Abstract
Alternative cleavage and polyadenylation (ApA) is known to alter untranslated region (3ʹUTR) length but can also recognize intronic polyadenylation (IpA) signals to generate transcripts that lose part or all of the coding region. We analyzed 46 3ʹ-seq and RNA-seq profiles from normal human tissues, primary immune cells, and multiple myeloma (MM) samples and created an atlas of 4927 high-confidence IpA events represented in these cell types. IpA isoforms are widely expressed in immune cells, differentially used during B-cell development or in different cellular environments, and can generate truncated proteins lacking C-terminal functional domains. This can mimic ectodomain shedding through loss of transmembrane domains or alter the binding specificity of proteins with DNA-binding or protein–protein interaction domains. MM cells display a striking loss of IpA isoforms expressed in plasma cells, associated with shorter progression-free survival and impacting key genes in MM biology and response to lenalidomide.
Highlights
Alternative cleavage and polyadenylation (ApA) is known to alter untranslated region (3ʹUTR) length but can recognize intronic polyadenylation (IpA) signals to generate transcripts that lose part or all of the coding region
We identified robust ApA events that occur in introns and quantified IpA isoform expression using 3ʹ-seq across human tissues, immune cells, and in multiple myeloma (MM) patient samples
We found that IpA isoforms are widely expressed, most prevalently in blood-derived immune cells, and that generation of IpA isoforms is regulated during B-cell development, between cellular environments, and in cancer
Summary
IpA isoform expression has previously been viewed as a form of alternative splicing involving alternative last exon usage[15,46], but the defining event is usage of an intronic alternative polyadenylation signal. In matching RNA-seq data, we did not find read evidence upstream of transcription start sites associated with our predicted non-coding RNAs. CLIP-sequencing data analysis showed that the exonized intronic sequence of the 5ʹIpA isoforms contains binding sites for RNA-binding proteins (RBPs); potentially, these non-coding RNAs serve as scaffolds for RBPs and thereby exert a regulatory role in trans on other RNAs. The majority of IpA isoforms (n = 2667), are predicted to generate truncated proteins that retain at least one domain and have the potential to be functional. We found that IpA is another potential mechanism to produce soluble versions of membrane-bound receptors, as IpA-generated truncations occur close to the TMD This suggests that developmental regulation of membrane-bound versus secreted molecules—first described for IgM—is widespread and can mimic proteolytic cleavage. In patients who retain expression of CUL4A-IpA, displaying similar IpA usage to PCs overall, we hypothesize that this truncated protein isoform may provide a mechanism of resistance to lenalidomide. Not all cancer cells show depletion of IpA isoforms, as we found increased IpA isoform expression in another B-cell malignancy, chronic lymphocytic leukemia, reported elsewhere
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