Episode Summary:

Technologies like next-generation sequencing allow us to understand which RNA transcripts and proteins are expressed in biological tissues. However, it’s often equally important to understand how cells or molecules are positioned relative to one another! Whether it be a cell changing its shape, an organelle ramping up a metabolic process, or a DNA molecule traveling across the nucleus, understanding spatial context is critical. Current approaches for spatial sequencing are limited by cost, complicated equipment, sample damage, or low resolution. Recognizing this challenge, Josie and team developed Light-seq, a cheap and accessible method to combine sequencing and imaging in intact biological samples. Not only is the method inexpensive, but Light-seq can also achieve unprecedented spatial resolution by using light to add genetic barcodes to any RNA, allowing scientists to determine exactly where sequencing should occur with extreme precision. By helping researchers to understand spatial context, Light-seq-driven insights may illuminate cancer, neurodegeneration, and autoimmunity.

Episode Notes:

About the Author

• Following her lifelong passion for computer programming, Josie studied Computer Science at Caltech and worked as a software engineering intern at Google. At Caltech, a biomolecular computation course introduced her to the field of biomolecular programming. Josie was quickly excited about the intersection of computers and biology and its potential to bring about positive change in the world. She pursued this interest in her graduate studies in the Wyss Institute for Biologically Inspired Engineering at Harvard, where – as first a postdoctoral fellow, and then the Technology Development Fellow – she developed platform technologies for DNA-based imaging and sequencing assays.

Key Takeaways 

• Next-generation sequencing is a powerful technology to read the transcriptomic state of biological tissues by surveying the RNA transcripts present.
• However, it’s important to understand not only what is being expressed but where this expression occurs! The spatial arrangement, structure, and interactions between molecules are critical to define the functions of biological systems.
• By linking imaging with -omics profiling, the field of spatial biology seeks to understand molecules like RNAs in their 2D and 3D contexts.
• Unfortunately, currently available spatial transcriptomics methods are limited in their ability to select individual cells with complex morphologies, require expensive instrumentation or complex microfluidics setups to the tune of several $100K, and often damage the samples.
• Further, rare cells are often missed due to lower sequencing throughput, even though they may be critical for biological activity.
• Recognizing this challenge, Josie and her collaborators developed Light-seq, a new, cheap, and accessible approach for single-cell spatial indexing and sequencing of intact biological samples.
• Using light-controlled nucleotide crosslinking chemistry, Light-seq can correlate multi-dimensional and high-resolution cellular phenotypes – like morphology, protein markers, spatial organization) – to transcriptomic profiles across diverse sample types.
• In particular, using the biological equivalent of photolithography, Light-seq can add genetic barcodes to any RNA by shining light on it, allowing scientists to control exactly where sequencing should occur with extreme precision – up to the subcellular level.
• Light-seq can operate directly on the sample: the method does not require cellular dissociation, microfluidic separation/sorting, or custom capture substrates or pre-patterned slides.
• Samples used for Light-seq remain intact for downstream analysis post-sequencing.
• Josie evaluated Light-seq on mouse retinal sections to barcode three different cell layers and study the rare dopaminergic amacrine cells (DACs).

Impact 

• Josie created a cheap, accessible, and powerful tool for scientists to perform spatial sequencing at unprecedented resolution without requiring expensive or complicated setups.
• By enabling new advances in spatial biology, Light-seq has the potential to help biologists discover biomarkers for disease, measure on and off target effects of therapeutic candidates, and illuminate poorly understood biological mechanisms where understanding spatial context makes all the difference.

Author: Josie Kishi

Paper: Light-Seq: Light-directed in situ barcoding of biomolecules in fixed cells and tissues for spatially indexed sequencing