Spatial mapping of splicing factor complexes involved in exon and intron definition

We have analyzed the interaction between serine/arginine-rich (SR) proteins and splicing components that recognize either the 5′ or 3′ splice site. Previously, these interactions have been extensively characterized biochemically and are critical for both intron and exon definition. We use fluorescence resonance energy transfer (FRET) microscopy to identify interactions of individual SR proteins with the U1 small nuclear ribonucleoprotein (snRNP)–associated 70-kD protein (U1 70K) and with the small subunit of the U2 snRNP auxiliary factor (U2AF35) in live-cell nuclei. We find that these interactions occur in the presence of RNA polymerase II inhibitors, demonstrating that they are not exclusively cotranscriptional. Using FRET imaging by means of fluorescence lifetime imaging microscopy (FLIM), we map these interactions to specific sites in the nucleus. The FLIM data also reveal a previously unknown interaction between HCC1, a factor related to U2AF65, with both subunits of U2AF. Spatial mapping using FLIM-FRET reveals differences in splicing factors interactions within complexes located in separate subnuclear domains

Identification of Nuclear and Cytoplasmic mRNA Targets for the Shuttling Protein SF2/ASF

The serine and arginine-rich protein family (SR proteins) are highly conserved regulators of pre-mRNA splicing. SF2/ASF, a prototype member of the SR protein family, is a multifunctional RNA binding protein with roles in pre-mRNA splicing, mRNA export and mRNA translation. These observations suggest the intriguing hypothesis that SF2/ASF may couple splicing and translation of specific mRNA targets in vivo. Unfortunately the paucity of endogenous mRNA targets for SF2/ASF has hindered testing of this hypothesis. Here, we identify endogenous mRNAs directly cross-linked to SF2/ASF in different subcellular compartments. Cross-Linking Immunoprecipitation (CLIP) captures the in situ specificity of protein-RNA interaction and allows for the simultaneous identification of endogenous RNA targets as well as the locations of binding sites within the RNA transcript. Using the CLIP method we identified 326 binding sites for SF2/ASF in RNA transcripts from 180 protein coding genes. A purine-rich consensus motif was identified in binding sites located within exon sequences but not introns. Furthermore, 72 binding sites were occupied by SF2/ASF in different sub-cellular fractions suggesting that these binding sites may influence the splicing or translational control of endogenous mRNA targets. We demonstrate that ectopic expression of SF2/ASF regulates the splicing and polysome association of transcripts derived from the SFRS1, PABC1, NETO2 and ENSA genes. Taken together the data presented here indicate that SF2/ASF has the capacity to co-regulate the nuclear and cytoplasmic processing of specific mRNAs and provide further evidence that the nuclear history of an mRNA ma

Holographic Duals of Quark Gluon Plasmas with Unquenched Flavors

We review the construction of gravitational solutions holographically dual to N=1 quiver gauge theories with dynamical flavor multiplets. We focus on the D3-D7 construction and consider the finite temperature, finite quark chemical potential case where there is a charged black hole in the dual solution. Discussed physical outputs of the model include its thermodynamics (with susceptibilities) and general hydrodynamic properties.Comment: Lecture presented at the Workshop "AdS/CFT and Novel Approaches to Hadron and Heavy Ion Physics", Kavli Institute of Theoretical Physics (KITPC), Beijing, China, 13 October 2010. Review article to be published in Communications in Theoretical Physics. 27 pages, 2 figure

Regulation of RUVBL1-RUVBL2 AAA-ATPases by the nonsense-mediated mRNA decay factor DHX34, as evidenced by Cryo-EM

