Thermospermine is Not a Minor Polyamine in the Plant Kingdom

Thermospermine is a structural isomer of spermine, which is one of the polyamines studied extensively in the past, and is produced from spermidine by the action of thermospermine synthase encoded by a gene named ACAULIS5 (ACL5) in plants. According to recent genome sequencing analyses, ACL5-like genes are widely distributed throughout the plant kingdom. In Arabidopsis, ACL5 is expressed specifically during xylem formation from procambial cells to differentiating xylem vessels. Loss-of-function mutants of ACL5 display overproliferation of xylem vessels along with severe dwarfism, suggesting that thermospermine plays a role in the repression of xylem differentiation. Studies of suppressor mutants of acl5 that recover the wild-type phenotype in the absence of thermospermine suggest that thermospermine acts on the translation of specific mRNAs containing upstream open reading frames (uORFs). Thermospermine is a novel type of plant growth regulators and may also serve in the control of wood biomass production

Improved antifouling properties of polyvinyl chloride blend membranes by novel phosphate based-zwitterionic polymer additive

To improve the antifouling properties of polymer membranes, a novel phosphate-based zwitterionic polymer (methacryloyloxyethylphosphorylcholine-co-poly(propylene glycol) methacrylate, i.e., MPC-PPGMA) is introduced into the polyvinyl chloride (PVC) membrane matrix. The solubility of this zwitterionic copolymer in common organic solvents and the compatibility between this copolymer and PVC are studied. A series of PVC blend membranes are prepared via non-solvent induced phase separation based on the good compatibility of the copolymer with PVC. Surface chemical compositions, wettability and porous structures of the membranes are characterized. Fluorescence-labeled proteins static adsorption and organic foulant filtration are conducted to evaluate the fouling resistance of the blend membranes. Antibiofouling properties of the membranes are also confirmed by the immersing method. Doping MPC-PPGMA in the casting solution can significantly improve the surface hydrophilicity and antifouling properties of the PVC blend membranes due to the strong hydration ability of the zwitterionic segments. The blend membranes show excellent and stable resistance to both organic fouling and biofouling. Overall, the novel zwitterionic copolymer MPC-PPGMA is a promising candidate to develop antifouling blend membranes due to its high effectiveness and long-term stability in the membranes.10 page(s

<i>sac53-d</i> is an allele of <i>RACK1A</i>.

<p><b>(A)</b> Exon/intron structure of <i>RACK1A</i>. Untranslated and coding regions are shown by white and black boxes, respectively. Arrowheads indicate the position of mutations. <b>(B)</b> Comparison of partial amino acid sequences of RACK1. <i>Arabidopsis thaliana</i> RACK1A (accession no. NP_173248.1), RACK1B (no. AEE32329), and RACK1C (no. AEE76051) are aligned with <i>Homo sapiens</i> RACK1 (no. AAH32006) and <i>Saccharomyces cerevisiae</i> RACK1 (no. DAA10013). Shaded boxes indicate conserved amino acids. Asterisks indicate a WD40 domain. <b>(C)</b> Gross morphology of 40-day-old plants. <b>(D)</b> Plant height of 40-day-old plants. Data show means ± SD (n = 6). Statistical significance was determined by a Student’s t test; significant difference (P < 0.05) from the wild type (Col-0) is indicated by different lowercase letters. <b>(E)</b> Gross morphology of 7-day-old etiolated seedlings.</p

Effect of <i>sac</i> mutations on gene expression.

<p><b>(A)</b> to <b>(D)</b> Relative mRNA levels of <i>ACL5</i><b>(A)</b>, <i>BUD2/AdoMetDC4</i><b>(B)</b>, <i>ATHB8</i><b>(C)</b>, and <i>VND7</i><b>(D)</b> were examined by quantitative RT-PCR. Seedlings were grown for 10 days in MS agar plates. mRNA levels were normalized to the <i>UBQ10</i> mRNA level and set to 1 in wild type (WT). Data show means ± SD (n = 3). Asterisks indicate significant differences (P < 0.05) from the wild type (Col-0).</p

<i>sac56-d</i> is an allele of <i>RPL4A</i>.

<p><b>(A)</b> Exon/intron structure of <i>RPL4A</i>. Untranslated and coding regions are shown by white and black boxes, respectively. Arrowheads indicate the position of mutations. Arrows indicate the position of primers used to amplify a <i>sac56-d</i> genomic fragment for transgenic recapitulation. <b>(B)</b> Comparison of partial amino acid sequences of RPL4. <i>Arabidopsis thaliana</i> RPL4A (accession no. AAP37854) and RPL4D (no. AED90529) are aligned with <i>Oryza sativa</i> RPL4 (no. NP_001059041), <i>Saccharomyces cerevisiae</i> RPL4A (no. NP_009587), <i>Homo sapiens</i> RPL4 (no. NP_000959), <i>Caenorhabditis elegans</i> RPL4 (no. CCD61249), <i>Drosophila melanogaster</i> RPL4 (no. AAG22173), and <i>Escherichia coli</i> RPL4 (no. ACI76839). Shaded boxes indicate conserved amino acids. <b>(C)</b> Gross morphology of 40-day-old plants. 56R #1 to #3 represent three <i>acl5–1</i> lines that were independently transformed with a <i>sac56-d</i> genomic fragment shown in <b>(A)</b>. <b>(D)</b> Plant height of 40-day-old plants. Data show means ± SD (n = 6). <i>rpl4a-2/sac56-d</i> indicates a genotype heterozygous for <i>rpl4a-2</i> and <i>sac56-d</i>. Statistical significance was determined by a Student’s t test; significant difference (P < 0.05) from the wild type (Col-0) is indicated by different lowercase letters.</p

Effect of thermospermine and <i>sac</i> mutations on the <i>SAC51</i> mRNA stability.

