scholarly journals Origin of a novel CYP20A1 transcript isoform through multiple Alu exaptations creates a potential miRNA sponge

2019 ◽  
Author(s):  
Aniket Bhattacharya ◽  
Vineet Jha ◽  
Khushboo Singhal ◽  
Mahar Fatima ◽  
Dayanidhi Singh ◽  
...  
Keyword(s):  
Heat Shock ◽  
Cortical Neurons ◽  
Regulatory Networks ◽  
Neuronal Development ◽  
Rna Seq ◽  
Orphan Gene ◽  
Mirna Sponge ◽  
Human Neurons ◽  
Single Nucleus ◽  
Conserved Gene

AbstractBackgroundPrimate-specific Alus contribute to transcriptional novelties in conserved gene regulatory networks. Alu RNAs are present at elevated levels in stress conditions and consequently leads to transcript isoform specific functional role modulating the physiological outcome. One of the possible mechanisms could be Alu nucleated mRNA-miRNA interplay.ResultUsing combination of bioinformatics and experiments, we report a transcript isoform of an orphan gene, CYP20A1 (CYP20A1_Alu-LT) through exaptation of 23 Alus in its 9kb 3’UTR. CYP20A1_Alu-LT, confirmed by 3’RACE, is an outlier in length and expressed in multiple cell lines. We demonstrate its presence in single nucleus RNA-seq of ∼16000 human cortical neurons (including rosehip neurons). Its expression is restricted to the higher primates. Most strikingly, miRanda predicts ∼4700 miRNA recognition elements (MREs; with threshold< −25kcal/mol) for ∼1000 miRNAs, which have majorly originated within the 3’UTR-Alus post exaptation. We hypothesized that differential expression of this transcript could modulate mRNA-miRNA networks and tested it in primary human neurons where CYP20A1_Alu-LT is downregulated during heat shock response and upregulated upon HIV1-Tat treatment. CYP20A1_Alu-LT could possibly function as a miRNA sponge as it exhibits features of a sponge RNA such as cytosolic localization and ≥10 MREs for 140 miRNAs. Small RNA-seq revealed expression of nine miRNAs that can potentially be sponged by CYP20A1_Alu-LT in neurons. Additionally, CYP20A1_Alu-LT expression was positively correlated (low in heat shock and high in Tat) with 380 differentially expressed genes that contain cognate MREs for these nine miRNAs. This set is enriched in genes involved in neuronal development and hemostasis pathways.ConclusionWe demonstrate a potential role for CYP20A1_Alu-LT as miRNA sponge through preferential presence of MREs within Alus in a transcript isoform specific manner. This highlights a novel component of Alu-miRNA mediated transcriptional modulation leading to physiological homeostasis.

scholarly journals Origin of a novel CYP20A1 transcript isoform through multiple Alu exaptations creates a potential miRNA sponge

2020 ◽  
Author(s):  
Aniket Bhattacharya ◽  
Vineet Jha ◽  
Khushboo Singhal ◽  
Mahar Fatima ◽  
Dayanidhi Singh ◽  
...  
Keyword(s):  
Heat Shock ◽  
Cortical Neurons ◽  
Regulatory Networks ◽  
Neuronal Development ◽  
Rna Seq ◽  
Orphan Gene ◽  
Mirna Sponge ◽  
Human Neurons ◽  
Single Nucleus ◽  
Conserved Gene

