Assistant Professor of Biochemistry, Biophysics and Structural Biology
The last decade has seen an explosion in the number of available macromolecular structures visualized at atomic resolution. In large part, cryo-electron microscopy (cryo-EM) has been the main technique delivering atomic structures of a diverse range of molecules. In parallel, machine learning algorithms such as Alphafold2 are more successfully predicting protein structures from their amino acid sequences. These developments have yielded an unprecedented databank of experimentally determined and predicted protein structures. This “structureome” could be a reference for the broader biological community, analogous to the genome. However, for this to become a reality, methods that can use structureomic data to understand how molecules work together to realize the plethora of biological processes inside cells, tissues and organisms are needed. The Lucas Lab develops new approaches to leverage the growing compendium of atomic-resolution structures and atomistic models to investigate the molecular details of life using cryo-EM.
Visualizing ribosome assembly in the nucleolus, nucleus and cytoplasm
Ribosomes are the modular molecular machines that translate the genetic code to produce every protein in the cell. The assembly of these large macromolecular machines consumes more than half of a growing cell's energy. In yeast >200 ribosome biogenesis factor proteins are required to assemble the 4 rRNAs and ~80 proteins to produce ~2,000 ribosomes per minute. Structures of purified intermediates in the assembly pathway have revealed how some of these 200 biogenesis factors aid assembly. However, in cells, ribosome assembly spans three subcellular compartments, the nucleolus, the nucleus and the cytoplasm. How the detailed molecular arrangements are coordinated in the cell is unclear. We make use of new cryo-EM methods to visualize intermediates of ribosome assembly from the nucleolus to the cytoplasm. Using this approach, we can correlated the detailed molecular structures of intermediates with their subcellular location. We aim to build a spatially resolved detailed molecular model for an entire biological pathway in yeast.
While there is a wealth of information about the key factors in yeast ribosome assembly, human ribosome assembly is comparatively poorly understood. Screens have identified hundreds of potential ribosome assembly factors without a yeast homolog and for which the function is unknown. We use in situ cryo-EM to characterize human intermediates of ribosome assembly with the goal to characterize the functions of these additional assembly factors.
Improved methods for FIB-milling
Cryo-EM can reveal the atomic details of cells. However, most cells are too thick to image directly with cryo-EM and need to be thinned. The application of Focused Ion Beam (FIB) milling to thin cryogenically frozen cells was a major development that allowed visualization of the internal architecture of almost any cell. However, how the milling process affects the quality of the sample is only beginning to be understood. Our lab uses 2DTM to quantitatively analyze the damage profiles caused by milling and uses this approach to identify and test alternate, possibly less damaging approaches.
*co-first author #co-corresponding author
- Lucas BA*, Himes BA*, Xue L, Grant T, Mahamid J, Grigorieff N. Locating macromolecular assemblies in cells by 2D template matching with cisTEM. eLife 10:e68946 (2021).
- Lucas BA#, Zhang K, Loerch S, Grigorieff N#. In situ single particle classification reveals distinct 60S maturation intermediates in cells. eLife 11:79272 (2022).
- Lucas BA# and Grigorieff N#. Quantification of gallium cryo-FIB milling damage in biological lamella. bioRxiv (2023). https://doi.org/10.1101/2023.02.01.526705
Lucas BA, Lavi E, Shuie L, Cho H, Katzman S, Miyoshi K, Siomi MC, Carmel L, Ares Jr. M, Maquat LE. Evidence for convergent evolution of SINE-directed Staufen-mediated mRNA decay. Proceedings of the National Academy of Sciences. 115: 968–973 (2018).
-Highlighted by Cedric Feschotte in Faculty Opinions (F1000)
- Elbarbary RA*, Lucas BA*, Maquat LE. Retrotransposons as regulators of gene expression. Science. 351: aac7247 (2016)
Last Updated 2023-04-18