Paired CRISPR screens to map gene regulation in and .

Recent massively-parallel approaches to decipher gene regulatory circuits have focused on the discovery of either -regulatory elements (CREs) or -acting factors. Here, we develop a scalable approach that pairs - and -regulatory CRISPR screens to systematically dissect how the key immune checkpoint is regulated. In human pancreatic ductal adenocarcinoma (PDAC) cells, we tile the locus using ∼25,000 CRISPR perturbations in constitutive and IFNγ-stimulated conditions.

Transcriptome-scale RNA-targeting CRISPR screens reveal essential lncRNAs in human cells.

Mammalian genomes host a diverse array of RNA that includes protein-coding and noncoding transcripts. However, the functional roles of most long noncoding RNAs (lncRNAs) remain elusive. Using RNA-targeting CRISPR-Cas13 screens, we probed how the loss of ∼6,200 lncRNAs impacts cell fitness across five human cell lines and identified 778 lncRNAs with context-specific or broad essentiality.

Anomalous coarsening of coalescing nucleoli in human cells.

Coarsening is a ubiquitous phenomenon in droplet systems near thermodynamic equilibrium-as an increase in droplet size lowers the system's free energy-however, coarsening of droplets in nonequilibrium systems, such as the cell nucleus, is far from understood. Liquid condensates in the cell nucleus, like nucleoli, form by liquid-liquid phase separation and play a key role in the nuclear organization. In human cells, nucleolar droplets are nucleated at the beginning of the cell cycle and coarsen with time by coalescing with each other.

CRISPR in cancer biology and therapy.

Over the past decade, CRISPR has become as much a verb as it is an acronym, transforming biomedical research and providing entirely new approaches for dissecting all facets of cell biology. In cancer research, CRISPR and related tools have offered a window into previously intractable problems in our understanding of cancer genetics, the noncoding genome and tumour heterogeneity, and provided new insights into therapeutic vulnerabilities. Here, we review the progress made in the development of CRISPR systems as a tool to study cancer, and the emerging adaptation of these technologies to improve diagnosis and treatment.

Mechanical stress affects dynamics and rheology of the human genome.

Material properties of the genome are critical for proper cellular function - they directly affect timescales and length scales of DNA transactions such as transcription, replication and DNA repair, which in turn impact all cellular processes the central dogma of molecular biology. Hence, elucidating the genome's rheology may help reveal physical principles underlying the genome's organization and function. Here, we present a novel noninvasive approach to study the genome's rheology and its response to mechanical stress in form of nuclear injection in live human cells.

Nucleolar dynamics and interactions with nucleoplasm in living cells.

Liquid-liquid phase separation (LLPS) has been recognized as one of the key cellular organizing principles and was shown to be responsible for formation of membrane-less organelles such as nucleoli. Although nucleoli were found to behave like liquid droplets, many ramifications of LLPS including nucleolar dynamics and interactions with the surrounding liquid remain to be revealed. Here, we study the motion of human nucleoli , while monitoring the shape of the nucleolus-nucleoplasm interface.

Surface Fluctuations and Coalescence of Nucleolar Droplets in the Human Cell Nucleus.

The nucleolus is a membraneless organelle embedded in chromatin solution inside the cell nucleus. By analyzing surface dynamics and fusion kinetics of human nucleoli in vivo, we find that the nucleolar surface exhibits subtle, but measurable, shape fluctuations and that the radius of the neck connecting two fusing nucleoli grows in time as r(t)∼t^{1/2}. This is consistent with liquid droplets with low surface tension ∼10^{-6}  N m^{-1} coalescing within an outside fluid of high viscosity ∼10^{3}  Pa s.