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RNA-peptide/protein interactions have been of utmost importance to life since its earliest forms, reaching even before the last universal common ancestor (LUCA). However, the ancient molecular mechanisms behind this key biological interaction remain enigmatic because extant RNA-protein interactions rely heavily on positively charged and aromatic amino acids that were absent (or heavily under-represented) in the early pre-LUCA evolutionary period. Here, an RNA-binding variant of the ribosomal L11 C-terminal domain was selected from a ∼1010 library of partially randomized sequences, all composed of 10 prebiotically plausible canonical amino acids. The selected variant binds to the cognate RNA with a similar overall affinity although it is less structured in the unbound form than the wild-type protein domain. The variant complex association and dissociation are both slower than for the wild-type, implying different mechanistic processes involved. The profile of the wild-type and mutant complex stabilities along with MD simulations uncover qualitative differences in the interaction modes. In the absence of positively charged and aromatic residues, the mutant L11 domain uses bridging ion (K+/Mg2+) interactions between the RNA sugar-phosphate backbone and glutamic acid residues as an alternative source of stabilization. This study presents experimental support to provide a new perspective on how early protein-RNA interactions evolved, where the lack of aromatic/basic residues was compensated by acidic residues plus metal ions.

CYC116 is a selective Aurora kinase inhibitor that has been tested in a Phase I study in patients with advanced solid tumors. Although CYC116 has shown desirable preclinical efficacy, the potential for emergence of resistance has not been explored. We established several CYC116 resistant clones from isogenic HCT116 p53+/+ and HCT116 p53−/− cell line pairs. We also generated resistant clones towards ZM447439 (quinazoline derivative), a model Aurora inhibitor. The selected clones were 10-80 fold resistant to CYC116 and cross-resistant to other synthetic Aurora inhibitors including AZD1152, VX-680, and MLN8054. Resistant clones displayed multidrug resistant phenotypes, tested by using 13 major cytostatics. All clones were highly resistant to etoposide followed by other drugs. Interestingly, all CYC116 clones but not ZM447439 became polyploid. ZM447439, but not CYC116 induced three novel mutations in Aurora B. Leu152Ser significantly affected ZM447439 binding, but not CYC116. Gene expression studies revealed differential expression of more than 200 genes. Some of these genes expression profiles were also observed in CYC116 resistant primary tumors. Bcl-xL (BCL2L1) was found to be overexpressed in CYC116 clones and its knockdown resensitized the p53+/+ resistant clone to CYC116. Finally Bcl-xL overexpressing p53+/+ CYC116 clones were highly sensitive to navitoclax (ABT-263) compared to parent cells. The data shed light on the genetic basis for resistance to Aurora kinase inhibitors which could be used to predict clinical response, to select patients who might benefit from therapy and to suggest suitable drug combinations for a particular patient population.