<p><b>(A)</b> Effect of <i>upf1–1</i> and <i>upf3–1</i> on <i>SAC51</i> expression. <b>(B)</b> Time-course assays of <i>SAC51</i> mRNA stability in wild type and <i>acl5–1</i>. Seedlings were treated with 0.6 mM cordycepin for indicated periods of time. In <i>acl5–1</i>, closed circles and open circles indicate data on seedlings treated with cordycepin and those pre-treated with 100 μM thermospermine for 1 h before addition of cordycepin, respectively. <b>(C)</b> Time-course assays of <i>SAC51</i> mRNA stability in <i>acl5–1 sac</i> double mutants. In <b>(A)</b> to <b>(C)</b>, seedlings were grown for 10 days in MS agar plates. Relative mRNA levels of <i>SAC51</i> were examined as shown in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0117309#pone.0117309.g005" target="_blank">Fig. 5</a> caption and set to 1 in wild type <b>(A)</b> and in each seedling before treatment (<b>[B]</b> and <b>[C]</b>).</p

Mutations in Ribosomal Proteins, RPL4 and RACK1, Suppress the Phenotype of a Thermospermine-Deficient Mutant of <i>Arabidopsis thaliana</i>

<div><p>Thermospermine acts in negative regulation of xylem differentiation and its deficient mutant of <i>Arabidopsis thaliana</i>, <i>acaulis5</i> (<i>acl5</i>), shows excessive xylem formation and severe dwarfism. Studies of two dominant suppressors of <i>acl5</i>, <i>sac51-d</i> and <i>sac52-d</i>, have revealed that <i>SAC51</i> and <i>SAC52</i> encode a transcription factor and a ribosomal protein L10 (RPL10), respectively, and these mutations enhance translation of the <i>SAC51</i> mRNA, which contains conserved upstream open reading frames in the 5’ leader. Here we report identification of <i>SAC53</i> and <i>SAC56</i> responsible for additional suppressors of <i>acl5. sac53-d</i> is a semi-dominant allele of the gene encoding a receptor for activated C kinase 1 (RACK1) homolog, a component of the 40S ribosomal subunit. <i>sac56-d</i> represents a semi-dominant allele of the gene for RPL4. We show that the GUS reporter activity driven by the CaMV 35S promoter plus the <i>SAC51</i> 5’ leader is reduced in <i>acl5</i> and restored by <i>sac52-d</i>, <i>sac53-d</i>, and <i>sac56-d</i> as well as thermospermine. Furthermore, the <i>SAC51</i> mRNA, which may be a target of nonsense-mediated mRNA decay, was found to be stabilized in these ribosomal mutants and by thermospermine. These ribosomal proteins are suggested to act in the control of uORF-mediated translation repression of <i>SAC51</i>, which is derepressed by thermospermine.</p></div

Effect of <i>sac</i> mutations on the <i>SAC51</i> 5’-<i>GUS</i> expression.

<p><b>(A)</b> Effect of <i>sac</i> mutations on <i>SAC51</i> expression. Relative mRNA levels of <i>SAC51</i> in 10-day-old seedlings were examined as shown in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0117309#pone.0117309.g005" target="_blank">Fig. 5</a> caption. An asterisk indicates a significant difference (P < 0.05) from the wild type (L<i>er</i>). <b>(B)</b> Effect of thermospermine on CaMV 35S promoter-driven <i>SAC51</i> 5’-<i>GUS</i> expression. Seedlings carrying the <i>SAC51</i> 5’ leader-<i>GUS</i> gene fused with the CaMV 35S promoter were grown for 10 days in MS agar plates and incubated for 24 h in MS solutions without (black bars) or with 100 μM thermospermine (white bars). Data show means ± SD (n = 3). Significant difference (P < 0.05) from the wild type (L<i>er</i>) is indicated by different lowercase letters. <b>(C)</b> to <b>(G)</b> GUS staining of wild-type <b>(C)</b>, <i>acl5–1</i><b>(D)</b>, <i>acl5–1 sac52-d</i><b>(E)</b>, <i>acl5–1 sac53-d</i><b>(F)</b>, and <i>acl5–1 sac56-d</i><b>(G)</b> seedlings carrying the <i>SAC51</i> 5’ leader-<i>GUS</i> gene fused with the CaMV 35S promoter. Seedlings were grown for 3 days in MS agar plates.</p