Template Activating Factor-I α Regulates Retroviral Silencing during Reprogramming

Bui, Phuong Linh; Nishimura, Ken; Mondejar, Gonzalo Seminario; Kumar, Arun; Aizawa, Shiho; Murano, Kensaku; Nagata, Kyosuke; Hayashi, Yohei; Fukuda, Aya; Onuma, Yasuko; Ito, Yuzuru; Nakanishi, Mahito; Hisatake, Koji

doi:10.1016/j.celrep.2019.10.010

Cited by 9 publications

(8 citation statements)

References 56 publications

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“…SeVdp(dCNBR) is based upon a replication-defective and persistent Sendai virus vector (SeVdp), which expresses multiple exogenous genes at high levels [ 20 ]. SeVdp remains stable in the cytoplasm of infected cells and is free from the silencing that often occurs when a gene is expressed from a retrovirus-based vector in mESCs [ 21 , 22 ].…”

Section: Resultsmentioning

confidence: 99%

Locus-Specific Isolation of the Nanog Chromatin Identifies Regulators Relevant to Pluripotency of Mouse Embryonic Stem Cells and Reprogramming of Somatic Cells

Burramsetty

Nishimura

Kishimoto

et al. 2022

IJMS

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Pluripotency is a crucial feature of pluripotent stem cells, which are regulated by the core pluripotency network consisting of key transcription factors and signaling molecules. However, relatively less is known about the molecular mechanisms that modify the core pluripotency network. Here we used the CAPTURE (CRISPR Affinity Purification in situ of Regulatory Elements) to unbiasedly isolate proteins assembled on the Nanog promoter in mouse embryonic stem cells (mESCs), and then tested their functional relevance to the maintenance of mESCs and reprogramming of somatic cells. Gene ontology analysis revealed that the identified proteins, including many RNA-binding proteins (RBPs), are enriched in RNA-related functions and gene expression. ChIP-qPCR experiments confirmed that BCLAF1, FUBP1, MSH6, PARK7, PSIP1, and THRAP3 occupy the Nanog promoter region in mESCs. Knockdown experiments of these factors show that they play varying roles in self-renewal, pluripotency gene expression, and differentiation of mESCs as well as in the reprogramming of somatic cells. Our results show the utility of unbiased identification of chromatin-associated proteins on a pluripotency gene in mESCs and reveal the functional relevance of RBPs in ESC differentiation and somatic cell reprogramming.

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Section: Resultsmentioning

confidence: 99%

Locus-Specific Isolation of the Nanog Chromatin Identifies Regulators Relevant to Pluripotency of Mouse Embryonic Stem Cells and Reprogramming of Somatic Cells

Burramsetty

Nishimura

Kishimoto

et al. 2022

IJMS

Self Cite

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“…The copyright holder for this preprint this version posted March 16, 2021. ; https://doi.org/10.1101/2021.03.15.435479 doi: bioRxiv preprint chromatin architecture during the transition between pluripotency and differentiation (Bui et al 2019). While we see RB misregulation in Nrmt1 -/mice, and both RB and NRMT1 deficient mice share many similar phenotypes, it is likely other targets are contributing to Nrmt1 -/neural phenotypes and future scRNA-seq will help elucidate these targets as well.…”

Section: Discussionmentioning

confidence: 99%

“…SOX5 plays a role in regulating neuroblast migration and post-migratory differentiation, and its loss results in immature neuronal differentiation (Kwan et al 2008). The verified NRMT1 target SET has also been show to play a role in regulating the rearrangement of chromatin architecture during the transition between pluripotency and differentiation (Bui et al 2019). While we see RB misregulation in Nrmt1 -/mice, and both RB and NRMT1 deficient mice share many similar phenotypes, it is likely other targets are contributing to Nrmt1 -/neural phenotypes and future scRNA-seq will help elucidate these targets as well.…”

Section: Discussionmentioning

confidence: 99%

Age-related neurodegeneration and cognitive impairments of NRMT1 knockout mice are preceded by misregulation of RB and expansion of the neural stem cell population

