The N-terminus of FILIA Forms an Atypical KH Domain with a Unique Extension Involved in Interaction with RNA

FILIA is a member of the recently identified oocyte/embryo expressed gene family in eutherian mammals, which is characterized by containing an N-terminal atypical KH domain. Here we report the structure of the N-terminal fragment of FILIA (FILIA-N), which represents the first reported three-dimensional structure of a KH domain in the oocyte/embryo expressed gene family of proteins. The structure of FILIA-N revealed a unique N-terminal extension beyond the canonical KH region, which plays important roles in interaction with RNA. By co-incubation with the lysates of mice ovaries, FILIA and FILIA-N could sequester specific RNA components, supporting the critical roles of FILIA in regulation of RNA transcripts during mouse oogenesis and early embryogenesis

Effects of Interaction between Dual Shaking Tables and Specimen and Force Feedback Compensation Control

The shaking table array system is composed of multiple shaking tables for seismic response simulation tests of large-span spatial structures, bridge structures, slender structures such as pipeline and aqueduct, complex structures, and so on. In the process of testing with the multiple shaking tables, the interaction between the shaking tables and specimen affects the output accuracy of the shaking tables. The characteristics and rules of the dual shaking tables-specimen interaction effects on the system performance were analyzed in this paper. In order to improve the output accuracy of the dual shaking tables, force feedback compensation was introduced into three-variable control to reduce the interaction effects. However, the measurement errors of the force in the actuator and the acceleration of the shaking tables existed in the process of force feedback compensation. In order to verify the effectiveness of force feedback compensation for interaction between the dual shaking tables and specimen, the error influences on the system performance were simulated

Differential Movement Synchronous Tracking Control Strategy of Double-Shaking Table System Loading with Specimen

Multisupport, multidimension, and nonuniform excitation seismic experiments have new requirements for shaking table array system in synchronous tracking control. Therefore, this article proposed a novel synchronous tracking strategy, differential movement synchronous tracking control (DMSTC) strategy, for double-shaking table system while taking the interaction between shaking tables and specimen into consideration. DMSTC Simulink model of the double-shaking table with specimen was established and simulations were conducted in various conditions. The results demonstrate the viability of the proposed DMSTC in that the frequency bandwidth of the double-shaking tables is expanded from 3.27 Hz to 64.57 Hz, the maximum value of differential movement synchronous error is decreased from 1.682 mm to 0.482 mm, and the maximum tracking errors of the two shaking tables decrease from 1.138 mm to 0.044 mm and from 1.030 mm to 0.497 mm, respectively

Study on the interaction between shaking table and eccentric load

The control-structure interaction (CSI) between shaking table and eccentric load is one of the most important reasons causing the accuracy degradation of shaking table test. At present, the eccentric ratio (ER) of load and the coupling between actuators pose challenges to study the CSI. Thus, this paper establishes an analytical transfer function matrix of a shaking table and eccentric load. Based on the transfer function matrix, a comprehensive study is conducted to analyse the CSI effect under different eccentric ratio conditions. The analysis proves the influence of the CSI, and the CSI amplifies the actuator coupling more than 22 times at 20.00 Hz. Furthermore, a real-time CSI compensation strategy considering the actuator coupling is proposed. With the adopting of the proposed strategy, the coupling between two actuators is fully eliminated, and the correlation coefficients of ground motion records of two actuators are improved by 14.75% and 5.48%, respectively

Interaction of FILIA with RNA.

<p>(A) The binding of FILIA or FILIA-N with poly-C or poly-U RNA. In panel (a), purified recombinant proteins (FILIA, FILIA-N, FILIA-NΔ12, FILIA-NΔ28 and FILIA-NΔ39) were detected by anti-6xHis antibody. Proteins are labeled on top and molecular mass of marker bands are shown on right. In panel (b), purified recombinant proteins were sequestered by poly-C ribonucleotide homopolymers and detected by anti-6xHis antibody. In panel (c), purified recombinant proteins were sequestered by poly-U ribonucleotide homopolymers and detected by anti-6xHis antibody. (B) Nova2-KH3/RNA complex (PDB code 1EC6) was superimposed onto FILIA-N. Proteins are colored as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030209#pone-0030209-g001" target="_blank">Fig. 1(C)</a>, and the RNA molecule is shown in ribbon representation.</p

Intrinsic RNA pulled down by FILIA-N, FILIA-NΔ12 and FILIA.

<p>(A). FILIA-N and FILIA pull-down intrinsic RNA. Total RNA was purified from mice ovaries and incubated with FILIA-N, FILIA-NΔ12, FILIA, GST and Ni-NTA beads. The pull-down RNA and total RNA were separated by urea denatured PAGE and stained with SYBR Green II. The results were scanned with FLA 7000. (B). Repeating pull-down experiments. Pull-down lane 1 was RNA pulled down by FILIA-N from total ovarian RNA; Pull-down lane 2 was RNA pulled down by FILIA-N from the first residual RNA; Pull-down lane 3 was RNA pulled down by FILIA-N from the second residual RNA; After pull-down lane 1 was the first residual RNA; After pull-down lane 2 was the second residual RNA; After pull-down lane 3 was the third residual RNA.</p

Dimerization of FILIA-N.

<p>(A) Sedimentation velocity analysis of FILIA-N. The peak corresponds to a molecular mass of 29 KDa, indicating a dimer in solution. (B) Dimer in an asymmetric unit. Monomer1 is colored green from residues 40–117, and pale green from residues 2–39; monomer2 is colored magenta from residues 40–114, and light pink from residues 4–39 AA. (C) Interaction surface within dimer. Monomer1 of FILIA-N is shown represented by electrostatic surface potential, while monomer2 is shown in ribbon representation. The residues involved in the interaction between monomer1 and monomer2 are shown in stick representation and depicted in detail on the right picture. (D) Superposition of monomer1 and monomer2 of FILIA-N, colored as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030209#pone-0030209-g002" target="_blank">Fig. 2(B)</a>. (E) Protein concentrations plotted versus radius for an AUC equilibrium experiment. 25.5, 15.3 and 10.2 µ<i>M</i> FILIA-N were spun at 15,000, 22,500 and 28,500 rpm at 4°C. The solid line showed a fit of the data to a model of a dimer with a molecular weight of 29,814 (rmsd of 0.005). The molecular weight of the monomer calculated from its sequence is 14,625 Da. Residuals are shown at the top of the plot. A SDS-PAGE gel of FILIA-N used in this experiment is also shown.</p

X-ray crystallographic data and refinement statistics for FILIA-N.

Φ<p>ASU = asymmetric unit.</p><p>*Values in parentheses are for the highest resolution shell.</p>†<p>Rsym  =  Σ|I−<i>|/Σ<i>, where I is the observed intensity, and <i> is the average intensity of multiple observations of symmetry related reflections.</i></i></i></p><i><i><i>‡<p>R  =  Σhkl||Fobs|−|Fcalc||/Σhkl|Fobs|.</p>§<p>Rfree is calculated from 5% of the reflections excluded from refinement.</p></i></i></i

Features of the N-terminal extension.

<p>(A) Sequence alignment of FILIA-N from different mammals. Residues are colored and labeled as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030209#pone-0030209-g001" target="_blank">Fig. 1(C)</a>. (B) Interaction of the N-terminal fragment (AA 2–12) with other parts of FILIA-N. The dominant and conserved residues within different mammals are shown in stick representation and labeled.</p