Overview

From January 2009 you can browse our genome-wide yeast CUTs abundance and localization results (Neil et al., Nature 2009)

We are interested in RNA metabolism and our results cover a large array of different cellular processes involving RNA, like the identification of novel transcripts, the assembly and the transport of ribonucleoprotein particles or the mechanisms of mRNA degradation in the cytoplasm. We use the yeast Saccharomyces cerevisiae as a model organism and try to develop novel genomic-scale tools for this model eukaryote. We spend our research time between different projects on several main topics:

Dynamics of assembly and export of the large ribosomal subunit in S. cerevisiae.

We identified novel factors involved in late 60S ribosomal subunit assembly steps, in the cytoplasm of yeast cells, like the conserved ATPase Drg1 (Pertschy et al., Mol Cell Biol 2007 - in collaboration with Helmut Bergler’s laboratory, Graz University, Austria). We also identified a network of functional and physical interactions between Kap121, Rei1, Arx1, Alb1 and Tif6 involved in the recycling of pre-ribosomal proteins that shuttle between the nucleus and the cytoplasm and required for proper 60S ribosomal subunit maturation (Lebreton et al., J Cell Biol 2006). The Jjj1 Hsp40 chaperone, a physical and genetical partner of Rei1, is also involved in the dissociation/recycling of the pre-60S shuttling factors. However, while the functions of Jjj1 and Rei1 partially overlap, Jjj1 is preferentially required for the release of Arx1 and Alb1 shuttling factors from the cytoplasmic pre-60S particles while Rei1 is preferentially involved in their recycling (Demoinet et al., RNA 2007). In the earlier steps of 60S biogenesis, we recently described the role of the nucleolar pre-60S WD40 repeats protein Mak11 in maintaining the levels of the ribosomal-like protein Rlp24 and its association with nascent ribosomal particles (Saveanu et al., Mol Cell Biol 2007). For the global description of the pathway, we used quantitative mass-spectrometry coupled with affinity purification of pre-ribosomal complexes from various mutant strains (Lebreton et al., Nucl Acids Res 2008)

Figure 2: A summary of sequential association to and dissociation from the nuclear and cytoplasmic pre-60S particles. The pre-60S factors studied in our group during the last four years are represented as colorized forms. Many others preribosomal factors (more than 80) are involved in the large ribosomal subunit biogenesis. They are symbolized by grey forms with a white contour. This Figure illustrates the complexity of the pathway and shows the existence of several classes of preribosomal factors. Some of them such as Drg1, Rei1 and Jjj1 are strictly cytoplasmic; others, such as Mak11 and Nsa2, are strictly nuclear and recycled within the nucleus; and finally others, such as Rlp24, Nog1, Arx1/Alb1 are shuttling factors. Their release from the preribosomal particles requires cytoplasmic pre-60S factors and their recycling involves karyopherins.

Currently, we try to understand the mechanistic bases of the observed phenotypes and genetic interactions and to identify novel proteins that are involved in eukaryotic ribosome maturation.

Mechanisms of mRNA degradation.

One of the major steps in mRNA degradation is the decapping reaction, usually triggered by a reduction in the size of the poly(A) tail. However, we could identify a novel mechanism of deadenylation-independent mRNA decapping and degradation that involves the protein Edc3 - a factor physically and genetically linked to the decapping machinery. The auto-regulation mechanism involved in the fine-tuning of the RPS28B mRNA levels depends on Edc3 (Badis et al., Mol Cell 2004). We currently analyze other candidate proteins that are involved either in general decapping or in the regulation of specific transcripts levels. Recently we validated new such candidates, identified by large-scale genetic screens in our lab (Decourty et al., PNAS 2008).

CUTs - Cryptic Unstable Transcripts.

CUTs cartoon
A few years ago, in collaboration with three other laboratories, we described the existence of a novel class of ubiquitous transcripts that are normally very efficiently degraded by the combined action of a poly-adenylation complex (TRAMP) and of the nuclear exosome (La Cava et al., Cell 2005; Wyers et al., Cell 2005). These transcripts, collectively known as CUTs - for Cryptic Unstable Transcripts, are very abundant in mutant strains that lack the degradation or poly-adenylation activities. The study of CUTs is likely to give hints to understand cellular processes as diverse as transcription initiation and termination, nucleo-cytoplasmic transport and translation. We have recently exhaustively mapped the position and abundance of yeast CUTs (Neil et al., Nature 2009). The association of CUTs with gene promoters strongly suggests that eucaryotic promoters are intrinsically bidirectional.

Genome-wide genetic screens by GIM (Genetic Interactions Mapping).

To explore the effects of combining mutations and thus, be able to better understand complex functional interactions in a cell, we developed a novel genetic screening method that combines the exhaustivity of the yeast systematic gene deletion collection, with a novel haploid specific selection marker and detection of the effects of combining gene mutations on growth by the use of microarrays (Decourty et al., PNAS 2008) [full text].

For further details, please look at the annual scientific activity summary for 2007. Our laboratory belongs to the “Genomes and Genetics” department.