Friday, August 11, 2017
Proteomics and lipidomics combines to reveal shocking membrane complexity in Plasmodium!
These authors sure know how to make my day! An amazingly in-depth analysis of the lipid membrane microdomains of anything would probably get my attention anyway, but this great new paper is extra awesome:
1) They combine proteomics and lipidomics
2) They do the work on the sexual stage parasites of the universally despised Plasmodium genus (in this case, the mouse infecting model organism Plasmodium berghei
3) They do top notch bioinformatics that not only leads to output showing enrichment of the proteins interacting in this awful-to-work-with mostly detergent-resistant mess of lipids, but also protein-lipid associations (what?!?)
4) They show how incredibly important designing an upstream experiment is in biology by pulling all of the proteomics with a linear ion trap and some really clever analysis and the lipidomics via TLC and GC.
How'd they pull this off? With an amazingly painful amount of upstream sample preparation. What? Another list? Sorry, I have to, to capture how glad I am that they did this and I didn't have to.
1) Synchronized the parasite (difficult)
2) Infected mouse.
3) Removed parasites when they reached their sexual stage (difficult to determine AND remove)
4) Separated infected from uninfected cells on Nycodenz gradient (blech. blech. blech.)
5) And then it gets fun. Lysing the infected red blood cells, determining the protein content at every stage with anti-parasite antibodies and more detergents and gradient ultracentrifugation and some homogenization.
6) I have to end the list. The amount of work here is truly just awe-inspiring. It just keeps going....
The amount of care that these authors spend on ensuring that they aren't going to the next awful painful step unnecessarily by carefully determining what they currently have is just amazing. I'm so glad there are people in this world who go to these lengths to help understand malaria so we can kill it.
Once they obtain their lipid rafts, they break them down with chloroform and methanol to get the peptides and take the lipids through a staggering amount of work to HPTLC and GC readouts.
And then?? then they make sense of all these measurements and more that I didn't mention!! The network analysis of the proteins is linked with the major lipid class identifications and paints a surprisingly complex picture of the parasite's membrane proteome -- and how it interacts with an also surprisingly heterogeneous lipid profile in the strains they work with.
This was a monumental amount of work, and I'm beyond impressed.
All data is uploaded at Massive and will be available at full paper release (the authors include access info now if you really want it)