Structural Co-evolution of the LARP Superfamily and its Role in Functional Plasticity

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PROJECT SUMMARY/ABSTRACT The long-term goal of our research program is to decipher the molecular mechanism of “non-canonical” protein- RNA interactions studying a particularly fascinating and disease-relevant family of proteins, called the La-related protein (LARP) superfamily, on a molecular level. Cytoplasmic LARPs play a pivotal role in post-transcriptional gene control by regulating the delicate balance between active translation, degradation, and storage of mRNAs. Hence, many LARPs are intimately implicated in various cancers and fibroproliferative diseases rendering them an important class of druggable targets. However, the development of therapies has been stunted by the lack of detailed molecular-level information. Our research will elucidate the molecular mechanism of RNA recognition exhibited by LARPs, in particular, to explain the intricacies of commonality and individuality, i.e. how their com- mon RNA-binding motif, called the La-module, has individually evolved to allow specific RNA recognition and thus achieve its distinct function. The investigation is based on our unique strength in solution and solid-state NMR spectroscopy and their close coupling with other biochemical, biophysical, computational, and functional approaches. Our initial efforts will follow two lines of inquiry, simultaneously focusing on two members of the LARP superfamily, hLARP6 and hLARP1. In the first line of inquiry, we will explore how the La-module of hLARP6 achieves the exclusive recognition of the highly conserved 5' stem-loop (5'SL) motif, which is found in all verte- brate mRNAs encoding type I collagens. This line of study will provide a detailed molecular-level map on how individual elements of the La-module contribute to the specificity and affinity of 5'SL binding. The detailed insights to be gained, together with currently available biochemical and biophysical data, will provide essential insights into the molecular “symbiosis” of the individual elements of the La-module and close this critical gap in knowledge required for the development of therapeutic strategies against fibroproliferative diseases. In the second line of inquiry, we will analogously dissect how the La-module of hLARP1 recognizes a distinctly different type of RNA compared to hLARP6. hLARP1 was found to be heavily involved in proliferation and cell cycle defects and to be significantly upregulated in malignant cells and tissues. Very recent biochemical studies revealed that the La- module of hLARP1 sequentially binds to the 3' poly(A) and then to the 5ʹ terminal oligopyrimidine (5ʹTOP) motifs of mRNAs. Notably, this peculiar two-step behavior has not yet been observed for any other LARP. We will uncover this unusual molecular mechanism by investigating the structural and dynamic changes of hLARP1 upon binding of the poly(A) and 5'TOP motifs. This study will for the first time reveal how the initial binding of one RNA to its La-module elicits structural and dynamic changes required for the binding of a second RNA target. Overall, these two lines of comparative investigations will synergistically lead us to understand the fundamental principles that connect the co-evolution of the individual elements of the La-module and their specific role in RNA recognition, thus explaining how structural and dynamic plasticity contribute to functional plasticity.