YB-1 promotes microtubule assembly in vitro through interaction with tubulin and microtubules

<p>Abstract</p> <p>Background</p> <p>YB-1 is a major regulator of gene expression in eukaryotic cells. In addition to its role in transcription, YB-1 plays a key role in translation and stabilization of mRNAs.</p> <p>Results</p> <p>We show here that YB-1 interacts with tubulin and microtubules and stimulates microtubule assembly <it>in vitro</it>. High resolution imaging via electron and atomic force microscopy revealed that microtubules assembled in the presence of YB-1 exhibited a normal single wall ultrastructure and indicated that YB-1 most probably coats the outer microtubule wall. Furthermore, we found that YB-1 also promotes the assembly of MAPs-tubulin and subtilisin-treated tubulin. Finally, we demonstrated that tubulin interferes with RNA:YB-1 complexes.</p> <p>Conclusion</p> <p>These results suggest that YB-1 may regulate microtubule assembly <it>in vivo </it>and that its interaction with tubulin may contribute to the control of mRNA translation.</p

XLF and APLF bind Ku80 at two remote sites to ensure DNA repair by non-homologous end joining

International audienceThe Ku70-Ku80 (Ku) heterodimer binds rapidly and tightly to the ends of DNA double-strand breaks and recruits factors of the non-homologous end-joining (NHEJ) repair pathway through molecular interactions that remain unclear. We have determined crystal structures of the Ku-binding motifs (KBM) of the NHEJ proteins APLF (A-KBM) and XLF (X-KBM) bound to a Ku-DNA complex. The two KBM motifs bind remote sites of the Ku80 alpha/beta domain. The X-KBM occupies an internal pocket formed by an unprecedented large outward rotation of the Ku80 alpha/beta domain. We observe independent recruitment of the APLF-interacting protein XRCC4 and of XLF to laser-irradiated sites via binding of A- and X-KBMs, respectively, to Ku80. Finally, we show that mutation of the X-KBM and A-KBM binding sites in Ku80 compromises both the efficiency and accuracy of end joining and cellular radiosensitivity. A- and X-KBMs may represent two initial anchor points to build the intricate interaction network required for NHEJ

Role of ATP in the single strand annealing activity of the "RecA core only" Sak4 of phage HK620

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Role of ATP in the single strand annealing activity of the "RecA core only" Sak4 of phage HK620

National audienc

The EcoRI−DNA Complex as a Model for Investigating Protein−DNA Interactions by Atomic Force Microscopy

International audienc

Multiple displacement amplification for complex mixtures of DNA fragments

International audienc

Method combining BAC film and positive staining for the characterization of DNA intermediates by dark-field electron microscopy

International audienceDNA intermediate structures are formed in all major pathways of DNA metabolism. Transmission electron microscopy(TEM) is a tool of choice to study their choreography and has led to major advances in the understanding of these mechanisms,particularly those of homologous recombination (HR) and replication. In this article, we describe specific TEM proceduresdedicated to the structural characterization of DNA intermediates formed during these processes. These particularDNA species contain single-stranded DNA regions and/or branched structures, which require controlling both the DNA moleculesspreading and their staining for subsequent visualization using dark-field imaging mode. Combining BAC (benzyl dimethylalkyl ammoniumchloride) film hyperphase with positive staining and dark-field TEM allows characterizing syntheticDNA substrates, joint molecules formed during not only in vitro assays mimicking HR, but also in vivo DNAintermediates

Smart DNA Vectors Based on Cyclodextrin Polymers: Compaction and Endosomal Release

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