Ribosomes are comprised of both rRNAs, which perform the catalytic function of protein synthesis, and a large number of proteins that form the structural scaffold. Transcription of the 35S rRNA precursor of 28S, 18S, and 5.8S by RNA polymerase I take place in the nucleolus where ribosomal assembly occurs. Only 5S rRNA is usually transcribed in the nucleoplasm by RNA polymerase III and brought into the nucleolus together with ribosomal protein L5. In the only well characterized eukaryotic system, Saccharomyes cerevisiae, two assembly factors, Rpf2 and Rrs1 as well as ribosomal protein, L11, are required to bring the early preribosomal (5S rRNA and L5) to the assembling ribosome. Our laboratory has defined the earliest preribosomal particle in trypanosomes which contains the conserved components 5S rRNA and L5 as well as the trypanosome-specific proteins, P34/P37. We have identified key determinants of this novel tri-molecular complex as well as unique features of the complex that we hypothesize are essential to its structure and function. We have also shown that both L5 and P34/P37 are essential to the formation of functional ribosomes and therefore, the viability of T. brucei. In this proposal we will identify and characterize the assembly components of the preribosomal particle and use our knowledge to define potential targets for chemotherapeutic approaches. Our hypothesis is that the interactions between the components of the essential preribosomal complex will provide valid targets for chemotherapeutic disruption of ribosomal assembly in T. brucei.
The specific aims of the project are to: 1. Determine the critical proteins that enable the association of the 5S rRNA-containing complex with the ribosome and their function in ribosome biogenesis. 2. Define the interaction network among the pre-ribosomal complex components and examine how the disruption affects the function of the complex within the cell. 3. Develop FRET approaches to study RNA-protein interaction, in vivo protein-protein interactions, and screen for small interfering molecules. Although the ribosome is highly conserved, subtle differences between the host and pathogen have enabled the development of drugs specifically targeting pathogen ribosome assembly. Many existing drugs for the treatment of bacterial infections specifically bind to the functionally relevant sites of the bacterial ribosome. The pre-ribosomal complex that is the focus of this proposal is essential to trypanosome ribosome assembly and function and to the survival of the parasite. Proteins within the complex are either unique to trypanosomes (P34/P37) or contain features clearly distinct from those of the host (L5). Our work will also continue to provide new insights that will impact the broader field of eukaryotic ribosomal biogenesis. Moreover, the unique features of the complex will allow us to define and exploit this target for chemotherapeutic development.
The protozoan parasites, the Kinetoplastids, include the organisms Trypanosoma brucei, Trypanosoma cruzi and Leishmania, which cause serious diseases in much of the world (African sleeping sickness, Chagas' disease and leishmaniasis, respectively). This study will examine a unique preribosomal complex occurring in these parasites with the goal of developing an assay to identify drugs that target the parasite but not host complex.
| Kamina, Anyango D; Williams, Noreen (2017) Ribosome Assembly in Trypanosomatids: A Novel Therapeutic Target. Trends Parasitol 33:256-257 |
