Alternative 3’UTRs play a widespread role in mRNA association with the endomembrane system

Transcripts encoding membrane and secreted proteins are known to have translation on endoplasmic reticulum (ER) through the signal recognition particle (SRP)-mediated mechanism. Translation-independent association of mRNAs with ER has also been reported, but poorly understood. Here, using 3’ end sequencing, we examine transcripts associated with polysomes on ER in mouse C2C12 cells. We identify transcript features that facilitate translation-independent ER association, including CDS size, 3’UTR size, and GC content. Interestingly, transcripts encoding nuclear proteins tend to have a higher ER association potential than other transcripts, suggesting localized translation to facilitate nuclear import. We show long 3’UTR isoforms are generally more likely to be associated with ER than short isoforms. Consistently, global shortening of 3’UTRs by knocking down CFI-25 expression leads to substantial alternation of mRNA association with ER. While ER association potentials of transcripts undifferentiated myoblasts are similar to those in differentiated myotubes, 3’UTR lengthening in differentiation leads to more transcripts associated with ER in myotubes. Moreover, translation-independent ER association correlates with low mRNA stability, which is exacerbated in cells with oxidative stress. Together, our data indicate that sequence and structure features of mRNAs facilitate ER association and translation, potentially impacting localization dynamics of protein products, and cellular stress remodels membrane-associated transcripts through mRNA stability regulation. Three RNAseq samples without replicate plus 37 3'READS+ samples with 1 or 2 replicates.