Mitochondrial ribosomal proteins developed unconventional mitochondrial targeting signals due to structural constraints

Mitochondrial ribosomes are complex molecular machines indispensable for respiration. Their assembly involves the import of several dozens of mitochondrial ribosomal proteins (MRPs), encoded in the nuclear genome, into the mitochondrial matrix. Available proteomic and structural data as well as computational predictions indicate that up to 25% of MRPs do not have a conventional N-terminal mitochondrial targeting signal (MTS). We characterized a set of 15 yeast MRPs in vivo and showed that 30% of them use internal mitochondrial targeting signals. We isolated a novel internal targeting signal from the conserved MRP Mrp17 (bS6). The Mrp17 targeting signal shares some properties as well as import components with conventional MTS-containing proteins but is not reliably predicted indicating that mitochondrial protein targeting is more versatile than expected. We hypothesize that internal targeting signals arose in MRPs when the N-terminus extension was constrained by ribosome assembly interfaces.