A synthetic growth switch based on controlled expression of RNA polymerase

International audienceThe ability to control growth is essential for fundamental studies of bacterial physiology and biotechnological applications. We have engineered an Escherichia coli strain in which the transcription of a key component of the gene expression machinery, RNA polymerase, is under the control of an inducible promoter. By changing the inducer concentration in the medium, we can adjust the RNA polymerase concentration and thereby switch bacterial growth between zero and the maximal growth rate supported by the medium. We show that our synthetic growth switch functions in a medium-independent and reversible way, and we provide evidence that the switching phenotype arises from the ultrasensitive response of the growth rate to the concentration of RNA polymerase. We present an application of the growth switch in which both the wild-type E. coli strain and our modified strain are endowed with the capacity to produce glycerol when growing on glucose. Cells in which growth has been switched off continue to be metabolically active and harness the energy gain to produce glycerol at a twofold higher yield than in cells with natural control of RNA polymerase expression. Remarkably, without any further optimization, the improved yield is close to the theoretical maximum computed from a flux balance model of E. coli metabolism. The proposed synthetic growth switch is a promising tool for gaining a better understanding of bacterial physiology and for applications in synthetic biology and biotechnology

Method for producing metabolites, peptides and recombinant proteins

Also published as EP3047031 (A1) US2016222428 (A1) WO2015036622 (A1)The present invention relates to a method for producing a molecule of interest in bacteria which is based on a reversible growth arrest of the bacteria at the cellular growth global control system level, thus allowing an improved yield of production of said molecule of interest

A genome-wide screen for identifying all regulators of a target gene

International audienceWe have developed a new screening methodology for identifying all genes that control the expression of a target gene through genetic or metabolic interactions. The screen combines mutant libraries with luciferase reporter constructs, whose expression can be monitored in vivo and over time in different environmental conditions. We apply the method to identify the genes that control the expression of the gene acs, encoding the acetyl coenzyme A synthetase, in Escherichia coli. We confirm most of the known genetic regulators, including CRP-cAMP, IHF and components of the phosphotransferase system. In addition, we identify new regulatory interactions, many of which involve metabolic intermediates or metabolic sensing, such as the genes pgi, pfkA, sucB and lpdA, encoding enzymes in glycolysis and the TCA cycle. Some of these novel interactions were validated by quantitative reverse transcriptase-polymerase chain reaction. More generally, we observe that a large number of mutants directly or indirectly influence acs expression, an effect confirmed for a second promoter, sdhC. The method is applicable to any promoter fused to a luminescent reporter gene in combination with a deletion mutant library

WellReader: a MATLAB program for the analysis of fluorescence and luminescence reporter gene data.

International audienceWellReader is available under a LGPL licence at http://prabi1.inrialpes.fr/trac/wellreader

Targeting the methyltransferase SETD8 impairs tumor cell survival and overcomes drug resistance independently of p53 status in multiple myeloma

International audienceAbstract Background Multiple myeloma (MM) is a malignancy of plasma cells that largely remains incurable. The search for new therapeutic targets is therefore essential. In addition to a wide panel of genetic mutations, epigenetic alterations also appear as important players in the development of this cancer, thereby offering the possibility to reveal novel approaches and targets for effective therapeutic intervention. Results Here, we show that a higher expression of the lysine methyltransferase SETD8, which is responsible for the mono-methylation of histone H4 at lysine 20, is an adverse prognosis factor associated with a poor outcome in two cohorts of newly diagnosed patients. Primary malignant plasma cells are particularly addicted to the activity of this epigenetic enzyme. Indeed, the inhibition of SETD8 by the chemical compound UNC-0379 and the subsequent decrease in histone H4 methylation at lysine 20 are highly toxic in MM cells compared to normal cells from the bone marrow microenvironment. At the molecular level, RNA sequencing and functional studies revealed that SETD8 inhibition induces a mature non-proliferating plasma cell signature and, as observed in other cancers, triggers an activation of the tumor suppressor p53, which together cause an impairment of myeloma cell proliferation and survival. However, a deadly level of replicative stress was also observed in p53-deficient myeloma cells treated with UNC-0379, indicating that the cytotoxicity associated with SETD8 inhibition is not necessarily dependent on p53 activation. Consistent with this, UNC-0379 triggers a p53-independent nucleolar stress characterized by nucleolin delocalization and reduction of nucleolar RNA synthesis. Finally, we showed that SETD8 inhibition is strongly synergistic with melphalan and may overcome resistance to this alkylating agent widely used in MM treatment. Conclusions Altogether, our data indicate that the up-regulation of the epigenetic enzyme SETD8 is associated with a poor outcome and the deregulation of major signaling pathways in MM. Moreover, we provide evidences that myeloma cells are dependent on SETD8 activity and its pharmacological inhibition synergizes with melphalan, which could be beneficial to improve MM treatment in high-risk patients whatever their status for p53

Roches ornées, roches dressées

Jean Abélanet peut être considéré comme le pionnier de l'archéologie actuelle sur les terres nord-catalanes. Ce rôle de précurseur dans la découverte de sites majeurs, mais aussi sa contribution savante à l'avancée des études préhistoriques, tant sur le mégalithisme en Pyrénées que sur l'art rupestre post-glaciaire en Europe occidentale, justifient l'hommage qui lui est rendu par la communauté des chercheurs. Cet hommage a pris la forme d'un colloque placé sous l'égide de l'Association Archéologique des Pyrénées-Orientales, dont il fut membre fondateur, et de l'Université de Perpignan, dont il fut le premier enseignant en Préhistoire. Sont rassemblées dans cet ouvrage les contributions de 74 auteurs et co-auteurs. Ces 576 pages, abondamment illustrées par près de 300 figures, abordent des sujets très divers qui reflètent les différents champs d'études balayés par son insatiable et humaniste curiosité. Une première partie, remontant aux sources des arts et des mythes, éclaire certains aspects de l'art rupestre et du mégalithisme, depuis leurs origines jusqu'à nos jours, à partir de recherches récentes menées dans l'Ancien monde, des terres australes d'Afrique jusqu'en Europe de l'Ouest. Le second thème, tout en laissant une large place à l'étude des arts et des traditions funéraires, rassemble des travaux pluridisciplinaires menés à l'orient des Pyrénées, travaux d'historiographie, de palynologie, de géologie, d'archéologie préhistorique et historique, d'histoire ou d'ethnologie