Structure Of Yeast 90s Pre-ribosome

The ribosome biogenesis of eukaryotes is an extremely complex and sophisticated process involving about two hundred assembly factors.90S-preribosome,or small subunit processome,is the earliest gigantic complex formed during ribosome biogenesis,with about 5 megadaltons.Using cryo-EM single particle reconstruction,we solved three structures of the nearly complete yeast 90 S with the resolution from 8.7 ? to 4.5 ?,among which,the highest resolution,4.5 ?,was obtained from the Mtr4 knock-out sample.Combined with known crystal structures,homology models and chemical cross-linking coupled with mass spectrometry,we build the model of most protein factors,including 36 assembly factors and 19 small subunit proteins.Eight of the ten double helixes of 5' ETS RNA were constructed manually.The 18 S RNA model was modified on the basis of mature 18 S.This dissertation will describe the structure of 90 S in the sequence of 5'ETS protein(UTP-A,UTP-B,U3 snoRNP),5' ETS RNA,18 S 5' domain,central domain,3' major domain,3' minor domain,respectively.In our model of the 90 S,we observed the similar heterogeneous tetramers consisted of ?-helix bundles formed in UTP-A and UTP-B subcomplex.U3 RNA could form four complementary sites with 5' ETS RNA and 18 S RNA.The last complementary site was reported at first time.Each of four domains of 18 S RNA has partially formed in the 90 S,but the relative position among them was quite different from that in mature 40 S.We tested the impact of Dhr1 and Mtr4,two RNA helicases involved in 90 S processing,on the structure of 90 S.In the Dhr1 knock-out sample,we observed three states of 90 S.Apart from the normal 90 S,two other states were without 5' ETS RNA.Combined with the evidences of biochemistry and mass-spectrometry,we determined the function of Mtr4 in recruiting exosome components and degrading 5' ETS RNA.After the degradation of 5' ETS RNA,the 90 S could still maintain the overall shape.Surprisingly,the 5' ETS proteins were still bound to the 90 S,demonstrating that 5' ETS RNA degradation and disintegration of the 90 S are two relatively independent processes.The processing steps transiting the 90 S to the 40 S are more complex than existing model.