Spatially Resolved Multibait Mapping of Stress Granule and Processing Body Transcriptome

Stress granules (SGs) are dynamic, membrane-less organelles that house complex RNA-protein networks. Although previous profiling methods have characterized SG RNAs as long, translation-repressed, and extensively epigenetically modified, it remains unclear whether these RNAs are evenly distributed within SGs. In this study, we genetically targeted the photocatalyst protein miniSOG to multiple SG core proteins, enabling the comprehensive CAP-seq profiling of SG-associated RNAs. Our results reveal that RNAs near different SG core proteins display heterogeneous distributions and distinct intrinsic features. We also employed CAP-seq to map RNAs associated with processing body (PB) marker protein DDX6 under both unstressed conditions and arsenite-induced stress. By comparing the transcriptomes proximal to SGs and PBs, our data suggest that m⁶A modification may promote RNA localization to SGs, whereas higher AU content may facilitate mRNA targeting to PBs. These findings point to potential regulatory mechanisms that determine the subcellular localization of mRNAs within membrane-less organelles.