Single spinal neuron proteomics of ALS victims

 

I've been looking forward to seeing this one out! 

Background: ALS fucking sucks and we still don't know anywhere near enough about it except there is appears to be a genetic component for a very small amount of people who will spontaneously get the disease and inevitably die from it. This stupid fucking disease stole one of our field's brightest stars a few years ago and I still have to stop myself from writing him for help when I'm stuck on something. We don't have diagnostics for it, except in maybe 3% of people with a TDP-43 mutation. Fragments of the TDP-43 protein seem to be very linked to the disease because bits of the protein with or without different PTMs appear to accumulate and may be linked to other diseases beyond ALS. 

Unfortunately, motor neurons are surrounded by all sorts of other cells, so homogenization screws up any signal that we should/could have when studying post-mortem tissue from ALS. 

Need a reason for ultra-sensitive mass spectrometry? Here's one! This big collaboration between BioGen and two great labs at BYU use laser capture microdissection of postmortem tissue from matched healthy controls and ALS victims to cut out the motor neurons and use nanoPOTs to get all the peptide they can out of the neurons. 

The analysis was performed on an Exploris 480 running what I'm pretty sure was a 20nL/min flow rate over 100 minutes (low flow was achieved using an RSLCnano with a split flow). A data dependent method was used (120,000 MS1 30,000 MS/MS with a 1e5 AGC target and 500ms max injection time) and the results were processed in Proteome Discoverer + Infernys. Wide windows weren't used in this one. I suspect this study started before some recent method development work from these labs that I've recently read and posted about here.  

End results? Around 500 proteins appear to be significantly different in ALS affected neurons! Way more than anyone has ever seen in an ALS proteomics study. Possible biomarkers? We can hope! Also, we've got a method now for going after other nefarious and poorly understood diseases.

Funny perspective on the study -- "Limitations of this study" includes something like "we were only able to identify 2,500 proteins". Oh. Is that it?  Our second attempt at mouse neurons got us to around 400 proteins/cell over the Xmas holidays but the group will probably do a lot better on the third attempt without me in the way. 

Beautiful amazing study with far reaching ramifications. I can't recommend it enough.