Biology subjects

Christiansen, J.

Publications and source records attributed to Christiansen, J..

2 recordsLinked to original sources

bioRxiv2020

Fluorescence Correlation and Cross-Correlation Spectroscopy Unveil Cytoplasmic mRNP Composition and Dynamics

Understanding the mRNA life cycle requires analysis of the dynamic macromolecular composition and stoichiometry of mRNPs. Fluorescence correlation and cross-correlation spectroscopy (FCS and FCCS) are appealing technologies to study mRNP complexes because they readily provide information about the molecular composition, stoichiometry, heterogeneity and dynamics of the particles. We developed FCS protocols for analysis of live cells and cellular lysates, and demonstrate the feasibility of analysing common cytoplasmic mRNPs composed of core factor YBX1, IMPs (or IGF2BPs) and their interactions with other RNA binding proteins such as PABPC1, ELAVL2 (HuB), STAU1 and FMRP. FCCS corroborated previously reported RNA dependent interactions between the factors and provided an estimate of the relative overlap between the factors in the mRNPs. In this way FCS and FCCS provide a new and useful approach for the quantitative and dynamic analysis of mRNP macromolecular complexes that may complement current biochemical approaches.

cell biology↗

bioRxiv2019

Single mRNP analysis by super-resolution microscopy and fluorescence correlation spectroscopy reveals that small mRNP granules represent mRNA singletons

Small cytoplasmic mRNP granules are implicated in mRNA transport, translational control and decay. Employing Super-resolution Microscopy and Fluorescence Correlation Spectroscopy, we analyzed the molecular composition and dynamics of single cytoplasmic YBX1_IMP1 mRNP granules in live cells. Granules appeared elongated and branched with patches of IMP1 and YBX1 distributed along mRNA, reflecting the attachment of the two RNA-binding proteins in cis. Particles form at the nuclear pore and are spatially segregated from translating ribosomes, so the mRNP is a repository for mRNAs awaiting translation. Individual mRNPs contain a single mRNA and 5 to 15 molecules of YBX1 and IMP1, which is in agreement with the average number of mRNA-binding sites calculated from CLIP analyses. We conclude that small cytoplasmic mRNP granules are mRNA singletons, thus depicting the cellular transcriptome. Consequently, expression of functionally related mRNAs in RNA regulons is unlikely to result from coordinated assembly. O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY Graphical abstract C_FIG_DISPLAY

molecular biology↗