Functional characterization of the chaperone network connected to the eucaryotic ribosome
Final Report Abstract
The mechanism of protein synthesis is highly conserved in all kingdoms of life. This does not only apply to the core translational machinery, but also to the network of ribosome-bound protein biogenesis factors (RPBs), which coordinate the early steps of protein biogenesis. For example, many of the RPBs of the yeast Saccharomyces cerevisiae possess close homologs in mammalian cell. However, on closer inspection there is significant differences with respect to the nature of RPBs, as well as with respect to the mechanism of RPB action. In the course of FOR 967 we have performed a comparative analysis of RPBs, which are conserved from yeast to man. First, we compared properties and function of the yeast dimeric ribosome-associated complex (RAC) with mRAC, its mammalian counterpart. Second, we analyzed the interplay of signal recognition particle (SRP) with nascent chain-associated complex (NAC) in yeast and the interplay of SRP with the argonaute protein Ago2 in mammalian cells. Yeast RAC and mammalian mRAC interact with ribosomes directly and affect the biogenesis of newly synthesized polypeptide chains. However, on a mechanistic level RAC and mRAC differ significantly. While RAC is a cochaperone partner of a specific ribosome-bound Hsp70 termed Ssb, mRAC is a cochaperone partner of cytosolic Hsc70 and Hsp70, which do not bind to ribosomes directly. We found that one function of mRAC in mammalian cells is to recruit specifically Hsp70 to translating ribosomes. Such a role is not performed by RAC, because Ssb binds to ribosomes independently of RAC. We found that one subunit of mRAC, termed MPP11/ZRF1, serves an additional, extra-ribosomal function in the nucleus of mammalian cells. Here, MPP11 cooperates with the histone H2A deubiquitinase USP21 in the regulation of polycomb-repressed genes. This role of MPP11 in transcriptional regulation strongly suggests that in mammalian cells the MPP11-subunit of the mRAC complex acquired novel, important functions during evolution. We found that a signal anchor (SA) sequence inside the ribosomal tunnel folds into α-helical conformation. Ribosomes, which contain such α-helical SA sequences, recruit SRP to the tunnel exit even before the SA sequence becomes accessible from the outside. We analyzed the interplay of yeast NAC and SRP on translating ribosomes and found that ribosomes carrying nascent chains containing SA sequences interact with NAC and SRP in a coordinated manner, giving rise to NAC•RNC•SRP complexes, which are targeted to the ER membrane. In the mammalian system we discovered a functional interplay between SRP and the argonaute protein Ago2, which is not conserved in Saccharomyces cerevisiae. In mammalian cells the two RPBs compete for ribosomes carrying secretory nascent chains. While SRP binds to ribosomes carrying nascent chains containing a wild type signal sequence with higher affinity, Ago2 preferentially binds if mutations within the signal sequence hamper the interaction with SRP. Ago2 binding then triggers the decay of the corresponding mutated mRNA molecule. By that Ago2 not only prevents the translation of proteins with defective targeting information but also prevents further rounds of translation of the defective mRNA.
Publications
- (2007) Association of protein biogenesis factors at the yeast ribosomal tunnel exit is affected by the translational status and nascent polypeptide sequence. J Biol Chem 282: 7809-7816
Raue U, Oellerer S, Rospert S
(See online at https://doi.org/10.1074/jbc.M611436200) - (2007) Functional characterization of the atypical Hsp70 subunit of yeast ribosome-associated complex. J Biol Chem 282: 33977-33984
Conz C, Otto H, Peisker K, Gautschi M, Wölfle T, Mayer MP, Rospert S
(See online at https://doi.org/10.1074/jbc.M706737200) - (2008) Ribosome-associated complex binds to ribosomes in close proximity of Rpl31 at the exit of the polypeptide tunnel in yeast. Mol Biol Cell 19: 5279-5288
Peisker K, Braun D, Wölfle T, Hentschel J, Fünfschilling U, Fischer G, Sickmann A, Rospert S
(See online at https://doi.org/10.1091/mbc.e08-06-0661) - (2009) A signal-anchor sequence stimulates signal recognition particle binding to ribosomes from inside the exit tunnel. Proc Natl Acad Sci USA 106: 1398-1403
Berndt U, Oellerer S, Zhang Y, Johnson AE, Rospert S
(See online at https://doi.org/10.1073/pnas.0808584106) - (2009) The Hsp70 homolog Ssb is essential for glucose sensing via the SNF1 kinase network. Genes Dev 23: 2102-2115
von Plehwe U, Berndt U, Conz C, Chiabudini M, Fitzke E, Sickmann A, Petersen A, Pfeifer D, Rospert S
(See online at https://doi.org/10.1101/gad.529409) - (2010) BIP modulates the affinity of its co-chaperone ERJ1 to ribosomes. J Biol Chem 285: 36427-35433
Benedix J, Lajoie P, Jaiswal H, Burgard C, Greiner M, Zimmermann R, Rospert S, Snapp EL, Dudek J
(See online at https://doi.org/10.1074/jbc.M110.143263) - (2010) The ribosome-bound Hsp70 homolog Ssb of Saccharomyces cerevisiae. Biochim Biophys Acta 1803: 662-672
Peisker K, Chiabudini M, Rospert S
(See online at https://doi.org/10.1016/j.bbamcr.2010.03.005) - (2010) Transcriptional activation of Polycomb-repressed genes by ZRF1. Nature 468: 1124-1128
Richly H, Rocha-Viegas L, Ribeiro JD, Demajo S, Gundem G, Lopez-Bigas N, Nakagawa T, Rospert S, Ito T, Di Croce L
(See online at https://doi.org/10.1038/nature09574) - (2011) The chaperone network connected to human ribosome-associated complex (mRAC). Mol Cell Biol 31: 1160-1173
Jaiswal H, Conz C, Otto H, Wölfle T, Fitzke E, Mayer MP, Rospert S
(See online at https://doi.org/10.1128/MCB.00986-10) - (2012) NAC functions as a modulator of SRP during the early steps of protein targeting to the ER. Mol Biol Cell 23: 3027-3040
Zhang Y, Berndt U, Gölz H, Tais A, Oellerer S, Wölfle T, Fitzke E, Rospert S
(See online at https://doi.org/10.1091/mbc.e12-02-0112) - (2013) Interaction of Nascent Chains with the Ribosomal Tunnel Proteins Rpl4, Rpl17, and Rpl39 of Saccharomyces cerevisiae. J Biol Chem 288: 33697-33707
Zhang Y, Wölfle T, Rospert S
(See online at https://doi.org/10.1074/jbc.M113.508283) - (2014) Inefficient SRP Interaction with a Nascent Chain Triggers a mRNA Quality Control Pathway. Cell 156: 146-157
Karamyshev AL, Patrick AE, Karamysheva ZN, Griesemer DS, Hudson H, Tjon-Kon-Sang S, Nilsson I, Otto H, Liu Q, Rospert S, Heijne G, Johnson AE, Thomas PJ
(See online at https://doi.org/10.1016/j.cell.2013.12.017)
