Biology subjects

Ennifar, E.

Publications and source records attributed to Ennifar, E..

7 recordsLinked to original sources

bioRxiv2026

SigA forms amyloid fibrils and supports the competition of Shigella with the microbiota

Shigella sonnei is an enteropathogen that causes bacillary dysentery. During the first step of its virulence cycle, it must outcompete the resident microbiota to establish its own niche. Here we report that SigA, the sole SPATE (Serine Protease Autotransporter of Enterobacteriaceae) family member in S. sonnei, plays an indirect but central role in this process. A genome-wide analysis showed that the SPATE family includes SigA, Pic, SepA, and Sat. We demonstrated that SigA self-assembles into amyloid fibrils (F-SigA) independently of its protease activity. F-SigA remains associated with the S. sonnei surface in vitro and in vivo. Purified F-SigA fibrils have a diameter of 17.7 {+/-} 3.2 nm, and their amyloid organization was confirmed using specific markers and biochemical methods. F-SigA is secreted into the lumen in vivo and localizes to the surface of the colonic epithelium. We found that colicin E1 (ColE1) interacts with F-SigA amyloid fibrils, and that F-SigA-ColE1 complexes display antimicrobial activity that promotes S. sonnei competition with other bacteria. Because Pic, another Shigella SPATE, also forms amyloid fibrils, we anticipate that this virulence mechanism may be relevant across a wide range of Shigella strains and enterobacteria and may serve additional roles during the Shigella virulence cycle.

microbiology↗

bioRxiv2025

Interaction of lncRNA LENT with DHX36 regulates translation and suppresses autophagy in melanoma.

The melanocyte lineage determining Microphthalmia-associated transcription factor (MITF) drives proliferation and survival of melanocytic melanoma cells through regulation of both coding genes and long non-coding RNAs (LncRNAs). Here we characterize LINC00520 (hereafter called LncRNA ENhancer of Translation, LENT) regulated by MITF and strongly expressed in melanocytic melanoma cells. LENT is essential for proliferation and survival of cultured melanocytic melanoma cells and xenograft tumours. LENT interacts with the G4 quadruplex resolvase DHX36 and both associate with the ribosome in the 80S and light polysome fractions. LENT modulates DHX36 association with a collection of mRNAs regulating their engagement with polysomes and fine-tuning their subsequent translation. These mRNAs encode proteins involved in endoplasmic reticulum (ER) and mitochondrial homeostasis as well as autophagy. Consequently, LENT silencing leads to extensive autophagy and mitophagy, compromised oxidative metabolic capacity accompanied by an accumulation and mis-localization of mitochondrial proteins leading to proteotoxic stress and apoptosis. The LENT-DHX36 axis therefore fine-tunes translation of proteins involved in ER and mitochondrial homeostasis suppressing autophagy and promoting survival and proliferation of melanoma cells.

cancer biology↗

bioRxiv2024

Principles of ion binding to RNA inferred from the analysis of a 1.55 A resolution bacterial ribosome structure - Part I: Mg2+

The importance of Mg2+ ions for RNA structure and function can difficultly be overstated. Several attempts were made to establish a comprehensive Mg2+ binding site classification. However, such descriptions were hampered by poorly modelled ion binding sites. Recently, ribosome cryo-EM structures with resolutions < 2.0 [A] allowed better descriptions of site-bound ions. However, in a recent cryo-EM 1.55 [A] E. coli ribosome structure, incomplete ion assignments prevented a full understanding of their binding modes. We revisited this model to establish general binding principles applicable to any RNA of sufficient resolution. These principles rely on the 2.9 [A] distance separating two Mg2+-bound cis-water molecules. By applying these rules, we could assign all Mg2+ ions bound with 2 to 4 non-water oxygens. We also uncovered unanticipated motifs where up to five adjacent nucleotides wrap around a single ion. The formation of these complex motifs involves a hierarchical dehydration of the Mg2+ ions, a process that plays a significant role in ribosome biogenesis and in the folding of large RNAs. These binding principles enhance our understanding of the roles of ions in RNA structure and will help refining the solvation shell of different ribosomes and of RNAs with complex topologies.

molecular biology↗

bioRxiv2022

Specific conformational dynamics of the ATPase head domains and DNA exit gate mediate the Cohesin ATPase cycle

