Publications by authors named "Gokul N Ramadoss"

5 Publications

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Genome-wide programmable transcriptional memory by CRISPR-based epigenome editing.

Cell 2021 Apr 9;184(9):2503-2519.e17. Epub 2021 Apr 9.

Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, CA 94158, USA; Whitehead Institute for Biomedical Research, Massachusetts Institute of Technology, Cambridge 02142, USA. Electronic address:

A general approach for heritably altering gene expression has the potential to enable many discovery and therapeutic efforts. Here, we present CRISPRoff-a programmable epigenetic memory writer consisting of a single dead Cas9 fusion protein that establishes DNA methylation and repressive histone modifications. Transient CRISPRoff expression initiates highly specific DNA methylation and gene repression that is maintained through cell division and differentiation of stem cells to neurons. Pairing CRISPRoff with genome-wide screens and analysis of chromatin marks establishes rules for heritable gene silencing. We identify single guide RNAs (sgRNAs) capable of silencing the large majority of genes including those lacking canonical CpG islands (CGIs) and reveal a wide targeting window extending beyond annotated CGIs. The broad ability of CRISPRoff to initiate heritable gene silencing even outside of CGIs expands the canonical model of methylation-based silencing and enables diverse applications including genome-wide screens, multiplexed cell engineering, enhancer silencing, and mechanistic exploration of epigenetic inheritance.
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http://dx.doi.org/10.1016/j.cell.2021.03.025DOI Listing
April 2021

SARS-CoV-2 infection of human iPSC-derived cardiac cells reflects cytopathic features in hearts of patients with COVID-19.

Sci Transl Med 2021 04 15;13(590). Epub 2021 Mar 15.

Gladstone Institutes, San Francisco, CA 94158, USA.

Although coronavirus disease 2019 (COVID-19) causes cardiac dysfunction in up to 25% of patients, its pathogenesis remains unclear. Exposure of human induced pluripotent stem cell (iPSC)-derived heart cells to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) revealed productive infection and robust transcriptomic and morphological signatures of damage, particularly in cardiomyocytes. Transcriptomic disruption of structural genes corroborates adverse morphologic features, which included a distinct pattern of myofibrillar fragmentation and nuclear disruption. Human autopsy specimens from patients with COVID-19 reflected similar alterations, particularly sarcomeric fragmentation. These notable cytopathic features in cardiomyocytes provide insights into SARS-CoV-2-induced cardiac damage, offer a platform for discovery of potential therapeutics, and raise concerns about the long-term consequences of COVID-19 in asymptomatic and severe cases.
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http://dx.doi.org/10.1126/scitranslmed.abf7872DOI Listing
April 2021

BRD2 inhibition blocks SARS-CoV-2 infection in vitro by reducing transcription of the host cell receptor ACE2.

bioRxiv 2021 Jan 19. Epub 2021 Jan 19.

SARS-CoV-2 infection of human cells is initiated by the binding of the viral Spike protein to its cell-surface receptor ACE2. We conducted an unbiased CRISPRi screen to uncover druggable pathways controlling Spike protein binding to human cells. We found that the protein BRD2 is an essential node in the cellular response to SARS-CoV-2 infection. BRD2 is required for ACE2 transcription in human lung epithelial cells and cardiomyocytes, and BRD2 inhibitors currently evaluated in clinical trials potently block endogenous ACE2 expression and SARS-CoV-2 infection of human cells. BRD2 also controls transcription of several other genes induced upon SARS-CoV-2 infection, including the interferon response, which in turn regulates ACE2 levels. It is possible that the previously reported interaction between the viral E protein and BRD2 evolved to manipulate the transcriptional host response during SARS-CoV-2 infection. Together, our results pinpoint BRD2 as a potent and essential regulator of the host response to SARS-CoV-2 infection and highlight the potential of BRD2 as a novel therapeutic target for COVID-19.
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http://dx.doi.org/10.1101/2021.01.19.427194DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7836110PMC
January 2021

SARS-CoV-2 infection of human iPSC-derived cardiac cells predicts novel cytopathic features in hearts of COVID-19 patients.

bioRxiv 2020 Sep 12. Epub 2020 Sep 12.

Gladstone Institutes, San Francisco, CA.

Although COVID-19 causes cardiac dysfunction in up to 25% of patients, its pathogenesis remains unclear. Exposure of human iPSC-derived heart cells to SARS-CoV-2 revealed productive infection and robust transcriptomic and morphological signatures of damage, particularly in cardiomyocytes. Transcriptomic disruption of structural proteins corroborated adverse morphologic features, which included a distinct pattern of myofibrillar fragmentation and numerous iPSC-cardiomyocytes lacking nuclear DNA. Human autopsy specimens from COVID-19 patients displayed similar sarcomeric disruption, as well as cardiomyocytes without DNA staining. These striking cytopathic features provide new insights into SARS-CoV-2 induced cardiac damage, offer a platform for discovery of potential therapeutics, and raise serious concerns about the long-term consequences of COVID-19.
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http://dx.doi.org/10.1101/2020.08.25.265561DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7491510PMC
September 2020

hPSC-Derived Striatal Cells Generated Using a Scalable 3D Hydrogel Promote Recovery in a Huntington Disease Mouse Model.

Stem Cell Reports 2018 05 5;10(5):1481-1491. Epub 2018 Apr 5.

Department of Chemical and Biomolecular Engineering, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Bioengineering, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; The Helen Wills Neuroscience Institute, University of California, Berkeley, Berkeley, CA 94720, USA. Electronic address:

Huntington disease (HD) is an inherited, progressive neurological disorder characterized by degenerating striatal medium spiny neurons (MSNs). One promising approach for treating HD is cell replacement therapy, where lost cells are replaced by MSN progenitors derived from human pluripotent stem cells (hPSCs). While there has been remarkable progress in generating hPSC-derived MSNs, current production methods rely on two-dimensional culture systems that can include poorly defined components, limit scalability, and yield differing preclinical results. To facilitate clinical translation, here, we generated striatal progenitors from hPSCs within a fully defined and scalable PNIPAAm-PEG three-dimensional (3D) hydrogel. Transplantation of 3D-derived striatal progenitors into a transgenic mouse model of HD slowed disease progression, improved motor coordination, and increased survival. In addition, the transplanted cells developed an MSN-like phenotype and formed synaptic connections with host cells. Our results illustrate the potential of scalable 3D biomaterials for generating striatal progenitors for HD cell therapy.
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http://dx.doi.org/10.1016/j.stemcr.2018.03.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5995679PMC
May 2018