Catlin

Rein

et al. 2021

Preprint

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N-terminal methylation is an important post-translational modification that regulates protein/DNA interactions and plays a role in many cellular processes, including DNA damage repair, mitosis, and transcriptional regulation. Our generation of a constitutive knockout mouse for the N-terminal methyltransferase NRMT1, demonstrated its loss results in severe developmental abnormalities and premature aging. As premature aging is often accompanied by neurodegeneration, we more specifically examined how NRMT1 loss affects neural pathology and cognitive behaviors. Here we find that Nrmt1-/- mice exhibit postnatal enlargement of the lateral ventricles, age-dependent striatal and hippocampal neurodegeneration, memory impairments, and hyperactivity. These morphological and behavior abnormalities are preceded by alterations in neural stem cell (NSC) development. Depletion of quiescent NSC pools in Nrmt1-/- mice is concurrent with expansion of intermediate progenitor and neuroblast pools. These phenotypes are similar to those seen with loss of the NRMT1 target retinoblastoma protein (RB), and we see that NRMT1 loss leads to derepression of RB target genes and abnormal RB phosphorylation and degradation. As also seen with RB loss, neurons in Nrmt1-/- mice fail to exit cell cycle and ultimately undergo NOXA-mediated apoptosis, indicating that early misregulation of RB in Nrmt1-/- mice promotes premature NSC proliferation and contributes to subsequent neurodegenerative phenotypes.

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“…The usefulness of synthetic genetic programs critically relies on the ability to robustly maintain and control the expression of transgenes. However, simple overexpression of transgenes in primary cells remains nontrivial, with large fractions of transduced cells silencing transgenes [107][108][109].…”

Section: Trends In Biotechnologymentioning

confidence: 99%

Synthetic gene circuits as tools for drug discovery

Beitz

Oakes

Galloway

2022

Trends in Biotechnology

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Within mammalian systems, there exists enormous opportunity to use synthetic gene circuits to enhance phenotype-based drug discovery, to map the molecular origins of disease, and to validate therapeutics in complex cellular systems. While drug discovery has relied on marker staining and high-content imaging in cell-based assays, synthetic gene circuits expand the potential for precision and speed. Here we present a vision of how circuits can improve the speed and accuracy of drug discovery by enhancing the efficiency of hit triage, capturing disease-relevant dynamics in cell-based assays, and simplifying validation and readouts from organoids and microphysiological systems (MPS). By tracking events and cellular states across multiple length and time scales, circuits will transform how we decipher the causal link between molecular events and phenotypes to improve the selectivity and sensitivity of cell-based assays. Enhancing phenotypic drug discovery with synthetic biologyDevelopment of new drugs requires identification and optimization of functional molecules that often proceeds sequentially through high-throughput discovery screening, hit validation, hit expansion and optimization, and preclinical disease modeling (see Glossary) (Figure 1, Key figure)[1]. Target-based drug discovery (TDD) programs aim to identify drug candidates that modulate a defined biological target that is hypothesized to play a causal role in initiating or sustaining a disease. TDD programs offer precision by directly screening for drug candidate efficacy using purified molecular targets [2]. By contrast, phenotypic drug discovery (PDD) programs aim to identify drug candidates that modulate a physiologically-relevant biological system or cellular signaling pathway and screen for drug efficacy using cell-based assays [2]. Cell-based assays are target-agnostic and provide the opportunity to identify therapeutics that resolve diseaseassociated phenotypic deficits even when the disease etiology remains obscure and targets are undefined. Thus, PDD has recently received renewed interest for its potential to identify first-in-class drugs that act via a novel mechanism of action (MoA) as well as identify drugs for disease with complex or unknown etiology [1][2][3][4]. In order to identify drug candidates that translate to effective therapeutics in the clinic, PDD programs require cell-based assays that faithfully recapitulate and report on disease-relevant processes. To efficiently and effectively screen in phenotypic models, assay development for initial discovery screens must balance throughput with the ability to capture these disease processes [1]. As PDD probes a larger biological space than TDD to identify potential therapeutics, the number of hits from an initial discovery screen in a PDD program may be very large [2]. Following initial discovery screens, hit validation and triage are required to produce a small number of high-confidence candidates to proceed to the resource-intensive stages of hit expansion, compound optimization, and hig...

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Template Activating Factor-I α Regulates Retroviral Silencing during Reprogramming

Cited by 9 publications

References 56 publications

Locus-Specific Isolation of the Nanog Chromatin Identifies Regulators Relevant to Pluripotency of Mouse Embryonic Stem Cells and Reprogramming of Somatic Cells

Locus-Specific Isolation of the Nanog Chromatin Identifies Regulators Relevant to Pluripotency of Mouse Embryonic Stem Cells and Reprogramming of Somatic Cells

Age-related neurodegeneration and cognitive impairments of NRMT1 knockout mice are preceded by misregulation of RB and expansion of the neural stem cell population

Synthetic gene circuits as tools for drug discovery

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