Abstract Background: Primate-specific Alus contribute to transcriptional novelties in conserved gene regulatory networks. Alu RNAs are present at elevated levels in stress conditions and consequently leads to transcript isoform specific functional role modulating the physiological outcome. One of the possible mechanisms could be Alu nucleated mRNA-miRNA interplay. Result: Using combination of bioinformatics and experiments, we report a transcript isoform of an orphan gene, CYP20A1 ( CYP20A1_Alu-LT ) through exaptation of 23 Alus in its 9kb 3’UTR. CYP20A1_Alu-LT , confirmed by 3’RACE, is an outlier in length and expressed in multiple cell lines. We demonstrate its presence in single nucleus RNA-seq of ~16000 human cortical neurons (including rosehip neurons). Its expression is restricted to the higher primates. Most strikingly, miRanda predicts ~4700 miRNA recognition elements (MREs; with threshold< -25kcal/mol) for ~1000 miRNAs, which have majorly originated within the 3’UTR-Alus post exaptation. We hypothesized that differential expression of this transcript could modulate mRNA-miRNA networks and tested it in primary human neurons where CYP20A1_Alu-LT is downregulated during heat shock response and upregulated upon HIV1-Tat treatment. CYP20A1_Alu-LT could possibly function as a miRNA sponge as it exhibits features of a sponge RNA such as cytosolic localization and ≥10 MREs for 140 miRNAs. Small RNA-seq revealed expression of nine miRNAs that can potentially be sponged by CYP20A1_Alu-LT in neurons. Additionally, CYP20A1_Alu-LT expression was positively correlated (low in heat shock and high in Tat) with 380 differentially expressed genes that contain cognate MREs for these nine miRNAs. This set is enriched in genes involved in neuronal development and hemostasis pathways. Conclusion: We demonstrate a potential role for CYP20A1_Alu-LT as miRNA sponge through preferential presence of MREs within Alus in a transcript isoform specific manner. This highlights a novel component of Alu-miRNA mediated transcriptional modulation leading to physiological homeostasis.

scholarly journals Multiple Alu exonization in 3’UTR of a primate specific isoform of CYP20A1 creates a potential miRNA sponge

2020 ◽  
Author(s):  
Aniket Bhattacharya ◽  
Vineet Jha ◽  
Khushboo Singhal ◽  
Mahar Fatima ◽  
Dayanidhi Singh ◽  
...  
Keyword(s):  
Heat Shock ◽  
Cortical Neurons ◽  
Regulatory Networks ◽  
Large Scale ◽  
Neuronal Development ◽  
Random Sets ◽  
Rna Seq ◽  
Orphan Gene ◽  
Mirna Sponge ◽  
Human Neurons

Abstract Alu repeats contribute to phylogenetic novelties in conserved regulatory networks in primates. Our study highlights how exonized Alus could nucleate large-scale mRNA-miRNA interactions. Using a functional genomics approach, we characterize a transcript isoform of an orphan gene, CYP20A1 (CYP20A1_Alu-LT) that has exonization of 23 Alus in its 3’UTR. CYP20A1_Alu-LT, confirmed by 3’RACE, is an outlier in length (9 kb 3’UTR) and widely expressed. Using publically available datasets, we demonstrate its expression in higher primates and presence in single nucleus RNA-seq of 15928 human cortical neurons. miRanda predicts ∼4700 miRNA recognition elements (MREs) for ∼1000 miRNAs, primarily originated within these 3’UTR-Alus. CYP20A1_Alu-LT could be a potential multi-miRNA sponge as it harbors ≥10 MREs for 140 miRNAs and has cytosolic localization. We further tested whether expression of CYP20A1_Alu-LT correlates with mRNAs harboring similar MRE targets. RNA-seq with conjoint miRNA-seq analysis was done in primary human neurons where we observed CYP20A1_Alu-LT to be downregulated during heat shock response and upregulated in HIV1-Tat treatment. 380 genes were positively correlated with its expression (significantly downregulated in heat shock and upregulated in Tat) and they harbored MREs for nine expressed miRNAs which were also enriched in CYP20A1_Alu-LT. MREs were significantly enriched in these 380 genes compared to random sets of differentially expressed genes (p = 8.134e-12). Gene ontology suggested involvement of these genes in neuronal development and hemostasis pathways thus proposing a novel component of Alu-miRNA mediated transcriptional modulation that could govern specific physiological outcomes in higher primates.

scholarly journals Multiple Alu exonizations in 3’UTR of a primate specific isoform of CYP20A1 creates a potential miRNA sponge

2020 ◽  
Author(s):  
Aniket Bhattacharya ◽  
Vineet Jha ◽  
Khushboo Singhal ◽  
Mahar Fatima ◽  
Dayanidhi Singh ◽  
...  
Keyword(s):  
Heat Shock ◽  
Cortical Neurons ◽  
Regulatory Networks ◽  
Large Scale ◽  
Neuronal Development ◽  
Random Sets ◽  
Orphan Gene ◽  
Mirna Sponge ◽  
Human Neurons ◽  
Physiological Outcomes

