Autophagy and pyroptosis are two major pathways of intracellular cell-autonomous host defense.
Autophagy and pyroptosis are two major pathways of intracellular cell-autonomous host defense.

Intracellular infection

The ability of most cell lineages to defend themselves individually against infection can be considered as the most ancestral form of defense against pathogen pressure. We are interested in the intriguing interplay of autophagy and intracellular bacteria, where the host employs autophagy for bacterial clearance while some pathogens re-direct autophagy as a pro-survival strategy. We work towards understanding the mechanisms of restricting intracellular pathogens by studying three-dimensional structures of the protein machinery involved in this form of cytosolic host defense.


Protein complexes mediate membrane tubulation, vesicle fusion and fission.
Protein complexes mediate membrane tubulation, vesicle fusion and fission.

Membrane remodelling

Intracellular trafficking is characterized by dynamic membrane remodeling events. Pathogens hijack autophagy and other intracellular membranes to subvert immune recogonition, replicate and to escape from host cells. We currently lack a detailed view of how the pathogen protein machinery assembles on membranes to remodel its replicative compartments. The overarching goal of this research program is to understand the structures and mechanism of these protein machineries, and those of the host involved in restricting intracellular pathogens. We use nanotechnology alongside biochemistry and cell biology to build reconstituted systems that we can manipulate precisely, and we use electron imaging methods to visualize them in molecular detail.


Method Development

We actively develop new methods and technology to support our research.

  • Sample preparation methods for cryo-EM:
    In collaboration with Andreas Engel and Murali Ghatkesar we are developing a novel method for single-cell nanobiopsy to selectively target subcellular structures for direct vitrification.

  • Density interpretation and model building:
    We develop methods to facilitate interpretation of cryo-EM density maps and improve the improve of accuracy models derived from them. We have introduced the concept of local sharpening to optimize contrast in maps with local resolution variation and adaptive restraint weigthing for atomic model refinement against a single map target.

  • Microdiffraction: We have developed protocols to utilize the spontaneous protein crystallization in yeast peroxisomes as a source of nanocrystals for in cellulo diffraction using XFELs and electron microdiffraction (microED) and demonstrated that collection of diffraction data in these cases is possible from intact cells.



Publications

Please see here for group publications


Collaborations

Kavli Institute for Nanoscience Delft
Nynke Dekker
Stan Brouns
Cees Dekker

External
Murali Ghatkesar, Microsystems Engineering, TU Delft (NL)
Federico Forneris, Armenise-Harvard Laboratory of Structural Biology, Pavia (IT)

LosScale
Carsten Sachse, EMBL
Tom Burnley, CCP-EM



We gratefully acknowledge support by