Cohesin is key to eukaryotic genome organization and acts throughout the cell cycle in an ATP- dependent manner. The molecular mechanisms underlying the Cohesin ATPase activity are poorly understood. Here, we have characterized distinct steps of the human Cohesin ATPase cycle and show that the SMC1A and SMC3 ATPase domains undergo specific but concerted structural rearrangements along this cycle. Specifically, while the proximal coiled coil of the SMC1A ATPase domain remains conformationally stable, that of SMC3 displays an intrinsic flexibility. The ATP-dependent formation of the heterodimeric SMC1A/SMC3 ATPase module (engaged state) favours this flexibility, while it is counteracted by binding of NIPBL and DNA (clamped state). Opening of the SMC3/RAD21 interface (open-engaged state) leads to a stiffening of the SMC3 proximal coiled coil that constricts, together with that of SMC1A, the DNA binding chamber of the ATPase module. Our results reveal that the plasticity of the ATP-dependent interface between the SMC1A and SMC3 ATPase domains enables the structural rearrangements occurring between the engaged, clamped and open-engaged states, while keeping the ATP gate shut.

biochemistry↗

bioRxiv2021

The RNA chaperone protein CspA stimulates translation during cold acclimation by promoting the progression of the ribosomes

CspA is an RNA binding protein expressed during cold-shock in Escherichia coli, capable of stimulating translation of several mRNAs - including its own - at low temperature. We used reconstituted translation systems to monitor the effects of CspA on the different steps of the translation process and probing experiments to analyze the interactions with its target mRNAs. We specifically focused on cspA mRNA which adopts a cold-induced secondary structure at temperatures below 20{degrees}C and a more closed conformation at 37{degrees}C. We show that at low temperature CspA specifically promotes the translation of the mRNA folded in the conformation less accessible to the ribosome (37{degrees}C form). CspA interacts with its mRNA without inducing large structural rearrangement, does not bind the ribosomal subunits and is not able to stimulate the formation of the translation initiation complexes. On the other hand, CspA promotes the progression of the ribosomes during translation of its mRNA at low temperature and this stimulation is mRNA structure-dependent. A similar structure-dependent mechanism may be responsible for the CspA- dependent translation stimulation observed with other probed mRNAs, for which the transition to the elongation phase is progressively facilitated during cold acclimation with the accumulation of CspA.

molecular biology↗

bioRxiv2020

Safe and easy evaluation of tmRNA-SmpB-mediated trans-translation in ESKAPE pathogenic bacteria

Bacteria cope with ribosome stalling thanks to trans-translation, a major quality control system of protein synthesis that is mediated by tmRNA, an hybrid RNA with properties of both a tRNA and an mRNA, and the small protein SmpB. Because trans-translation is absent in eukaryotes but necessary for bacterial fitness or survival, it is a promising target for the development of novel antibiotics. To facilitate screening of chemical libraries, various reliable in vitro and in vivo systems have been created for assessing trans-translational activity. However, none of these permits the safe and easy evaluation of trans-translation in pathogenic bacteria, which are obviously the ones we should be targeting. Based on green fluorescent protein (GFP) reassembly during active trans-translation, we have created a cell-free assay adapted to the rapid evaluation of trans-translation in ESKAPE bacteria, with 24 different possible combinations. It can be used for easy high-throughput screening of chemical compounds as well as for exploring the mechanism of trans-translation in these pathogens.

biochemistry↗

bioRxiv2020

Deflating the RNA Mg2+ bubble. Stereochemistry to the rescue!

Proper evaluation of the ionic structure of biomolecular systems remains challenging in X-ray and cryo-EM techniques but is essential for advancing our understanding of complex structure/activity/solvent relationships. However, numerous studies overestimate the number of Mg2+ in the deposited structures and underrate the importance of stereochemical rules to correctly assign these ions. Herein, we re-evaluate the PDBid 6QNR and 6SJ6 models of the ribosome ionic structure and establish that stereochemical principles should always be considered when evaluating ion binding features, even when K+ anomalous signals are available as it is the case for 6QNR. Assignment errors can result in misleading conceptions of the solvent structure of ribosomes and other RNA systems and should therefore be avoided. Our analysis resulted in a significant decrease of bound Mg2+ ions in the 6QNR structure, suggesting that K+ and not Mg2+ is the prevalent ion in the ribosome 1st solvation shell. We stress that the use of proper stereochemical guidelines is critical for deflating the current Mg2+ bubble witnessed in many ribosome and other RNA structures. Herewith, we would like to draw the attention of the researchers interested in the ionic structure of biomolecular systems on the importance and complementarity of stereochemistry and other ion identification techniques such as those pertaining to the detection of anomalous signals of transition metals and K+. We also stress that for the identification of lighter ions such as Mg2+, Na+, ..., stereochemistry coupled with high resolution structures remain the best currently available option.

biochemistry↗