Stephen D. Carter ( CalTech)

Stephen D. Carter ( CalTech)

Special Seminar

Event Date/Location

January 9, 2019 -
12:00 pm to 1:00 pm
Schultz Laboratory 107

Speaker

  • Stephen D. Carter

    California Institute of Technology

Topic

Looking at autophagy and the unfolded protein response pathway inside cells using cryogenic correlated light and electron cryotomography

Cryogenic correlated light and electron cryo-microscopy (cryo-CLEM) is the optimal approach to study structural cellular biology in situ, because imaging the same sample by both light and electron cryo-microscopy (cryo-EM) combines the strengths of both techniques to locate unique objects and reveals their structures natively inside cells. Cryo-CLEM was used to visualize TRIM5a-YFP bodies inside HeLa cells. More specifically, I used cryo-fluorescence microscopy to locate TRIM5a-YFP bodies inside HeLa cells and then imaged them to “macromolecular” (~4 nm) resolution in their native state with electron cryotomography. We found that TRIM5α-YFP bodies are present within all the cellular ultrastructures involved in the autophagy pathway, including the sequestosome, phagophore, autophagosome and auto-lysosome. This approach was then applied to the look at activated IRE1α in the unfolded protein response pathway. These results show that IRE1α oligomers induce membrane deformations, leading to the protrusion of narrow 30 nm ribosome-free tubes that remain connected to the ER and are twisted into glomeruli of complex topology. The tubes contain double-helical IRE1α luminal filaments in their lumen revealed by sub-tomogram averaging at 15 Å. Taken together, these findings define a previously unrecognized subdomain of the ER membrane and shed new light on the structure and organization of active mammalian IRE1α inside the cell.

Audience

Free and open to the university community and the public.

Host

N. Yan, Department of Molecular Biology