eL15 and eL22 proteins and their functional environments during ribosome assembly in Saccharomyces cerevisiae
Ribosomes are ribonucleoprotein complexes in charge of protein translation. Translation is an essential process in all organisms that involves decoding the information contained in mRNAs (messenger RNAs) to generate proteins. Similarly, the process of ribosome biogenesis represents a critical pathway for all cells. Ribosomes are comprised of two different subunits, one large and one small, also named 60S and 40S, respectively, in eukaryotes, among them Saccharomyces cerevisiae, which is the model organism employed in this thesis. As ribosome biogenesis is well conserved among different eukaryotes, S. cerevisiae is a perfectly appropriate organism for this study. Ribosomes of S. cerevisiae are composed of 79 ribosomal proteins and 4 ribosomal RNAs (rRNAs), three of them in the large (25S, 5.8S y 5S) and one in the small ribosomal subunit (18S). The biogenesis of ribosomes is an extraordinarily complex process, in which in addition of the four rRNAs and the 79-80 ribosomal proteins, take part about 100 small nucleolar RNAs (snoRNAs) and more than 300 assembly protein factors, which are not present in the mature subunits and therefore are also known as trans-acting factors. The synthesis of ribosomes is a sequential process highly regulated and organized that starts in the nucleolus and orderly progresses through the nucleoplasm; almost mature ribosomal particles are exported to the cytoplasm, where the last steps of maturation take place, previously to their involvement in the translation process. The role of the different components participating in the biogenesis of the ribosomal subunits has been broadly studied through last decades. However, there are numerous assembly factors and some ribosomal proteins which still await characterization. This was the case for the ribosomal proteins eL22 and eL15 during the assembly of the 60S ribosomal subunit before the study performed in this doctoral thesis. With respect to the eL22 ribosomal protein, in this thesis, we have observed that, although, in laboratory conditions, this protein is not essential for survival, cell growth and ribosome production are mildly affected in strains lacking eL22. In this context, we have proved that the synthesis of ribosomes lacking of eL22 is partially blocked in the nucleus. Thus, our results indicate that the presence of eL22 is necessary for the correct processing of the 27S pre-rRNAs. Moreover, we have observed that eL22 performs an important role in the cytoplasm, where its presence is likely needed for the efficient recycling of assembly factors Arx1 and Alb1. With respect to the eL15 ribosomal protein, in this thesis, we have shown that this protein is essential for the production of mature ribosomes and, thus, for the cell survival and cell growth. Specifically, we have demonstrated that eL15 is necessary for the precise processing of the 27SA3 and 27SB pre-rRNAs in the nucleolus. This is due to the role of eL15 in the stabilization of early ribosomal particles which permits the stable assembly of proteins of its own functional cluster (eL8, eL36 y eL13) and association of numerous assembly proteins, known as A3- and B-factors, which are necessary for the processing of the 27SA3 and 27SB pre-rRNAs, respectively.
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