So, what happens if we restore pyrimidines to these cells? A complete loss of the ASP rebound and a return to the (very reasonable) phenotype of monotonic ASP depletion. So, SDH loss causes a disproportionate effect on pyrimidine synthesis by impairing ATCase, but why? - ThreadSky

lucasbsullivan.bsky.social • 4 days ago

So, what happens if we restore pyrimidines to these cells?

A complete loss of the ASP rebound and a return to the (very reasonable) phenotype of monotonic ASP depletion.

So, SDH loss causes a disproportionate effect on pyrimidine synthesis by impairing ATCase, but why?

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lucasbsullivan.bsky.social•4 days ago

SDH inhibition is distinct from other mitochondrial impairments due to the accumulation of succinate, which we hypothesized to play a role. First, we decreased succinate production by cotreating with the CI inhibitor rotenone. Result: restoration of pyrimidine synthesis, even with comparable ASP.

lucasbsullivan.bsky.social•4 days ago

A literature search found a goldmine: E Coli ATCase was a workhorse of biochemistry for decades (e.g. ⬇️) One compound used to impair its function in vitro? Succinate! While the concentrations needed to impair ATCase in vitro seemed unrealistic at ~1+ mM...

https://pubmed.ncbi.nlm.nih.gov/3894357/

lucasbsullivan.bsky.social•4 days ago

We found SDH impaired cells can reach >10 mM!

Conclusion: succinate competes with diminished ASP at ATCase, blocking pyrimidine synthesis. To our knowledge the first description of this relationship in cells + revealing another potential regulatory function of succinate.

lucasbsullivan.bsky.social•4 days ago

The full paper has plenty more (ASP flux measurements, succinate sufficiency, replication stress), so I invite you to give it a full read. There are also fun phenotypes that epitomize some of the logical pitfalls that we metabolism nerds often warn about (e.g. levels =/= flux).

lucasbsullivan.bsky.social•4 days ago

TLDR – we find that, upon SDH loss, the combined effects of ASP depletion and succinate accumulation profoundly inhibit ATCase. Pyrimidine loss impairs ASP consumption for biosynthesis, causing ASP to accumulate until it outcompetes succinate to restart pyrimidines/biosynthesis once again.

lucasbsullivan.bsky.social•4 days ago

Thanks to Madeleine for wanting to stick around post-graduation to figure out “the boomerang” and the rest of the team for support, esp Kristian for quantitative efforts. Thanks to NIGMS and NCI for funding. Also, watch out for Madeleine's next act with @saranowinski.bsky.social!

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