Molecular Mechanisms that Regulate the E3 Ubiquitin Ligase CHIP

Abstract

Neurodegenerative diseases are incurable diseases that are characterized by an accumulation of protein aggregates caused by a loss of protein quality control pathways. Among protein quality control pathways, molecular chaperones and the ubiquitin-proteosome system (UPS) play an important role in suppressing protein aggregation. Molecular chaperones are responsible for the refolding of misfolded proteins, whereas the UPS is important for targeting misfolded proteins for degradation. The E3 ubiquitin ligase Carboxy-terminus of Hsc70 Interacting Protein (CHIP) plays a unique role bridging the chaperone and ubiquitin pathways to protect against neurodegeneration. CHIP accomplishes this by binding molecular chaperones via an N-terminal tetratricopeptide repeat (TPR) domain and recruiting ubiquitin-conjugating enzymes via a C-terminal U-box domain. Together these interactions allow CHIP to prevent the accumulation of misfolded proteins, and thus is neuroprotective.To identify proteins and pathways that regulate CHIP function I utilized a proximity labeling dataset in our laboratory to identify proteins that are in proximity to CHIP. Using this dataset I found that TPR domain-containing peptidyl prolyl cis-trans isomerases (PPIases) were enriched in proximity to CHIP. Among these PPIases, I found that FK506 Binding Protein 51 (FKBP51) interacted with CHIP and induced unfolding of the CHIP TPR domain in a manner that was inhibited by the FKBP inhibitor FK506 and required prolines in CHIP. I further found that the interaction between FKBP51 and CHIP was mediated by affinities between the TPR domains in these proteins. In addition, I found that mutations in the TPR domain of CHIP causing SCA48 induce similar structural changes as the addition of FKBP51 does, and that the thermodynamic lability of the CHIP TPR domain is likely required for these induced structural changes Finally, I found that in addition to CHIP, FKBP51 can induce similar structural changes in additional TPR domain containing co-chaperones. Together, these data provide evidence of the first validated substrates of FKBP51 to my knowledge and identifies how SCA48-causing mutations in CHIP induce similar structural changes.