Abstract
Ribosomal proteins are highly expressed, and the quality of ribosomal proteins must be rigorously controlled to build up a functional ribosome. Rpl43, ribosomal protein large subunit 43, is located nearby the E-site of ribosomes. In our previous study, we found that Puf6, Loc1, and Rpl43 form a trimeric complex in Saccharomyces cerevisiae. Rpl43 protein levels are under-accumulated in the absence of PUF6 or LOC1. However, why the loss of Puf6 or Loc1 decreased the protein levels of Rpl43 remained unclear. In the present study, we further dissected the connections among these three proteins and found that the processing defects of pre-ribosomal RNA in puf6Δ and loc1Δ are similar to those of the mutant with depletion of Rpl43. The stability of newly synthesized Rpl43 protein decreased slightly in puf6Δ and significantly in loc1Δ. We also found that Puf6 and Loc1 could interact with nascent Rpl43 co-translationally via the N-terminus of Rpl43. While the association and dissociation of Rpl43 with karyopherins did not depend on Puf6 and Loc1, Puf6 and Loc1 interacted with nascent Rpl43 in collaboration. While the N-terminus of Puf6 contained nuclear localization signals for transport, the PUF (Pumilio) domain was essential to interaction with Loc1, Rpl43, and 60S subunits. The C-terminus of Loc1 is more important for interaction with Puf6 and Rpl43. In this study, we found that Puf6 and Loc1 are the dedicated chaperones of ribosomal protein Rpl43 and also analyzed the potential interaction domains among the three proteins. Correct formation of the Puf6, Loc1, and Rpl43 ternary complex is required to properly proceed to the next step in 60S biogenesis.
Highlights
Ribosomes are massive complexes composed of ribosomal proteins and RNAs
Pre-90S and pre-60S ribosomal subunits were immunoprecipitated by transacting factors at different stages to examine whether they are loaded onto the corresponding ribosomal subunits at similar stages (Figure 1A)
Ribosomal protein large subunit 43 (Rpl43) is a small ribosomal protein with only 92 amino acids; it contains a conserved C4-type Zn finger domain made up of four beta-sheets at the center and two helices connected at the Nand C-termini
Summary
Ribosomes are massive complexes composed of ribosomal proteins and RNAs. Ribosome biogenesis is necessary for cell growth and proliferation and represents one of the major energy cost pathways of cells. Growing yeast cells generate about 2000 ribosomal subunits per minute [1]. About 60% of the total transcription in a cell is devoted to ribosomal RNA synthesis to support such large-scale synthesis. The multiple steps of ribosome biogenesis allow cells to precisely regulate the balance between supply and demand. Cells can regulate at the RNA transcriptional and ribosomal protein synthesis levels [1,2,3,4]
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