Biology subjects

Garcia-Perez, J. L.

Publications and source records attributed to Garcia-Perez, J. L..

6 recordsLinked to original sources

bioRxiv2024

DGCR8 haploinsufficiency leads to primate-specific RNA dysregulation and pluripotency defects

The 22q11.2 deletion syndrome (22qDS) is caused by a microdeletion in chromosome 22, including DGCR8, an essential gene for miRNA production. The contribution of human DGCR8 hemizygosity to the disease is still unclear. In this study, we generated two human pluripotent cell models containing a single functional DGCR8 allele to elucidate its role on 22qDS. DGCR8+/-cells show increased apoptosis as well as self-renewal and differentiation defects in both the naive and primed states. The expression of primate-specific miRNAs was largely affected, due to impaired miRNA processing and chromatin accessibility. DGCR8+/- cells also displayed a pronounced reduction in human endogenous retrovirus class H (HERVH) expression, a primate-specific retroelement essential for pluripotency maintenance. Importantly, the reintroduction of primate-specific miRNAs as well as the miR-371-3 cluster rescued the cellular and molecular phenotypes of DGCR8+/-cells. Our results suggest that DGCR8 is haploinsufficient in humans and that miRNAs and transposable elements may have co-evolved in primates as part of an essential regulatory network to maintain stem cell identity.

molecular biology↗

bioRxiv2024

Variable patterns of retrotransposition in different HeLa strains provide mechanistic insights into SINE RNA mobilization processes

Alu elements are non-autonomous Short INterspersed Elements (SINEs) derived from the 7SL RNA gene that are present at over one million copies in human genomic DNA. Alu mobilizes by a mechanism known as retrotransposition, which requires the Long INterspersed Element-1 (LINE-1 or L1) ORF2-encoded protein (ORF2p). Here, we demonstrate that HeLa strains differ in their capacity to support Alu retrotransposition. Human Alu elements retrotranspose efficiently in HeLa-HA and HeLa-CCL2 (Alu-permissive) strains, but not in HeLa-JVM or HeLa-H1 (Alu-nonpermissive) strains. A similar pattern of retrotransposition was observed for other 7SL RNA-derived SINEs and tRNA-derived SINEs. In contrast, mammalian LINE-1s, a zebrafish LINE, a human SINE-VNTR-Alu (SVA) element, and an L1 ORF1-containing messenger RNA can retrotranspose in all four HeLa strains. Using an in vitro reverse transcriptase-based assay, we show that Alu RNAs associate with ORF2p and are converted into cDNAs in both Alu-permissive and Alu-nonpermissive HeLa strains, suggesting that 7SL- and tRNA-derived SINE RNAs use strategies to hijack' L1 ORF2p that are distinct from those used by SVA elements and ORF1-containing mRNAs. These data further suggest ORF2p associates with the Alu RNA poly(A) tract in both Alu-permissive and Alu-nonpermissive HeLa strains, but that Alu retrotransposition is blocked after this critical step in Alu-nonpermissive HeLa strains.

molecular biology↗

bioRxiv2023

Vitamin C activates young LINE-1 elements in mouse embryonic stem cells via H3K9me3 demethylation

Vitamin C (vitC) enhances the activity of 2-oxoglutarate-dependent dioxygenases, including TET enzymes, which catalyse DNA demethylation, and Jumonji-domain histone demethylases. The epigenetic remodelling promoted by vitC improves the efficiency of induced pluripotent stem cell derivation, and is required to attain a ground-state of pluripotency in embryonic stem cells (ESCs) that closely mimics the inner cell mass of the early blastocyst. However, genome-wide DNA and histone demethylation can lead to upregulation of transposable elements (TEs), and it is not known how vitC addition in culture media affects TE expression in pluripotent stem cells. Here we show that vitC increases the expression of evolutionarily young LINE-1 (L1) elements in mouse ESCs. We find that TET activity is dispensable for these effects, and that instead L1 upregulation occurs largely as a result of H3K9me3 loss mediated by KDM4A/C histone demethylases. Despite increased L1 levels, we did not detect increased somatic insertion rates in vitC-treated cells. Notably, treatment of human ESCs with vitC also increases L1 protein levels, which could impact the genetic and epigenetic stability of human pluripotent stem cells.

