Integration of hyperspectral imaging and transcriptomics from individual cells with HyperSeq

Microscopy and omics are complementary approaches to probe the molecular state of cells in health and disease, combining granularity with scalability. While important advances have been achieved over the last decade in each area, integrating both imaging- and sequencing-based assays on the same cell has proven challenging. In this study, a new approach called HyperSeq that combines hyperspectral autofluorescence imaging with transcriptomics on the same cell is demonstrated. HyperSeq was applied to Michigan Cancer Foundation 7 (MCF-7) breast cancer cells and identified a subpopulation of cells exhibiting bright autofluorescence rings at the plasma membrane in optical channel 13 ({lambda}ex = 431 nm,{lambda} em = 594 nm). Correlating the presence of a ring with the gene expression in the same cell indicated that ringed cells are more likely to express hallmark genes of apoptosis and less likely to express genes associated with ATP production. Further, correlation of cell morphology with gene expression suggested that multiple members of the spliceosome were downregulated in larger MCF-7 cells. Multiple genes were evenly expressed across cell sizes but also exhibited higher usage of specific exons in larger or smaller cells. Finally, correlation between gene expression and fluorescence within the spectral range of Nicotinamide adenine dinucleotide hydrogen (NADH) provided insight into the metabolic states of MCF-7 cells. These observations provided a link between the cells optical spectrum and its internal molecular state, demonstrating the utility of HyperSeq to study cell biology at single cell resolution by integrating spectral, morphological and transcriptomic analyses into a single, streamlined workflow.