Abstract Background: Alu repeats contribute to phylogenetic novelties in conserved regulatory networks in primates. Exaptation of Alus in transcript isoforms could nucleate large-scale mRNA-miRNA interactions and modulate cellular outcomes. Result: Using a functional genomics approach, we report a transcript isoform of an orphan gene, CYP20A1 (CYP20A1_Alu-LT) that arise through exonization of 23 Alus in 3’UTR and is expressed in higher primates. CYP20A1_Alu-LT, confirmed by 3’RACE, is an outlier in length (9kb) and is expressed in multiple cell lines. Using publicly available datasets, we demonstrate its presence in single nucleus RNA-seq of 15928 human cortical neurons (including rosehip neurons). miRanda predicts ~4700 miRNA recognition elements (MREs; with threshold< -25kcal/mol) for ~1000 miRNAs, which have primarily originated within the 3’UTR-Alus post exonization. CYP20A1_Alu-LT could be a potential multi-miRNA sponge as it harbours ≥10 MREs for 140 miRNAs and has cytosolic localization. In order to test this further, we explored whether expression of CYP20A1_Alu-LT correlates with genome wide mRNAs harboring similar MRE targets. We carried out RNAseq with conjoint miRNA-seq analysis in primary human neurons as we observed CYP20A1_Alu-LT to be downregulated during heat shock response and upregulated in HIV1-Tat treatment. CYP20A1_Alu-LT expression was positively correlated with 380 genes that were significantly downregulated in heat shock and upregulated in Tat and harboured MREs for a set of nine expressed miRNAs that were also enriched in CYP20A1_Alu-LT. The enrichment of MREs in the 380 genes were significant compared to random sets of expressed (p=4.716e-12) as well as differentially expressed genes (p=8.134e-12). Gene ontology suggested involvement of these genes in neuronal development and hemostasis pathways. Conclusion: Our study suggests a potential role for CYP20A1_Alu-LT as miRNA sponge due to significant enrichment of MREs within Alus in a transcript isoform specific manner. This highlights a novel component of Alu-miRNA mediated transcriptional modulation that could govern specific physiological outcomes in higher primates.

scholarly journals Multiple Alu exonizations in 3’UTR of a primate specific isoform of CYP20A1 creates a potential miRNA sponge

2020 ◽  
Author(s):  
Aniket Bhattacharya ◽  
Vineet Jha ◽  
Khushboo Singhal ◽  
Mahar Fatima ◽  
Dayanidhi Singh ◽  
...  
Keyword(s):  
Heat Shock ◽  
Cortical Neurons ◽  
Regulatory Networks ◽  
Large Scale ◽  
Neuronal Development ◽  
Random Sets ◽  
Orphan Gene ◽  
Mirna Sponge ◽  
Human Neurons ◽  
Physiological Outcomes

Abstract Background: Alu repeats contribute to phylogenetic novelties in conserved regulatory networks in primates. Exaptation of Alus in transcript isoforms could nucleate large-scale mRNA-miRNA interactions and modulate cellular outcomes. Result: Using a functional genomics approach, we report a transcript isoform of an orphan gene, CYP20A1 (CYP20A1_Alu-LT) that arise through exonization of 23 Alus in 3’UTR and is expressed in higher primates. CYP20A1_Alu-LT, confirmed by 3’RACE, is an outlier in length (9kb) and is expressed in multiple cell lines. We demonstrate its presence in single nucleus RNA-seq of ~16000 human cortical neurons (including rosehip neurons). Most strikingly, miRanda predicts ~4700 miRNA recognition elements (MREs; with threshold< -25kcal/mol) for ~1000 miRNAs, which have primarily originated within the 3’UTR-Alus post exonization. CYP20A1_Alu-LT could be a potential multi- miRNA sponge as it harbours - ≥10 MREs for 140 miRNAs and has cytosolic localization. In order to test this further, we explored whether expression of CYP20A1_Alu-LT correlates with genome wide mRNAs harboring similar MRE targets. We carried out RNAseq with conjoint miRNAseq analysis in primary human neurons as we observed CYP20A1_Alu-LT to be downregulated during heat shock response and upregulated in HIV1-Tat treatment. CYP20A1_Alu-LT expression was positively correlated with 380 genes that were significantly downregulated in heat shock and upregulated in Tat and harboured MREs for a set of nine expressed miRNAs that were also enriched in CYP20A1_Alu-LT. The enrichment of MREs in the 380 genes were significant compared to random sets of expressed (p=4.716e-12) as well as differentially expressed genes (p=8.134e-12). Gene ontology revealed involvement of these genes in neuronal development and hemostasis pathways. Conclusion: We demonstrate a potential role for CYP20A1_Alu-LT as miRNA sponge due to significant enrichment of MREs within Alus in a transcript isoform specific manner. This highlights a novel component of Alu-miRNA mediated transcriptional modulation that could govern specific physiological outcomes in higher primates.

