On transposons and totipotency

Abstract

Our perception of the role of the previously considered 'selfish' or 'junk' DNA has been dramatically altered in the past 20 years or so. A large proportion of this non-coding part of mammalian genomes is repetitive in nature, classified as either satellites or transposons. While repetitive elements can be termed selfish in terms of their amplification, such events have surely been co-opted by the host, suggesting by itself a likely altruistic function for the organism at the subject of such natural selection. Indeed numerous examples of transposons regulating the functional output of the host genome have been documented. Transposons provide a powerful framework for large-scale relatively rapid concerted regulatory activities with the ability to drive evolution. Mammalian totipotency has emerged as one key stage of development in which transposon-mediated regulation of gene expression has taken centre stage in the past few years. During this period, large-scale (epigenetic) reprogramming must be accomplished in order to activate the host genome. In mice and men, one particular element murine endogenous retrovirus with leucine tRNA primer (MERVL) (and its counterpart human ERVL (HERVL)) appears to have acquired roles as a key driving force in this process. Here, I will discuss and interpret the current knowledge and its implications regarding the role of transposons, particularly of long interspersed nuclear elements (LINE-1s) and endogenous retroviruses (ERVs), in the regulation of totipotency. This article is part of a discussion meeting issue 'Crossroads between transposons and gene regulation'.

Keywords: 2-cell-like cells; LINE-1; MERVL; pluripotency; reprogramming; transposable elements.

Figure 1.

Summary of the pathways shown so far to induce 2CLC in ESC cultures. Ectopic expression of the TF Dux and Dppa2/4 as well as culturing in hypoxia conditions or with TSA, increase the 2CLC population and therefore are positive regulators (green arrow). Downregulation of CAF-1, specific chromatin modifiers and components of the splicesosome also induces 2CLC, and therefore, these are negative regulators (red arrow). 2CLC emergence involves a transient DNA hypomethylation state, and a Zscan4-positive intermediate state. Global features of 2CLC are listed on the right.

Figure 2.

Experimental approaches to investigate potential totipotency. In (a,b), typical chimera assays are shown. In (a), incorporation into 8-cell stage pre-implantation mouse embryos is done by aggregation, typically referred to as ‘morula aggregation’. In this assay, contribution to lineage is based on 3D position, but should be complemented with co-immunostaining to establish at least partial molecular identity. EGFP, enhanced green fluorescent protein; FACS, fluorescence activated cell sorting; H2B, histone H2B; NLS, nuclear localization signal. In (b), incorporation is achieved through microinjection of cells into the blastocoel of early blastocysts, followed by implantation and analysis of the conceptus, typically at embryonic day (E) 9.5. In this assay, contribution is based on expression of a fluorescent reporter in the placenta. These are difficult experiments, often hindered by the fact that the placenta is highly autofluorescent, and often it is not straightforward to distinguish between placenta and the yolk sac, which is an ICM derivative. These analyses should be accompanied by a stringent analysis through sections and molecular analysis of markers from the trophoblastic derivatives of the placenta. scRNAseq, single cell RNA sequencing. In (c), cell culture strategies are shown. In (c), as suggested by Baker and Pera [53], the ability of a single cell to give rise to stem cells from the three lineages of the mature blastocyst is depicted. XEN cells, primitive endoderm-derived stem cells; TS cells, trophoblast stem cells, derived from the trophectoderm. Molecular analysis of each of these cell types for the known relevant markers should be performed. (d) A potential design to promote self-aggregation of 2CLC, and derivation of cyst-containing structures also referred to as blastocyst-like. As in (c), a molecular analysis and exploration of lineage markers should be performed. In (e,f), nuclear transfer (NT) strategies are shown. (e) The rationale behind using somatic cell nuclear transfer as an assay to test cellular plasticity, based on the observations that nuclei derived from early embryos show a highest success in generating embryos and pups upon cloning, as opposed to pluripotent stem cells. Accordingly, 2CLC nuclei show a higher success in producing clone embryos upon NT [31]. SCNT, somatic cell nuclear transfer. In (f), the schematic of an NT experiment, including the potential outcomes and implications, is shown.