Yeast de novo proteins integrate into cellular systems using ancient protein targeting and degradation pathways

Recent evidence demonstrates that eukaryotic genomes encode thousands of evolutionarily novel proteins that originate de novo from non-coding DNA and can contribute to species-specific adaptations. Yet, it remains unclear how these incipient proteins--whose sequences are entirely new to nature--navigate the cellular environment to bring about phenotypic change. Here, we conduct a systematic in vivo investigation of yeast de novo proteins with enhanced growth phenotypes, revealing the early stages of cellular integration. We find that these proteins are strongly enriched at the endoplasmic reticulum (ER) relative to conserved proteins, and that they integrate into cellular systems through conserved membrane targeting, trafficking, and degradation pathways. Despite having unrelated sequences, ER-localized de novo proteins share a common molecular signature: a C-terminal transmembrane domain that likely enables recognition by conserved post-translational ER insertion pathways. After insertion, ER-localized de novo proteins traffic from the ER and their homeostasis is regulated by conserved proteasomal and vacuolar degradation pathways. Our findings demonstrate that ancient targeting and degradation pathways can accommodate young de novo proteins sharing a convergent molecular signature. These pathways may act as selective filters, biasing which young de novo proteins persist. Significance StatementDeeply conserved genes shape the core structure and function of cells, but novel genes are key drivers of biodiversity. Novel genes can arise by divergence from ancient genes, or de novo from non-genic DNA. For de novo genes to develop novel functions, it is necessary for them to be processed by the cell so that their protein levels and localization are regulated. However, little is known about how novel de novo genes obtain the capacity to engage the cellular machinery needed for this regulation. Here, we experimentally assess how a set of endoplasmic reticulum (ER)-localized proteins encoded by recently-evolved de novo genes are localized and degraded in yeast cells. We discover that, despite having entirely unique amino acid sequences, these proteins share biochemical signatures allowing them to engage the same ancient cellular machinery and localize to the ER membrane. Interestingly though, this machinery is not the one that targets most ancient proteins to the ER. These results indicate that even recently emerged proteins without an extensive period of evolutionary adaptation can be recognized by specific ancient cellular pathways, facilitating their localization and homeostasis.