scholarly journals Species-specific mitochondria dynamics and metabolism regulate the timing of neuronal development.

2021 ◽  
Author(s):  
Ryohei Iwata ◽  
Pierre Casimir ◽  
Emir Erkol ◽  
Leila Boubakar ◽  
Melanie Planque ◽  
...  
Keyword(s):  
Cortical Neurons ◽  
Neuronal Development ◽  
Brain Evolution ◽  
Complex Cells ◽  
Human Neurons ◽  
Mitochondria Dynamics ◽  
Species Specific ◽  
Human And Mouse ◽  
Human Specific ◽  
Stimulation Of

The evolution of species involves changes in the timeline of key developmental programs. Among these, neuronal development is considerably prolonged in the human cerebral cortex compared with other mammals, leading to brain neoteny. Here we explore whether mitochondria influence the species-specific properties of cortical neuron maturation. By comparing human and mouse cortical neuronal maturation at high temporal and cell resolution, we found a slower pattern of mitochondria development in human cortical neurons compared with the mouse, together with lower mitochondria metabolic activity, particularly oxidative phosphorylation. Stimulation of mitochondria metabolism in human neurons resulted in accelerated maturation, leading to excitable and complex cells weeks ahead of time. Our data identify mitochondria as important regulators of the pace of neuronal development underlying human-specific features of brain evolution.

scholarly journals EPEN-21. IMPAIRED NEURONAL-GLIAL FATE SPECIFICATION IN PEDIATRIC EPENDYMOMA REVEALED BY SINGLE-CELL RNA-SEQ

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii311-iii312
Author(s):  
Bernhard Englinger ◽  
Johannes Gojo ◽  
Li Jiang ◽  
Jens M Hübner ◽  
McKenzie L Shaw ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Single Cell ◽  
Regulatory Networks ◽  
Target Genes ◽  
Target Identification ◽  
Rna Seq ◽  
Cell Models ◽  
Pediatric Ependymoma ◽  
Glial Fate ◽  
Anatomic Locations

Abstract Ependymoma represents a heterogeneous disease affecting the entire neuraxis. Extensive molecular profiling efforts have identified molecular ependymoma subgroups based on DNA methylation. However, the intratumoral heterogeneity and developmental origins of these groups are only partially understood, and effective treatments are still lacking for about 50% of patients with high-risk tumors. We interrogated the cellular architecture of ependymoma using single cell/nucleus RNA-sequencing to analyze 24 tumor specimens across major molecular subgroups and anatomic locations. We additionally analyzed ten patient-derived ependymoma cell models and two patient-derived xenografts (PDXs). Interestingly, we identified an analogous cellular hierarchy across all ependymoma groups, originating from undifferentiated neural stem cell-like populations towards different degrees of impaired differentiation states comprising neuronal precursor-like, astro-glial-like, and ependymal-like tumor cells. While prognostically favorable ependymoma groups predominantly harbored differentiated cell populations, aggressive groups were enriched for undifferentiated subpopulations. Projection of transcriptomic signatures onto an independent bulk RNA-seq cohort stratified patient survival even within known molecular groups, thus refining the prognostic power of DNA methylation-based profiling. Furthermore, we identified novel potentially druggable targets including IGF- and FGF-signaling within poorly prognostic transcriptional programs. Ependymoma-derived cell models/PDXs widely recapitulated the transcriptional programs identified within fresh tumors and are leveraged to validate identified target genes in functional follow-up analyses. Taken together, our analyses reveal a developmental hierarchy and transcriptomic context underlying the biologically and clinically distinct behavior of ependymoma groups. The newly characterized cellular states and underlying regulatory networks could serve as basis for future therapeutic target identification and reveal biomarkers for clinical trials.

