant with a decreased number of undifferentiated colonies, Yy2-depleted mESC expressed higher levels of Zscan4 but no differences in the expression of TEs or other pluripotency markers including Sox2, Oct4, Nanog and Esrrb were observed. These results confirm the contribution of Yy2 to the maintenance of mouse embryonic stem cells and show the preimplantation expression of YY2. These functions are discussed in relation to mammalian-specific functions of YY1 and REX1.

Altered DNA methylation in human placenta after (suspected) preterm labor.

Aim: The aim of this study was to determine if alterations in DNA methylation in the human placenta would support suspected preterm labor as a pathologic insult associated with diminished placental health. Methods: We evaluated placental DNA methylation at seven loci differentially methylated in placental pathologies using targeted bisulfite sequencing, in placentas associated with preterm labor (term birth after suspected preterm labor [n = 15] and preterm birth [n = 15]), and controls (n = 15). Results: DNA methylation levels at the NCAM1 and PLAGL1 loci in placentas associated with preterm labor did differ significantly (p < 0.05) from controls. Discussion: Specific alterations in methylation patterns indicative of an unfavourable placental environment are associated with preterm labor per se and not restricted to preterm birth.

In Vivo Chromatin Targets of the Transcription Factor Yin Yang 2 in Trophoblast Stem Cells.

BACKGROUND: Yin Yang 2 (YY2) is a zinc finger protein closely related to the well-characterized Yin Yang 1 (YY1). YY1 is a DNA-binding transcription factor, with defined functions in multiple developmental processes, such as implantation, cell differentiation, X inactivation, imprinting and organogenesis. Yy2 has been treated as a largely immaterial duplication of Yy1, as they share high homology in the Zinc Finger-region and similar if not identical in vitro binding sites. In contrast to these similarities, gene expression alterations in HeLa cells with attenuated levels of either Yy1 or Yy2 were to some extent gene-specific. Moreover, the chromatin binding sites for YY2, except for its association with transposable retroviral elements (RE) and Endogenous Retroviral Elements (ERVs), remain to be identified. As a first step towards defining potential Yy2 functions matching or complementary to Yy1, we considered in vivo DNA binding sites of YY2 in trophoblast stem (TS) cells. RESULTS: We report the presence of YY2 protein in mouse-derived embryonic stem (ES) and TS cell lines. Following up on our previous report on ERV binding by YY2 in TS cells, we investigated the tissue-specificity of REX1 and YY2 binding and confirm binding to RE/ERV targets in both ES cells and TS cells. Because of the higher levels of expression, we chose TS cells to understand the role of Yy2 in gene and chromatin regulation. We used in vivo YY2 association as a measure to identify potential target genes. Sequencing of chromatin obtained in chromatin-immunoprecipitation (ChIP) assays carried out with αYY2 serum allowed us to identify a limited number of chromatin targets for YY2. Some putative binding sites were validated in regular ChIP assays and gene expression of genes nearby was altered in the absence of Yy2. CONCLUSIONS: YY2 binding to ERVs is not confined to TS cells. In vivo binding sites share the presence of a consensus binding motif. Selected sites were uniquely bound by YY2 as opposed to YY1, suggesting that YY2 exerts unique contributions to gene regulation. YY2 binding was not generally associated with gene promoters. However, several YY2 binding sites are linked to long noncoding RNA (lncRNA) genes and we show that the expression levels of a few of those are Yy2-dependent.