Trempe Lab

We are back on Facebook! Here is our latest story on the mechanism of PINK1 activation via autophosphorylation. Congratulations to PhD student Shafqat Rasool, with critical contribution from postdoc Simon Veyron and collaborators from the Lukacs and Fon labs. Here is a little story about it.

This project stemmed from the observation that PINK1 autophosphorylates upon accumulating on damaged mitochondria. This step is important for efficient Parkin recruitment. Later on, we and others showed that PINK1 autophosphorylates at an invariant serine (S228 in human), and that this step was critical for binding its substrate ubiquitin. Now, folks who know a thing or two about kinases would see that this is a very unusual autophos site. We knew it was in "trans", i.e. one PINK1 molecule with ATP phosphorylates Ser228 in another PINK1 molecule. But the mechanism was unclear.

To solve that problem, we engineered an insect PINK1 variant with solubilizing mutations and co-expressed with a phosphatase to make the completely dephosphorylated protein in vitro. And it crystallized! Crystal contacts revealed in all its glory the autophosphorylation dimer, with the target serine from one molecule reaching out nicely into the active site of the other. PINK1 possesses three unique "insertions" compared to other kinases. We knew that "Insert-3" was critical for Ub binding, but the new dimer structure revealed that "insert-2" is critical for mediating autophosphorylation.

Now, the structure had more to offer. Just upstream of the "canonical" kinase domain is an N-term helix that binds to the C-term extension. The helix harbours Parkinson's mutations, like C125G and A126P, but how these mutations impair PINK1 was unclear. We find that the NT helix and CTE form a module that is essential for PINK1 to form a ~750 kDa complex with the translocase of the outer membrane on mitos. Mutations that disrupt the NT helix basically disrupts PINK1's ability to form that complex. We hope this work will help elucidate how PINK1 acts as a "damage sensor" by integrating and relaying stress signals. There is still much to be learned, especially regarding its intriguing mitochondrial targeting sequences and interactions with the import machinery.

Mechanism of PINK1 activation by autophosphorylation and insights into assembly on the TOM complex Rasool et al. solved crystal structures of the cytosolic domain of PINK1, revealing how dimerization allows for trans autophosphorylation at the conserved Ser205 site. The NT and CTE regions flanking the kinase domain form a module that is essential for PINK1 stabilization on the TOM complex upon m...

A bit late, but congratulations to our own Shafqat Rasool for winning a well-deserved CRBS scholarship.

Congratulations to Andrew (featured in the article below), as well as Nathalie Croteau and Simon Veyron, and all the other CRBS members who worked on the COVID testing kits!
https://mcgill.ca/channels/article/part-something-big

Very interesting paper, for a tool I’m using quite a fair bit.

Evaluating drug targets through human loss-of-function genetic variation Analysis of predicted loss-of-function variants from 125,748 human exomes and 15,708 whole genomes in the Genome Aggregation Database (gnomAD) provides a roadmap for human ‘knockout’ studies and a guide for future research into disease biology and drug-target selection.

Made-in-Canada COVID-19 test developed at McGill receives government support McGill and RI-MUHC team partnering with National Research Council has obtained funding from Innovation, Science, and Economic Development Canada to develop capability to produce millions of SARS-CoV-2 tests to make Canada self-sufficient.

Great talks, well done.

Having fun at lab meetings!

Amazing... may help in designing neutralizing antibodies.

Structural basis for the recognition of SARS-CoV-2 by full-length human ACE2 Scientists are racing to learn the secrets of severe acute respiratory syndrome–coronavirus 2 (SARS-CoV-2), which is the cause of the pandemic disease COVID-19. The first step in viral entry is the binding of the viral trimeric spike protein to the human receptor angiotensin-converting enzyme 2 (A...

Well, that's cool. We've published the structure of Ddi1 back in 2016. I'm intrigued as to how the HDD domain contributes to polyubiquitin-chain recognition, structurally.

Ddi1 is a ubiquitin-dependent protease Many proteins in the cell are tagged with a polyubiquitin chain, which serves as a recognition signal for degradation by the proteasome. Some tagged substrates bind directly to the proteasome, but others are delivered through shuttling factors. Yeast Ddi1 and its homologs in other eukaryotic cells h...

Yep

Un guide simple:

Y a t’il un ingénieur dans la salle?

Code Life Ventilator Challenge To design a low-cost, simple, easy-to-use and easy-to-build ventilator that can serve the COVID patients, in an emergency timeframe.