DNA binding preferences of S. cerevisiae RNA polymerase I Core Factor reveal a preference for the GC-minor groove and a conserved binding mechanism.

In Saccharomyces cerevisiae, Core Factor (CF) is a key evolutionarily conserved transcription initiation factor that helps recruit RNA polymerase I (Pol I) to the ribosomal DNA (rDNA) promoter. Upregulated Pol I transcription has been linked to many cancers, and targeting Pol I is an attractive and emerging anti-cancer strategy. Using yeast as a model system, we characterized how CF binds to the Pol I promoter by electrophoretic mobility shift assays (EMSA). Synthetic DNA competitors along with anti-tumor drugs and nucleic acid stains that act as DNA groove blockers were used to discover the binding preference of yeast CF. Our results show that CF employs a unique binding mechanism where it prefers the GC-rich minor groove within the rDNA promoter. In addition, we show that yeast CF is able to bind to the human rDNA promoter sequence that is divergent in DNA sequence and demonstrate CF sensitivity to the human specific Pol I inhibitor, CX-5461. Finally, we show that the human Core Promoter Element (CPE) can functionally replace the yeast Core Element (CE) in vivo when aligned by conserved DNA structural features rather than DNA sequence. Together, these findings suggest that the yeast CF and the human ortholog Selectivity Factor 1 (SL1) use an evolutionarily conserved, structure-based mechanism to target DNA. Their shared mechanism may offer a new avenue in using yeast to explore current and future Pol I anti-cancer compounds.

Combinatorial approaches in post-polymerization modification for rational development of therapeutic delivery systems.

The combinatorial polymer library approach has been proven to be effective for the optimization of therapeutic delivery systems. The library of polymers with chemical diversity has been synthesized by (i) polymerization of functionalized monomers or (ii) post-polymerization modification of reactive polymers. Most scientists have followed the first approach so far, and the second method has emerged as a versatile approach for combinatorial biomaterials discovery. This review focuses on the second approach, especially discussing the post-modifications that employ reactive polymers as templates for combinatorial synthesis of a library of functional polymers with distinct structural diversity or a combination of different functionalities. In this way, the functional polymers have a consistent chain length and distribution, which allows for systematic optimization of therapeutic delivery polymers for the efficient delivery of genes, small-molecule drugs, and protein therapeutics. In this review, the modification of representative reactive polymers for the delivery of different therapeutic payloads are summarized. The recent advances in rational design and optimization of therapeutic delivery systems based on reactive polymers are highlighted. This review ends with a summary of the current achievements and the prospect on future directions in applying the approach of post-polymerization modification of polymers to accelerate the development of therapeutic delivery systems. STATEMENT OF SIGNIFICANCE: A strategy to rationally design and systematically optimize polymers for the efficient delivery of specific therapeutics is highly needed. The combinatorial polymer library approach could be an effective way to this end. The post-polymerization modification of reactive polymer precursors is applicable for the combinatorial synthesis of a library of functional polymers with distinct structural diversity across a consistent degree of polymerization. This allows for parallel comparison and systematic evaluation/optimization of functional polymers for efficient therapeutic delivery. This review summarizes the key elements of this combinatorial polymer synthesis approach realized by post-polymerization modification of reactive polymer precursors towards the development and identification of optimal polymers for the efficient delivery of therapeutic agents.