genetics↗

bioRxiv2019

LINE-1 retrotransposition impacts the genome of human pre implantation embryos and extraembryonic tissues

Long Interspersed Element 1 (LINE-1/L1) is an abundant retrotransposon that has greatly impacted human genome evolution. LINE-1s are responsible for the generation of millions of insertions in the current human population. The characterization of sporadic cases of mosaic individuals carrying pathogenic L1-insertions, suggest that heritable insertions occurs during early embryogenesis. However, the timing and potential genomic impact of LINE-1 mobilization during early embryogenesis is unknown. Here, we demonstrate that inner cell mass of human pre-implantation embryos support the expression and retrotransposition of LINE -1s. Additionally, we show that LINE-1s are expressed in trophectoderm cells of embryos, and identify placenta-restricted endogenous LINE-1 insertions in newborns. Using human embryonic stem cells as a model of post-implantation epiblast cells, we demonstrate ongoing LINE-1 retrotransposition, which can impact expression of targeted genes. Our data demonstrate that LINE-1 retrotransposition starts very shortly after fertilization and may represent a previously underappreciated factor in human biology and disease.

genomics↗

bioRxiv2018

The phylogenetically distinct early human embryo

Is the human early embryo unique in lacking an inner cell-mass (ICM) and having parallel development? We reanalyse single-cell transcriptomic data and stain human embryos in situ to reveal both classical step-wise development and a transcriptomically homologous ICM. This apparent classicism obscures phylogenetic singularity: unlike mice, human epiblast has self-renewal hallmarks and we have abundant blastocyst non-committed cells (NCCs), part of an apoptosis-mediated purging process. The transcriptomes of the pluripotent cells are fast evolving, in large part owing to endogenous retrovirus H (ERVH) activity, rendering all primate embryos unique. Each species is characterised by the ERVHs that are active and the neighbour genes whose expression are modulated. ERVH is associated with recent major gene expression gain and loss events of pluripotency{-}associated genes. Not least through lack of HERVH expression, the current portfolio of naive cultures, putative in vitro mimics of pluripotent cells, are both developmentally and phylogenetically "confused". O_LIAnalysis of single cell transcriptomics and in situ stainings uncover, and enable characterization of, human inner cell mass (ICM) C_LIO_LICell purging via apoptosis defines a phylogenetically restricted class of blastocyst non-committed cells (NCCs), whereas HERVs in conjunction with host defence mark the committed cells of ICM C_LIO_LIFast transcriptome evolution is particular to the pluripotent epiblast and is mostly due to the primate-specific transposable element, HERVH C_LIO_LICurrent naive cultures dont reflect human uniqueness being phylogenetically and developmentally "confused". C_LI

developmental biology↗

bioRxiv2017

Impact of non-LTR retrotransposons in the differentiation and evolution of Anatomically Modern Humans

Transposable Elements are biologically important components of eukaryote genomes. In particular, non-LTR retrotransposons (N-LTRrs) extensively shaped the human genome throughout evolution. In this study, we compared retrotransposon insertions differentially present in the genomes of Anatomically Modern Humans, Neanderthals, Denisovans and Chimpanzees, in order to assess the possible impact of retrotransposition in the differentiation of the human lineage. Briefly, we first identified species-specific N-LTRrs and established their distribution in present day human populations. These analyses shortlisted a group of N-LTRr insertions that were found exclusively in Anatomically Modern Humans. Notably, these insertions targeted genes more frequently than randomly expected and are associated with an increase in the number of transcriptional/splicing variants of those genes they inserted in. The analysis of the functionality of genes targeted by human-specific N-LTRr insertions seems to reflect phenotypic changes that occurred during human evolution. Furthermore, the expression of genes containing the most recent N-LTRr insertions is enriched in the brain, especially in undifferentiated neurons, and these genes associate in networks related to neuron maturation and migration. Additionally, we also identified candidate N-LTRr insertions that have likely produced new functional variants exclusive to modern humans, which show traces of positive selection and are now fixed in all present-day human populations. In sum, our results strongly suggest that N-LTRr impacted our differentiation as a species and have been a constant source of genomic variability all throughout the evolution of the human lineage.

genomics↗