Thread by @connor_rosen

Incredible paper from the Elledge group in @CellCellPress, cell.com/cell/fulltext/…, discovering new regulation of protein stability by “DesCEND” degrons and shaping of eukaryotic proteome... 1/18

Starts with proteome-scale discovery of linear peptide degrons (sequences mediating degradation) - peptide fusion with GFP, co-expressed with DsRED as internal control - sort based on GFP/DsRED ratio, low ratio = unstable GFP = peptide acting as degron... 2/18

CRL (cullin ring E3 ligase) inhibition stabilized a fraction of peptide-GFP fusions - shown to be mostly Cul2 and Cul5 dependent. CRISPR screening leads to some CRL2 adaptors... 3/18

Examination of different CRL2 adaptors revealed KLHDC3 targets proteins ending in glycine - returning to full-proteome scale showed proteins ending in “G” were generally CRL substrates... 4/18

And, glycine as a C-terminal residue is underrepresented across all proteins in multiple eukaryotic genomes (from yeast to plants to mammals!) - as are -2 positions that further favor CRL adaptor binding / degradation... 5/18

Expanding to other C-terminii using a new library consisting of all human protein C-terminii revealed other C-terminal degrons - RxxG, -EE, R-3 - also depleted from protein C-terminii across the proteome!... 6/18

This is a real “tour de force” paper - great systems biology, fantastic assay design, excellent detail on mechanism of degradation, discovery of a multiple branches of new degrons… “DesCEND” as a new degradation rule... 7/18

Companion paper in @MolecularCell, cell.com/molecular-cell… on some particular substrates and further exploration... 8/18

Mol Cell paper answers the first question I had - what about ubiqutin (my favorite protein with a C-terminal glycine!) - not a substrate, but extending C-terminal tail allows for degradation... 9/18

Further questions… Across intracellular proteases, do P-1 positions show preference for stabilizing or destabilizing residues? Does DesCEND primarily clear out N-terminal fragments of cleaved proteins (Mol Cell paper says it can, at least for Usp1)?... 10/18

Mol Cell paper also discusses CRL2 degradation of “gibberish” sequences, early truncations, etc… Maybe acting as a QC mechanism on protein synthesis. Is this a (major / minor) source of DRiPs? Is CRL2 co-localized with ribosome in position to mediate QC?... 11/18

Ubiquitin results suggest level of access required for C-terminal degrees to be efficiently recognized - is that a general rule (both N- and C-terminus tend to be more disordered, but N-terminus may be often buried in complexes - what’s the trend for C-terminus?)... 12/18

What is the extent of dynamic regulation of protein C-terminii? Interesting to note most carboxypeptidases are not cytoplasmic (extracellular or in secretory pathway) - and many preferentially cleave off “stable” C-terminal residues... 13/18

Do proteins that come back out of ER / Golgi / endosomes that have been cleaved by carboxypeptidases get degraded more by DesCEND? (I’m thinking cross-presentation for endocytosed proteins!) Do carboxypeptidases have P-1 preference related to DesCEND rules?... 14/18

Is nuclear localization of Carboxypeptidase E, instead of normal secretion, leading to abnormal destabilization of proteins - hence cancer association with nuclear form (jci.org/articles/view/…)?... Or are there more relevant cytoplasmic carboxypeptidases?... 15/18

Viral proteomes seem to have "responded" to DesCEND by depleting C-terminal G - bacteria haven’t, but have pathogenic effectors secreted into host cells? I suspect probably yes, but will be interesting to see... 16/18

There will be so much to explore - starting with these rules (some C-terminal degrons have unknown non-CRL E3s!), applying GPS methodology to discover more classes of degrons, understanding regulation of particular substrates, evolutionary pressures on proteome... 17/18

Overall an incredible paper I’m tremendously excited to re-read frequently and in greater depth, reshaping how I think about proteome regulation! #SystemsBiology #CellBiology #Ubiquitin 18/18