Hsp100 family chaperones of microorganisms and plants cooperate with the Hsp70/Hsp40/NEF system to resolubilize and reactivate stress-denatured proteins. In yeast this machinery also promotes propagation of prions by fragmenting prion polymers. We previously showed the bacterial Hsp100 machinery cooperates with the yeast Hsp40 Ydj1 to support yeast thermotolerance and with the yeast Hsp40 Sis1 to propagate [ PSI + ] prions. Here we find these Hsp40s similarly directed specific activities of the yeast Hsp104-based machinery. By assessing the ability of Ydj1-Sis1 hybrid proteins to complement Ydj1 and Sis1 functions we show their C-terminal substrate-binding domains determined distinctions in these and other cellular functions of Ydj1 and Sis1. We find propagation of [URE3] prions was acutely sensitive to alterations in Sis1 activity, while that of [ PIN + ] prions was less sensitive than [URE3], but more sensitive than [ PSI + ]. These findings support the ideas that overexpressing Ydj1 cures [URE3] by competing with Sis1 for interaction with the Hsp104-based disaggregation machine, and that different prions rely differently on activity of this machinery, which can explain the various ways they respond to alterations in chaperone function. Author Summary The cellular chaperone machinery helps proteins adopt and maintain native conformations and protects cells from stress. The yeast Hsp40s Ydj1 and Sis1 are co-chaperones that regulate Hsp70s, which are key components of many chaperone complexes. Both of these Hsp40s are crucial for growth and Ydj1 directs disaggregation activity of the Hsp100-based machinery to provide stress protection while Sis1 directs this activity to promote prion replication. Ydj1 also cures yeast of certain prions when overexpressed. We show that C-terminal domains that possess substrate-binding function of Ydj1 and Sis1 can mediate these and other functional distinctions and that the degree that prions depend on Sis1 activities could underlie differences in how they respond to alterations of chaperones. These findings support a view that Hsp40s regulate and specify functions of the chaperone machinery through substrate discrimination and cooperation with Hsp70. The disproportionate evolutionary expansion of Hsp40s (J-proteins) relative to their Hsp70 partners led to a proposal that this amplification allows increased regulation and fine-tuning of chaperone machines for increasingly complex processes. Our findings support this idea and provide insight into fundamental aspects of this cooperation.
【초록키워드】 Stress, complement, Proteins, activity, Replication, polymers, Protein, amplification, plants, microorganisms, plant, cooperation, thermotolerance, yeast, function, Bacterial, cellular, microorganism, Interaction, C-terminal domain, regulate, Propagation, specific activities, Support, Complexes, Regulation, growth, alteration, complex, domain, conformation, help, activities, functions, component, chaperones, Hsp70, chaperone, Cell, PROTECT, functional, less, promote, maintain, explain, overexpressed, respond, competing, C-terminal, increasingly, PSI, underlie, overexpressing, 【제목키워드】 function, machine,