Human pericentromeric tandemly repeated DNA is transcribed at the end of oocyte maturation and is associated with membraneless mitochondria-associated structures

Abstract

Most of the human genome is non-coding. However, some of the non-coding part is transcriptionally active. In humans, the tandemly repeated (TR) pericentromeric non-coding DNA—human satellites 2 and 3 (HS2, HS3)—are transcribed in somatic cells. These transcripts are also found in pre- and post-implantation embryos. The aim of this study was to analyze HS2/HS3 transcription and cellular localization of transcripts in human maturating oocytes. The maternal HS2/HS3 TR transcripts transcribed from both strands were accumulated in the ooplasm in GV-MI oocytes as shown by DNA–RNA FISH (fluorescence in-situ hybridization). The transcripts’ content was higher in GV oocytes than in somatic cumulus cells according to real-time PCR. Using bioinformatics analysis, we demonstrated the presence of polyadenylated HS2 and HS3 RNAs in datasets of GV and MII oocyte transcriptomes. The transcripts shared a high degree of homology with HS2, HS3 transcripts previously observed in cancer cells. The HS2/HS3 transcripts were revealed by a combination of FISH and immunocytochemical staining within membraneless RNP structures that contained DEAD-box helicases DDX5 and DDX4. The RNP structures were closely associated with mitochondria, and are therefore similar to membraneless bodies described previously only in oogonia. These membraneless structures may be a site for spatial sequestration of RNAs and proteins in both maturating oocytes and cancer cells.

Human Satellite 1 (HSAT1) analysis provides novel evidence of pericentromeric transcription

“…3’RACE allowed us to perform a deeper characterization of HSAT1 transcripts by proving the accumulation of polyadenylated ncRNAs. Other reports show that both polyadenylated and non-polyadenylated pericentromeric transcripts can be detected in human (Dobrynin et al 2020). Polyadenylation of Pol II ncRNA transcripts can be tightly related with transcription termination, and intensively regulated in the presence of cellular stresses and/or cancer-associated mutations (Proudfoot 2016; Tian and Manley 2017).…”

“…Centromeric αSAT (Alpha Satellite) transcripts in particular have been considered vital for kinetochore stabilization and centromere cohesion (Liu et al 2015; McNulty et al 2017; Smurova and De Wulf 2018; Chen et al 2021). The progressive description of pericentromeric HSAT2 and HSAT3 transcripts has elevated their status to essential in several cellular contexts (Eymery et al 2009a), like the formation and regulation of heterochromatin (Saksouk et al 2015; Johnson et al 2017), aging (Enukashvily et al 2007; Mendez-Bermudez et al 2021), response to stress (Jolly et al 2004; Goenka et al 2016; Ninomiya et al 2020), differentiation (Yandim and Karakülah 2019; Dobrynin et al 2020), and cancer (Hall et al 2017; Nogalski and Shenk 2020; Chatterjee and Sengupta 2021).…”

Pericentromeric Non-Coding DNA Transcription Is Associated with Niche Impairment in Patients with Ineffective or Partially Effective Multiple Myeloma Treatment

“…HS sequences are tandemly repeated, therefore many HS primers amplify several sequences and thus are unsuitable for qPCR. We designed primers that amplify a 112 bp fragment of the transcript we described earlier [ 34 ] (see Supplementary Figure S2 in the cited reference), which was identical to a transcript (Acc No AY845701.1) that had been revealed in cancer cells by Valgardsdottir et al (2005) [ 35 ]. The amplification product appears as a single band of estimated length both in genomic DNA and polyT cDNA ( Figure 7 a).…”

“…mRNA expression levels were calculated by the 2 −ΔΔCt method with the levels of gene transcription normalized to the housekeeping genes GAPDH encoding glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Primers for HS2/HS3 amplification were targeted to the most abundant HS2/HS3 transcript we described earlier in oocyte transcriptomes [ 34 ], in senescent fibroblasts and some cancer lines [ 18 ], and which was also detected by [ 35 ]. The following HS2/HS3 primers were designed to amplify a 112 bp fragment at the 3′end of the transcript: ZhF, 5′-CGT TTC CTT TCG ATG GCG TT-3′ and ZhR, 5′-TGA AAT CCA ATA TGA TCA TCA TCG AA-3′.…”

Characterising genome architectures using genome decomposition analysis

“…However, it is clear that GDA can be run effectively on large genomes with resources commonly available on bioinformatics compute clusters, even including time-intensive repeat finding. In an analysis of the human genome with 50kbp windows, GDA clearly identified centromeres and pericentromeric repeat-rich regions known to be important features of chromosome architecture [ 27 ] (Sup Fig. 4 ).…”