scholarly journals Methyltransferase-directed orthogonal tagging and sequencing of miRNAs and bacterial small RNAs

BMC Biology ◽  
2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Milda Mickutė ◽  
Kotryna Kvederavičiūtė ◽  
Aleksandr Osipenko ◽  
Raminta Mineikaitė ◽  
Saulius Klimašauskas ◽  
...  
Keyword(s):  
Rna Sequencing ◽  
Regulatory Networks ◽  
Library Preparation ◽  
Rna Seq ◽  
Basic Principles ◽  
Cofactor Binding ◽  
Sequencing Library ◽  
Sequencing Library Preparation ◽  
Target Rna

Abstract Background Targeted installation of designer chemical moieties on biopolymers provides an orthogonal means for their visualisation, manipulation and sequence analysis. Although high-throughput RNA sequencing is a widely used method for transcriptome analysis, certain steps, such as 3′ adapter ligation in strand-specific RNA sequencing, remain challenging due to structure- and sequence-related biases introduced by RNA ligases, leading to misrepresentation of particular RNA species. Here, we remedy this limitation by adapting two RNA 2′-O-methyltransferases from the Hen1 family for orthogonal chemo-enzymatic click tethering of a 3′ sequencing adapter that supports cDNA production by reverse transcription of the tagged RNA. Results We showed that the ssRNA-specific DmHen1 and dsRNA-specific AtHEN1 can be used to efficiently append an oligonucleotide adapter to the 3′ end of target RNA for sequencing library preparation. Using this new chemo-enzymatic approach, we identified miRNAs and prokaryotic small non-coding sRNAs in probiotic Lactobacillus casei BL23. We found that compared to a reference conventional RNA library preparation, methyltransferase-Directed Orthogonal Tagging and RNA sequencing, mDOT-seq, avoids misdetection of unspecific highly-structured RNA species, thus providing better accuracy in identifying the groups of transcripts analysed. Our results suggest that mDOT-seq has the potential to advance analysis of eukaryotic and prokaryotic ssRNAs. Conclusions Our findings provide a valuable resource for studies of the RNA-centred regulatory networks in Lactobacilli and pave the way to developing novel transcriptome and epitranscriptome profiling approaches in vitro and inside living cells. As RNA methyltransferases share the structure of the AdoMet-binding domain and several specific cofactor binding features, the basic principles of our approach could be easily translated to other AdoMet-dependent enzymes for the development of modification-specific RNA-seq techniques.

scholarly journals Gene regulatory networks controlling differentiation, survival, and diversification of hypothalamic Lhx6-expressing GABAergic neurons

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Dong Won Kim ◽  
Kai Liu ◽  
Zoe Qianyi Wang ◽  
Yi Stephanie Zhang ◽  
Abhijith Bathini ◽  
...  
Keyword(s):  
Cortical Neurons ◽  
Regulatory Networks ◽  
Molecular Mechanisms ◽  
Gabaergic Neurons ◽  
Zona Incerta ◽  
Long Distance ◽  
Hypothalamic Neurons ◽  
Cortical Interneuron ◽  
Tangential Migration ◽  
Innate Behaviors

AbstractGABAergic neurons of the hypothalamus regulate many innate behaviors, but little is known about the mechanisms that control their development. We previously identified hypothalamic neurons that express the LIM homeodomain transcription factor Lhx6, a master regulator of cortical interneuron development, as sleep-promoting. In contrast to telencephalic interneurons, hypothalamic Lhx6 neurons do not undergo long-distance tangential migration and do not express cortical interneuronal markers such as Pvalb. Here, we show that Lhx6 is necessary for the survival of hypothalamic neurons. Dlx1/2, Nkx2-2, and Nkx2-1 are each required for specification of spatially distinct subsets of hypothalamic Lhx6 neurons, and that Nkx2-2+/Lhx6+ neurons of the zona incerta are responsive to sleep pressure. We further identify multiple neuropeptides that are enriched in spatially segregated subsets of hypothalamic Lhx6 neurons, and that are distinct from those seen in cortical neurons. These findings identify common and divergent molecular mechanisms by which Lhx6 controls the development of GABAergic neurons in the hypothalamus.

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