| Gene expression is a process that involves three major steps: transcription, RNA processing and translation. It is well known that the ribosome is a macromolecular machinery responsible of translation, but we still know very little about the regulation of ribosomal proteins constituting this machinery. They actually have been implicated in many different cellular processes such as cell adhesion, cell proliferation, and apoptosis. An abnormal regulation of ribosomal proteins can cause different diseases such as cancers, anemia, and ribosomopathies. Therefore, the expression of ribosomal proteins has to be carefully regulated in order to maintain homeostasis in the cell.;In yeast, there is evidence that ribosomal proteins could be involved in the regulation of their paralogs as well as their own. Previous work in the lab has shown that Rp130-1 negatively regulates its paralog rp130-2 in an intron dependant manner. My studies in yeast Schizoaccharomyces pombe were to clarify the mechanism by which Rp130-1 regulates rp130-2. In order to do so, we wanted to verify if Rp130-1 binds to rp130-2's intron. Results from gel shift assays showed that Rp130-1 did bind to the intron of rp130-2. I was to make different constructs of rp130-2 intron to find which part of the intron was responsible for the binding of Rp130-1. Unfortunately, we encountered many problems with the constructs which caused us to put this project on hold.;There are a few examples of auto-regulatory mechanism of ribosomal proteins. Our current work has led us to propose a new regulation pathway of the ribosomal protein S2 (rpS2). A stable cell line was generated allowing us to control the expression of a GFP-tagged version of rpS2 by adding doxycycline to the medium. Interestingly, a reduction of endogenous rpS2 can be observed when we induce the expression of GFP-rpS2. Yet, endogenous RPS2 mRNA levels remain unchanged in conditions of excess GFP-RPS2, suggesting a post-transcriptional mechanism of regulation. Our results suggest a mechanism that is independent of 5' and 3' untranslated regions. We also tested the possibility of an auto-regulatory pathway that involves rpS2 protein turnover. In this model, excess rpS2 would be ubiquitinated and sent to degradation. Our results show no evidence of uibquitination of rpS2 and no change in protein stability was observed in overexpression of rpS2. Further experiments led us to show that GFP-rpS2 could bind to RPS2 mRNA. We propose a model in which rpS2 protein can bind to its mRNA transcript and repress its translation.;Keywords: Ribosomal proteins, RPL30-1, RPL30-2 , RPS2, autoregulation, regulation of gene expression. |
