Gaublomme Lab shares CRISPRmap in Nature Biotechnology

In an article published last week in Nature Biotechnology, graduate student Jiacheng Gu, along with other members Gaublomme lab, shared CRISPRmap, a novel optical CRISPR screening approach that maps optical properties of single cells to targeted genetic perturbations. In other words, the approach takes advantage of how cells interact with light to reveal how they respond to changes that researchers introduce at the genetic level. By observing how disabling or modifying different genes affects the cells, researchers can identify previously unknown functions of specific genes, including those implicated in a wide array of diseases.

In collaboration with the Ciccia lab at Columbia University Irving Medical Center, the new study used CRISPRmap to explore the effects of genetic variants of unknown significance (VUSs) on the DNA damage response in breast cancer. Specifically, the researchers explored the functional consequences of 292 single nucleotide variants in 27 DNA damage response genes. After treating the cells with ionizing radiation, or DNA-damaging agents commonly used as chemotherapeutic drugs in breast cancer, the researchers visualized the subcellular localization of dozens of proteins and mRNA species in approximately one million cells. "This method allowed us to identify missense variants of uncertain clinical significance whose response resembles known pathogenic variants,” says Gu. “As such, our approach can provide a framework for annotating human variants in a treatment-specific manner and help prioritize therapeutic strategies." For breast cancer patients, a better understanding of these variants and their drug responses could lead to better outcomes.  

CRISPRmap offers solutions to two significant problems. First, while pooled CRISPR screens that introduce genetic perturbations into many cells at once have empowered scientists to analyze gene function with impressively high throughput, such screens rely on next-generation sequencing—a technology used to determine the order of nucleotides in entire genomes, or in targeted regions of DNA or RNA—to evaluate the effects. Next-generation sequencing based screens typically require isolating and destroying cells, or cell lysis, which means screens can't capture how cells are organized in space, especially in tissue contexts. As an optical approach CRISPRmap, however, doesn’t rely on cell lysis, enabling the study how proteins and cells behave and alter location. It combines optical barcode detection, which enables us to know which gene got altered in any given cell, with visualization of numerous proteins and RNA transcripts to quantify the cells response to the alteration. The efficiency of barcode readout enabled for the first time to perform optical CRISPR screens in challenging cell types and tissue contexts. Collaborating with the Chan lab, Gu and other members Gaublomme lab profiled tumor sections with CRISPRmap to visualize angiogenesis (the development of blood vessels that feed tumors) extracellular matrix formation around tumor domains (a network of macromolecules that provide structural and biochemical support to cells and plays a critical role in the development and progression of cancer), and transcription factor nuclear translocation in the transplanted cells. Dr. Gaublomme is enthusiastic about the new approach. “Tissue based perturbations studies enable mapping of cell-extrinsic effects that cannot be captured by sequencing-based methods based on tissue dissociation and cell lysis.”

Second, Gu and his collaborators optimized their approach for broad accessibility, beyond even the broad application of CRISPRmap in the context of cells and tissue. The method does not rely on third-party sequencing reagents for barcode detection, and the readout dyes can be customized to match microscopes available to researchers. Because the approach eliminates the need for sequencing, it also helps cut costs.

The development of CRISPRmap is supported by a NIH Director's New Innovator Award, and designed to be cost-effective, accessible, and compatible with concurrent multiplexed protein and mRNA profiling. The new paper from the Gaublomme lab has demonstrated CRISPRmap as a powerful tool for broad biological and medical discovery.