Institut de Biologie StructuraleGrenoble / France

Contact person(s) related to this article / PELLEQUER Jean-Luc

Importance of lncRNA tertiary structure in key cellular processes

The IBS AFM team determined by single lncRNA molecule imaging, the conformation of MEG3 under several experimental conditions (native, destabilizing, and denaturing). The attached figure provides a sample of 100 isolated molecules of native MEG3 deposited on mica and air-imaged by atomic force microscopy (AFM) with Peak-Force mode. Compared to AFM imaging of unstructured (poly-A) or known to be structured (group II intron) RNA molecules, the results demonstrate the presence of a particular folding for MEG3 which is consistent with the biochemical and biophysical results of this study led by Marco Marcia from EMBL Grenoble.
The AFM imaging was performed by Jean-Marie Teulon with RNA samples from Tina Uroda. Tina also participated in defining the sample deposition protocol on mica.

Uroda T, Anastasakou E, Rossi A, Teulon J-M, Pellequer J-L, Annibale P, Pessey O, Inga A, Chillon I and Marcia M (2019) Conserved pseudoknots in lncRNA MEG3 are essential for stimulation of the p53 pathway.Mol. Cell 75 : 1-14.

A detailed experimental protocol for studying the structure of long non-coding RNA

A protocol combining atomic force microscopy with small angle X-ray scattering appeared in the review Nature Protocols. To know more, consult the IBS highlights.

Uroda T, Chillon I, Annibale P, Teulon J-M, Pessey O, Karuppasamy M, Pellequer JL and Marcia M (2020) Visualizing the functional 3D shape and topography of long noncoding RNAs by single-particle atomic force microscopy and in-solution hydrodynamic techniques. Nat. Protoc. 15 : 2107-2139.

Finally, the HU protein of Deinococcus radiodurans imaged by AFM

The AFM team of the MEM group, in collaboration with the DNA Damage and Repair team of the VIC group, has made it possible to image for the first time the HU protein of Deinococcus radiodurans by Atomic Force Microscopy. HU is an important, even essential, protein in bacterial nucleotides. A new AFM image processing allows us to observe the positioning of the DrHU protein on native plasmids produced by E. coli as well as on linearized plasmids (image on the left which illustrates the application of the Laplacian-weight filter used in this work). This work highlights two major functions of DrHU in the condensation, but also in the decondensation, of double-stranded DNA. The self-compacting dynamics of naked DNA as well as the concentration of DrHU are important parameters in the cellular function of DrHU.
Image processing and structural interpretations were done by Wendy Chen. AFM imaging was conducted by Jean-Marie Teulon with the participation of Anne-Sophie Banneville on samples prepared by Anne-Sophie.

Chen SWW, Banneville AS, Teulon JM, Timmins J and Pellequer JL (2020) Nanoscale surface structures of DNA bound to Deinococcus radiodurans HU unveiled by atomic force microscopy. Nanoscale 12 : 22628-22638