CluMPS: Novel Method For Protein Aggregates Identification

Proteins on accumulation form clusters, these clusters play an important role in cell physiology- transmembrane receptor signalling, transcription factor activation and cellular stress response. The protein assembly enhances cell signal processing by increasing or decreasing biochemical reaction rates. However, overaccumulation of these proteins has been linked with numerous pathologies like Alzheimer’s, ALS and even cancer in some cases. 

Currently it is difficult to understand protein clustering as there has not been any direct method developed to observe it. The toxic proteins or soluble oligomers found in neurodegenerative diseases are roughly of 10-100 nm in size, the direct observation of such oligomers is not possible with simple fluorescent protein fusions as they do not concentrate enough fluorophores to emit fluorescence. Specialised techniques like fluorescence resonance energy transfer (FRET), fluorescence correlation spectroscopy and super-resolution imaging can be used to observe such clusters but the drawbacks like low signal-to-noise ratio, considerable optimisation for each target and long acquisition times are predominant.

CluMPS (clusters magnified by phase separation) is a fluorescent reporter strategy that detects protein clusters with high sensitivity. A CluMPS reporter consists of three domains:

(1) a binding domain for a target protein, 

(2) a fluorescent protein, and 

(3) an amplification domain that provides multivalency. 

CluMPS Is activated by forming visible blobs in the presence of target protein clusters as small as a few nanometers. In essence, CluMPS acts as an on/off switch, responding to the presence of protein clusters that it is programmed to detect. CluMPS reporter detects and visually amplifies even small clusters of a binding partner, generating large, quantifiable fluorescence condensates. Using computational modelling and optogenetic clustering it is demonstrated that CluMPS can detect small oligomers.

CluMPS identified small masses of pathogenic proteins when the GFP (Green Fluorescent Protein) fusions appeared dispersed. CluMPS can detect and track unmodified and tagged endogenous protein clusters, allowing for multiplexing of orthogonal probes within cells. CluMPS offers a simple way to examine higher-order protein assembly in its biological setting.

Reference:

Mumford TR, Rae D, Brackhahn E, Idris A, Gonzalez-Martinez D, Pal AA, Chung MC, Guan J, Rhoades E, Bugaj LJ. Simple visualization of submicroscopic protein clusters with a phase-separation-based fluorescent reporter. Cell Systems. 2024 Feb 5.

Image Source:

https://www.cell.com/cms/attachment/b463b852-127d-4b19-b367-bd2991b8bd84/fx1.jpg

https://scitechdaily.com/images/CluMPS-in-Action.jpg