ZIA BC 008382 (ZIA) | |||
---|---|---|---|
Title | Computational Approaches for RNA Structure/Function Determination | ||
Institution | NCI, Bethesda, MD | ||
Principal Investigator | Shapiro, Bruce | NCI Program Director | N/A |
Cancer Activity | N/A | Division | CCR |
Funded Amount | $377,524 | Project Dates | null - null |
Fiscal Year | 2018 | Project Type | Intramural |
Research Topics w/ Percent Relevance | Cancer Types w/ Percent Relevance | ||
Cancer (100.0%) Digestive Diseases (10.0%) |
Brain (10.0%) Liver Cancer (10.0%) Nervous System (10.0%) |
||
Research Type | |||
N/A | |||
Abstract | |||
Our previous discovery of the structure of the turnip crinkle virus tRNA-like translational enhancer (TCV TSS) has permitted us to pursue the use of a relatively new technique for understanding the structural characteristics of an RNA when optical tweezers are applied to pull the molecular structure apart. Essentially a force is applied to the 5 and 3 prime ends of the molecule, which is then monitored. Force changes are then correlated with structural features. The pulling experiments, in collaboration with Anne Simon, were correlated to simulated steered molecular dynamics, which enabled the visualization of the unfolding events of the molecule as a function of the pulling speed and forces applied. These results shed light on the significant rearrangement that occurs in the translational enhancer tRNA-like structure as a result of RNA dependent RNA polymerase (RdRp) binding to this region to produce the negative strand of the virus.Some unexpected structural relationships were found that at least in part, showed how major rerangements occur in the TSS as a result of RdRp binding. Coarse-grained and explicit solvent techniques were used to elucidate the structural characteristics. This technique offers a unique methodology for understanding RNA structure and the characteristics of various RNA motifs found in the structure. ---We have also pursued, in collaboration with Shuo Gu, a comprehensive examination of potential RNA-RNA interactions that are found across 4 different cell lines. MySeq reads were examined and correlated with computational bioinformatic analysis of potential interactions. The prevalence or lack thereof was determined to enable a better understanding of how cellular RNA interacts with its cellular environment. Interestingly the major finding was that there are very few RNA-RNA interactions found across the 4 cell lines, thus indicating the such interactions are avoided. The number of interactions went up significantly when proteins were removed from the lysates and the sequences re-annealed. Cells presumably avoid such interactions to prevent activation of innate immune responses, which are normally reserved for viruses. ---The functionality of Drosha in cellular systems is important for understanding the processing of microRNAs and how they relate to normal cellular activity as well as diseases such as cancer. In another collaboration with Shuo Gu the relationship of Drosha targeted stem-loop structures and the type of microRNA isforms that are produced was examined. Experimental and computational approaches were applied to determine these relationships. Results indicate that bent, distorted and/or flexible structures in the targeted Drosha stem seem to facilitate the production of alternate forms of microRNA. Structural predictions and experimental results were compared and correlated. ---A collaboration with Esta Sterneck's laboratory is ongoing. Her lab investigates cell signaling pathways involved in breast and glioblastoma tumorigenesis with a focus on the transcription factor CCAAT/enhancer binding protein delta (CEBPD) using in vitro cell culture and in vivo mouse model systems. Using a transgenic mouse model of breast cancer, her group has shown that CEBPD exhibits a dual role in mammary tumorigenesis. On the one hand, CEBPD prevents tumor multiplicity and on the other hand, CEBPD promotes distant lung metastases. In addition, CEBPD promotes stem-like cancer cells, which have been implicated in tumor metastasis and treatment resistance, in breast and glioblastoma tumor cells through regulation of various signaling pathways and stemness. In addition, strategies for targeting the message of CEBPD are necessary to down regulate CEBPD-mediated tumor progression signaling. As a tie-in to our nanobiology project, our laboratory is developing approaches for RNAi therapeutics to knock down the CEBPD mRNA by delivering strategically designed RNA nanostructures as their own entities or in combination with li |