Cotranslational signal independent SRP preloading during membrane targeting

Ribosome-associated factors must faithfully decode the limited information available in nascent polypeptides to direct them to their correct cellular fate(1). It is unclear how the low complexity information exposed by the nascent chain suffices for accurate recognition by the many factors competing for the limited surface near the ribosomal exit site(2,3). Questions remain even for the well-studied cotranslational targeting cycle to the endoplasmic reticulum (ER), involving recognition of linear hydrophobic Signal Sequences (SS) or Transmembrane Domains (TMD) by the Signal Recognition Particle (SRP)(4,5). Intriguingly, SRP is in low abundance relative to the large number of ribosome nascent chain complexes (RNCs), yet it accurately selects those destined to the ER(6). Despite their overlapping specificities, SRP and the cotranslational Hsp70 SSB display exquisite mutually exclusive selectivity in vivo for their cognate RNCs(7,8). To understand cotranslational nascent chain recognition in vivo, we interrogated the cotranslational membrane targeting cycle using ribosome profiling (herein Ribo-seq)(9) coupled with biochemical fractionation of ribosome populations. Unexpectedly, SRP preferentially binds secretory RNCs before targeting signals are translated. We show non-coding mRNA elements can promote this signal-independent SRP pre-recruitment. Our study defines the complex kinetic interplay between elongation and determinants in the polypeptide and mRNA modulating SRP-substrate selection and membrane targeting.