The ATP-powered gymnastics of TRiC/CCT: an asymmetric protein folding machine with a symmetric origin story

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Highlights

  • The eukaryotic chaperonin TRiC/CCT is a large hetero-oligomeric ring-shaped complex.

  • TRiC uses ATP to fold many essential cellular proteins within its central chamber.

  • TRiC undergoes a complex conformational cycle driven by ATP binding and hydrolysis.

  • Unfolded substrates bind TRiC through polyvalent subunit-specific contacts.

  • TRiC subunit diversity creates an asymmetric cycle essential for folding activity.

The eukaryotic chaperonin TRiC/CCT is a large hetero-oligomeric complex that plays an essential role assisting cellular protein folding and suppressing protein aggregation. It consists of two rings, and each composed of eight different subunits; non-native polypeptides bind and fold in an ATP-dependent manner within their central chamber. Here, we review recent advances in our understanding of TRiC structure and mechanism enabled by application of hybrid structural methods including the integration of cryo-electron microscopy with distance constraints from crosslinking mass spectrometry. These new insights are revealing how the different TRiC/CCT subunits create asymmetry in its ATP-driven conformational cycle and its interaction with non-native polypeptides, which ultimately underlie its unique ability to fold proteins that cannot be folded by other chaperones.

Introduction

All kingdoms of life contain a large double ring chaperonin complex that uses ATP binding and hydrolysis to bind and fold non-native polypeptides within its central chamber. Chaperonins have diverged into two evolutionarily distinct groups, differing substantially in their structure and mechanism. Prokaryotes and organelles of endosymbiotic origin contain Group I chaperonins, including the well characterized Escherichia coli chaperonin GroEL/GroES (reviewed in Ref. [1]). GroEL consists of two stacked homo-heptameric rings. In response to ATP-binding, the GroEL chamber is capped by the homo-heptameric GroES complex, which acts as a detachable lid required for substrate encapsulation and subsequent folding. Eukaryotes and archaea contain Group II chaperonins, which differ significantly from their prokaryotic counterparts in their architecture and mechanisms. First, the two rings of Group II chaperonins are generally eight membered and are stacked in a different inter-ring geometry than bacterial chaperonins. In addition, Group II chaperonins lack a GroES-like cofactor, using instead a flexible protrusion at the top of each subunit to create a built-in-lid that opens and closes in an ATP-dependent manner. Because of their distinct architecture, Group II chaperonins also have diverged in their conformational cycle, allosteric regulation, and in their substrate-recognition and folding mechanisms. In addition, while most prokaryotic chaperonins are homo-oligomeric group II chaperonins have undergone a progressive diversification of subunit composition. In archaea, chaperonins typically consist of 1–3 paralog subunits, while the eukaryotic chaperonin TRiC (TCP1-ring complex, also called CCT for cytosolic chaperonin containing TCP1) has eight different subunits per ring. The underlying causes and consequences of subunit diversification, which have long been enigmatic, are now beginning to emerge, as discussed in this review.

Section snippets

TRiC architecture and overall structure

TRiC/CCT is a 1 MDa complex composed of eight paralogous subunits that assemble into a double ring hexadecamer. Each subunit has a characteristic three domains architecture; an apical domain at the top of the ring which contains the substrate recognition site and the lid-forming loop, an equatorial domain that contains the ATP-binding site, and an intermediate domain which contains a conserved Aspartic acid required for ATP hydrolysis and communicates ATP cycling at the equatorial domain to

ATP regulation of the TRiC/CCT conformational cycle

Group II chaperonins use ATP binding and hydrolysis to alternate between an open state, where the lid forming segments are unstructured and the substrate binding sites in each subunit are accessible and a closed state, where the lid segments form a beta-stranded iris (schematic in Figure 2a). The conformational cycle, allosteric regulation, and ATP-driven structural changes are largely conserved between TRiC and its simpler archaeal homologues. Many of the early insights into the ATP cycle came

The cellular functions and substrates of TRiC/CCT

TRiC is essential for viability as many essential proteins, such as actin, tubulin, and cell cycle regulators, exhibit an obligate TRiC requirement to achieve the native state. TRiC binds co-translationally and post-translationally to ∼10% of the proteome [26,27]. Why TRiC selects these particular set of proteins as substrates and how it facilitates their folding are key unanswered questions for future research. TRiC substrates tend to be topologically complex with aggregation prone β -sheet

TRiC substrate interactions: how subunit diversity impacts polypeptide binding and folding

Substrate folding is coordinated with the ATPase cycle. In the open state TRiC binds unfolded polypeptides through the apical domains; ATP-induced lid formation releases the substrate to the closed chamber where folding is thought to occur. While a detailed understanding of how TRiC assists substrate folding has remained elusive, recent studies revealed a key role of subunit diversity in both substrate binding and folding.

The important question of substrate recognition, and how TRiC can

Perspectives

The wealth of new structural and mechanistic insight emerging in recent years has completely changed our view of TRiC/CCT. Initially thought to be a variation on the theme of GroEL-ES, this chaperonin has emerged as a highly asymmetrical and complex machine, where subunit diversification has created unique substrate binding and folding capabilities and a unique ATP-driven cycle unlike that of any other chaperone. This must clearly be linked to TRiC’s unique ability to fold eukaryotic proteins.

Conflict of interest statement

Nothing declared.

References and recommended reading

Papers of particular interest, published within the period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

Acknowledgements

Work in the Frydman lab is supported by N.I.H. (R01GM074074 to JF; F32GM103124 to DG). We thank members of the Frydman lab for discussions.

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