Crystal Structures of Lsm3, Lsm4 and Lsm5/6/7 from <em>Schizosaccharomyces pombe</em>

<div><p>Sm-like (Lsm) proteins are ubiquitous and function in many aspects of RNA metabolism, including pre-mRNA splicing, nuclear RNA processing, mRNA decay and miRNA biogenesis. Here three crystal structures including Lsm3, Lsm4 and Lsm5/6/7 sub-complex from <em>S. pombe</em> are reported. These structures show that all the five individual Lsm subunits share a conserved Sm fold, and Lsm3, Lsm4, and Lsm5/6/7 form a heptamer, a trimer and a hexamer within the crystal lattice, respectively. Analytical ultracentrifugation indicates that Lsm3 and Lsm5/6/7 sub-complex exist in solution as a heptamer and a hexamer, respectively while Lsm4 undergoes a dynamic equilibrium between monomer and trimer in solution. RNA binding assays show that Lsm2/3 and Lsm5/6/7 bind to oligo(U) whereas no RNA binding is observed for Lsm3 and Lsm4. Analysis of the inter-subunit interactions in Lsm5/6/7 reveals the organization order among Lsm5, Lsm6 and Lsm7.</p> </div

Details of sedimentation velocity data analysis.

a<p>S20, w is the sedimentation coefficient with the parameter being corrected to 20.0°C and the density of water.</p>b<p>RMSD is the root mean square deviation from SEDFIT program fitting.</p>c<p>MW is molecular weight in Dalton.</p

Electrostatic potential of Lsm and Sm proteins viewed from the helix and loop faces.

<p><i>Archaeoglobus fulgidus</i> Sm1 protein (AF-Sm1) (PDB code 1I4K); <i>Pyrococcus abyssi</i> Sm1 (PA-Sm1) (PDB code 1M8V); <i>Homo sapiens</i> Sm complex (HS-Sm) (PDB code 2Y9A); <i>Staphylococcus aureus</i> Hfq (SA-Hfq) (PDB code 1KQ1). The figure was generated with GRASP2.</p

Sedimentation velocity study of Lsm proteins in solution at 0.75 mg/ml.

<p>The Lsm proteins including SpLsm3, SpLsm5/6/7, SpLsm4N and ScLsm3 were analyzed by sedimentation velocity and fitted based on the c(M) and c(S) size-distribution functions. The corresponding molecular weights obtained from the c(M) size-distribution function for SpLsm3, SpLsm5/6/7, SpLsm4N and ScLsm3 were 75.0 kD, 62.6 kD, 80.3 kD and 11.8 kD, respectively.</p

Subunit interfaces in SpLsm5/6/7, SpLsm3 and ScLsm3.

<p>Residues involved in interface interaction are shown in stick model. All subunit interfaces are shown in similar orientations. (A) Stereo view of the interface between SpLsm5 and SpLsm6. (B) Stereo view of the interface between SpLsm5 and SpLsm7. (C) Stereo view of the interface between SpLsm6 and SpLsm7. (D) Stereo view of subunit interfaces of SpLsm3. One subunit is colored as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036768#pone-0036768-g001" target="_blank">Figure 1</a> while the other subunit is shown in grey. (E) Stereo view of subunit interfaces of ScLsm3 (PDB code 3BW1). The coloring scheme of the two subunits is as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036768#pone-0036768-g007" target="_blank">Figure 7D</a>.</p

Data collection and refinement statistics.

a<p>Values in the highest resolution shell are shown in parentheses.</p>b<p><i>R_work</i>  = <i>Σ||Fobs| - |Fcalc||/Σ|Fobs|</i>. <i>R_free</i> is calculated identically with 5% of randomly chosen reflections omitted from the refinement.</p>c<p>Fractions of residues in most favoured/allowed/generously allowed/disallowed regions of the Ramachandran plot were calculated according to PROCHECK.</p

Overall architectures of SpLsm3, SpLsm4N, SpLsm5, SpLsm6 and SpLsm7.

<p>The monomeric structures of SpLsm3, SpLsm4N, SpLsm5, SpLsm6 and SpLsm7 are shown in cartoon with similar orientations. Each monomer is colored as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036768#pone-0036768-g001" target="_blank">Figure 1</a>. The disordered loop 4 region in SpLsm3 and SpLsm7 is shown as dotted lines.</p

Analysis of U₁₅ binding activity of SpLsm2/3, SpLsm3, SpLsm5/6/7 and SpLsm4N.

<p>(A) Sensorgrams of surface plasmon resonance analysis using 5′-end biotin-labeled U15. Fluorescence anisotropy analysis using 5′-end FAM-labeled U15 showed that the fitted K_d value of SpLsm2/3 is 4.0 ± 0.5 µM (B) and the fitted K_d value of SpLsm5/6/7 is 52.5 ± 10.0 µM while no K_d values could be determined for SpLsm3 and SpLsm4N (C).</p

Oligomeric state of studied Lsm proteins determined from three different concentrations.

a<p>MW is molecular weight in Dalton.</p

The underlying processes of a soil mite metacommunity on a small scale

<div><p>Metacommunity theory provides an understanding of how ecological processes regulate local community assemblies. However, few field studies have evaluated the underlying mechanisms of a metacommunity on a small scale through revealing the relative roles of spatial and environmental filtering in structuring local community composition. Based on a spatially explicit sampling design in 2012 and 2013, this study aims to evaluate the underlying processes of a soil mite metacommunity on a small spatial scale (50 m) in a temperate deciduous forest located at the Maoershan Ecosystem Research Station, Northeast China. Moran’s eigenvector maps (MEMs) were used to model independent spatial variables. The relative importance of spatial (including trend variables, i.e., geographical coordinates, and broad- and fine-scale spatial variables) and environmental factors in driving the soil mite metacommunity was determined by variation partitioning. Mantel and partial Mantel tests and a redundancy analysis (RDA) were also used to identify the relative contributions of spatial and environmental variables. The results of variation partitioning suggested that the relatively large and significant variance was a result of spatial variables (including broad- and fine-scale spatial variables and trend), indicating the importance of dispersal limitation and autocorrelation processes. The significant contribution of environmental variables was detected in 2012 based on a partial Mantel test, and soil moisture and soil organic matter were especially important for the soil mite metacommunity composition in both years. The study suggested that the soil mite metacommunity was primarily regulated by dispersal limitation due to broad-scale and neutral biotic processes at a fine-scale and that environmental filtering might be of subordinate importance. In conclusion, a combination of metacommunity perspectives between neutral and species sorting theories was suggested to be important in the observed structure of the soil mite metacommunity at the studied small scale.</p></div