Revealing protein-RNA-interactions using RNA-immunopecipitation

In recent years, the involvement of long non-coding RNAs (ncRNAs) in epigenetic processes has been discovered and examined. However, the function of the majority of these ncRNAs remains still elusive. There is evidence that some long ncRNAs participate in histone modification processes. Based on a genome-wide tiling array study, we found several long ncRNAs to be differentially expressed upon cytokine stimulation and cell cycle status. To identify ncRNAs associated with chromatin remodeling machinery, we isolate relevant multi-protein complexes from cell lysates by RNA immunoprecipitation (RIP). Interacting RNAs are coprecipitated and identified using customized microarrays interrogating high numbers of regulated ncRNAs. Subsequently, molecular studies will be performed to reveal the function of those ncRNAs found to be associated with multi-protein complexes. In this work, different RIP methods are tested on exogenous and endogenous RNA conditions.