Nonsense-mediated mRNA decay (NMD) is a surveillance pathway that degrades aberrant mRNAs and also regulates the expression of a wide range of physiological transcripts. RUVBL1 and RUVBL2 AAA-ATPases form an hetero-hexameric ring that is part of several macromolecular complexes such as INO80, SWR1, and R2TP. Interestingly, RUVBL1-RUVBL2 ATPase activity is required for NMD activation by an unknown mechanism. Here, we show that DHX34, an RNA helicase regulating NMD initiation, directly interacts with RUVBL1-RUVBL2 in vitro and in cells. Cryo-EM reveals that DHX34 induces extensive changes in the N-termini of every RUVBL2 subunit in the complex, stabilizing a conformation that does not bind nucleotide and thereby down-regulates ATP hydrolysis of the complex. Using ATPase-deficient mutants, we find that DHX34 acts exclusively on the RUVBL2 subunits. We propose a model, where DHX34 acts to couple RUVBL1-RUVBL2 ATPase activity to the assembly of factors required to initiate the NMD response.Spanish Ministry of Science and Innovation SAF2017-82632-P Andres Lopez-Perrote Carlos F Rodriguez Marina Serna Oscar Llorca. Autonomous Government of Madrid Y2018/BIO4747 Ana Gonzalez-Corpas Oscar Llorca. Autonomous Government of Madrid P2018/NMT4443 Ana Gonzalez-Corpas Oscar Llorca MRC Core funding Javier F Caceres Spanish Ministry of Science and Innovation BES-2015-071348 Carlos F Rodriguez The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.S

D3-D7 Quark-Gluon Plasmas at Finite Baryon Density

We present the string dual to SU(Nc) N=4 SYM, coupled to Nf massless fundamental flavors, at finite temperature and baryon density. The solution is determined by two dimensionless parameters, both depending on the 't Hooft coupling $\lambda_h$ at the scale set by the temperature T: $\epsilon_h\sim\lambda_h Nf/Nc$, weighting the backreaction of the flavor fields and $\tilde\delta\sim\lambda_h^{-1/2}nb/(Nf T^3)$, where $nb$ is the baryon density. For small values of these two parameters the solution is given analytically up to second order. We study the thermodynamics of the system in the canonical and grand-canonical ensembles. We then analyze the energy loss of partons moving through the plasma, computing the jet quenching parameter and studying its dependence on the baryon density. Finally, we analyze certain "optical" properties of the plasma. The whole setup is generalized to non abelian strongly coupled plasmas engineered on D3-D7 systems with D3-branes placed at the tip of a generic singular Calabi-Yau cone. In all the cases, fundamental matter fields are introduced by means of homogeneously smeared D7-branes and the flavor symmetry group is thus a product of abelian factors.Comment: 27 pages; v2: 29 pages, 1 (new) figure, new section 4.4 on optical properties, references, comments added; v3: eq. (3.19), comments and a reference adde

Nucleo-cytoplasmic shuttling of splicing factor SRSF1 is required for development and cilia function

Shuttling RNA-binding proteins coordinate nuclear and cytoplasmic steps of gene expression. The SR family proteins regulate RNA splicing in the nucleus and a subset of them, including SRSF1, shuttles between the nucleus and cytoplasm affecting post-splicing processes. However, the physiological significance of this remains unclear. Here, we used genome editing to knock-in a nuclear retention signal (NRS) in Srsf1 to create a mouse model harboring an SRSF1 protein that is retained exclusively in the nucleus. Srsf1NRS/NRS mutants displayed small body size, hydrocephalus, and immotile sperm, all traits associated with ciliary defects. We observed reduced translation of a subset of mRNAs and decreased abundance of proteins involved in multiciliogenesis, with disruption of ciliary ultrastructure and motility in cells and tissues derived from this mouse model. These results demonstrate that SRSF1 shuttling is used to reprogram gene expression networks in the context of high cellular demands, as observed here, during motile ciliogenesis

D3/D7 Quark-Gluon Plasma with Magnetically Induced Anisotropy

We study the effects of the temperature and of a magnetic field in the setup of an intersection of D3/D7 branes, where a large number of D7 branes is smeared in the transverse directions to allow for a perturbative solution in a backreaction parameter. The magnetic field sources an anisotropy in the plasma, and we investigate its physical consequences for the thermodynamics and energy loss of particles probing the system. In particular we comment on the stress-energy tensor of the plasma, the propagation of sound in the directions parallel and orthogonal to the magnetic field, the drag force of a quark moving through the medium and jet quenching.Comment: 29 pages + appendices, 5 figures. v2 Version to appear in JHEP, with minor revisions, references added and typos correcte