Cytosolic Hsp70 and Hsp40 chaperones enable the biogenesis of mitochondrial β-barrel proteins

Abstract

Mitochondrial β-barrel proteins are encoded in the nucleus, translated by cytosolic ribosomes, and then imported into the organelle. Recently, a detailed understanding of the intramitochondrial import pathway of β-barrel proteins was obtained. In contrast, it is still completely unclear how newly synthesized β-barrel proteins reach the mitochondrial surface in an import-competent conformation. In this study, we show that cytosolic Hsp70 chaperones and their Hsp40 cochaperones Ydj1 and Sis1 interact with newly synthesized β-barrel proteins. These interactions are highly relevant for proper biogenesis, as inhibiting the activity of the cytosolic Hsp70, preventing its docking to the mitochondrial receptor Tom70, or depleting both Ydj1 and Sis1 resulted in a significant reduction in the import of such substrates into mitochondria. Further experiments demonstrate that the interactions between β-barrel proteins and Hsp70 chaperones and their importance are conserved also in mammalian cells. Collectively, this study outlines a novel mechanism in the early events of the biogenesis of mitochondrial outer membrane β-barrel proteins.

Pex19 is involved in importing dually targeted tail‐anchored proteins to both mitochondria and peroxisomes

“…One is C90 that corresponds to the C‐terminal region of Hsp90 and binds to the import receptor Tom70 and thus interferes with Tom70‐chaperones interactions. The second is CBag, which corresponds to the C‐terminal domain of the cochaperone Bag and inhibits directly the function of Hsp70 . In accordance with the results of Figure , adding CBag to extract from control cells resulted in a reduction of about 20% in the import efficiency of Fis1‐TMC.…”

“…The fact that inhibitors of the Hsp40‐stimulated activity of Hsp70 hamper the in vitro integration of Fis1 to mitochondria suggests a direct involvement of Hsp70 and possibly of its Hsp40 co‐chaperones in the import process. In line with this possibility, very recently, we could demonstrate an involvement of cytosolic Hsp70 and Hsp40 chaperones in the early stages of the biogenesis of MOM β‐barrel proteins . Hsp40 co‐chaperones regulate the ATPase activity of Hsp70 chaperones and, thereby, influence its substrate binding cycle .…”

“…Then, import reactions were incubated at 25°C for 10 minutes. Alternatively, inhibition of Hsp70‐mediated import was achieved by adding the polypeptide inhibitors C90 and CBag to the import reactions according to published procedures …”

Interplay between the Ubiquitin Proteasome System and Mitochondria for Protein Homeostasis

“…The classical cytosolic Hsp70 and Hsp90 chaperones, their co-factors Sti1 and Ydj1, and ubiquilins (chaperone-like factors), associate with mitochondrial pre-proteins and also physically interact with the outer membrane components of the mitochondrial import channel (Deshaies et al, 1988;Young et al, 2003;Hoseini et al, 2016;Zanphorlin et al, 2016;Jores et al, 2018;Opaliñski et al, 2018). Supporting these physical interaction evidences, a genetic synthetic growth defect was observed between TOM and STI1, which encode protein forming an important scaffold, by simultaneously binding to Hsp70 and Hsp90 (Hoseini et al, 2016).…”

Inferring translational heterogeneity from ribosome profiling data

“…Moreover, while this tRNA modification is normally pervasive and applied to most corresponding cytoplasmic tRNAs (60), it may be less present in cases of localized translation known to facilitate the synthesis of proteins destined to the major organelles. Of note, mitochondrial proteins often fold in the cytosol and their export to the mitochondria often share components of the cytoplasmic protein quality control network (69)(70)(71), in S.cerevisiae for instance the Hsp70 chaperone SSA1 (72). In contrast, translocation to the ER is directly coupled to translation as ER-destined proteins usually cannot fold inside the cytoplasm.…”