Research Article: Structure and evolution of the 4-helix bundle domain of Zuotin, a J-domain protein co-chaperone of Hsp70

Date Published: May 15, 2019

Publisher: Public Library of Science

Author(s): Om Kumar Shrestha, Ruchika Sharma, Bartlomiej Tomiczek, Woonghee Lee, Marco Tonelli, Gabriel Cornilescu, Milena Stolarska, Lukasz Nierzwicki, Jacek Czub, John L. Markley, Jaroslaw Marszalek, Szymon J. Ciesielski, Elizabeth A. Craig, Narayanaswamy Srinivasan.


The J-domain protein Zuotin is a multi-domain eukaryotic Hsp70 co-chaperone. Though it is primarily ribosome-associated, positioned at the exit of the 60S subunit tunnel where it promotes folding of nascent polypeptide chains, Zuotin also has off-ribosome functions. Domains of Zuotin needed for 60S association and interaction with Hsp70 are conserved in eukaryotes. However, whether the 4-helix bundle (4HB) domain is conserved remains an open question. We undertook evolutionary and structural approaches to clarify this issue. We found that the 4HB segment of human Zuotin also forms a bundle of 4 helices. The positive charge of Helix I, which in Saccharomyces cerevisiae is responsible for interaction with the 40S subunit, is particularly conserved. However, the C-termini of fungal and human 4HBs are not similar. In fungi the C-terminal segment forms a plug that folds back into the bundle; in S. cerevisiae it plays an important role in bundle stability and, off the ribosome, in transcriptional activation. In human, C-terminal helix IV of the 4HB is extended, protruding from the bundle. This extension serves as a linker to the regulatory SANT domains, which are present in animals, plants and protists, but not fungi. Further analysis of Zuotin sequences revealed that the plug likely arose as a result of genomic rearrangement upon SANT domain loss early in the fungal lineage. In the lineage leading to S. cerevisiae, the 4HB was subjected to positive selection with the plug becoming increasingly hydrophobic. Eventually, these hydrophobic plug residues were coopted for a novel regulatory function—activation of a recently emerged transcription factor, Pdr1. Our data suggests that Zuotin evolved off-ribosome functions twice—once involving SANT domains, then later in fungi, after SANT domain loss, by coopting the hydrophobic plug. Zuotin serves as an example of complex intertwining of molecular chaperone function and cell regulation.

Partial Text

J-domain proteins (JDPs) are obligate co-chaperones of Hsp70-based molecular chaperone systems [1, 2]. Such chaperone machineries are ubiquitous, functioning in diverse biological processes [3–5]. Zuotin is the JDP of the eukaryotic ribosome-associated Hsp70 system, which facilitates folding of nascent polypeptide chains [6, 7]. Zuotin binds near the exit site of the ribosome tunnel of the 60S subunit from which nascent chains emerge [8]. Like other JDP/Hsp70 systems, Zuotin’s J-domain is required for stimulation of the ATPase activity of its partner Hsp70, which facilitates efficient Hsp70 interaction with its substrate proteins [9, 10]. Zuotin is called Zuo1 in fungi and ZRF1 (Zuotin-related factor), Mpp11 or DNAJC2 in animals. In animals, plants and protists, but not fungi, Zuotin orthologs have a C-terminal, ~200 residue, segment containing SANT domains [11, 12]—domains typically involved in protein-protein interactions [13]. An early phylogenetic analysis of Zuotin established that fungal Zuotin evolved from a progenitor that had SANT domains, strongly suggesting that these domains were lost during fungal evolution [11]. Changes that occurred in Zuotin upon SANT domain loss is one focus of this report.

New information regarding the relationship between human and fungal Zuotin, and the evolution of Zuo1’s off-ribosome regulatory function in transcriptional activation emerged from the analyses reported here. Notably, the 4HB is conserved in most eukaryotes. However, while Helix I is particularly conserved, the C-terminal end of the 4HB differs substantially between animals and fungi—in the former it is a linker to the SANT domains, while in a subset of fungi related to S. cerevisiae, a regulator of transcriptional activity.




0 0 vote
Article Rating
Notify of
Inline Feedbacks
View all comments