Constitutive expression of two apple (Malus x domestica Borkh.) homolog genes of LIKE HETEROCHROMATIN PROTEIN1 affects flowering time and whole-plant growth in transgenic Arabidopsis

Fruit trees, such as apple (Malus × domestica Borkh.), are woody perennial plants with a long juvenile phase. The biological analysis for the regulation of flowering time provides insights into the reduction of juvenile phase and the acceleration of breeding in fruit trees. In Arabidopsis, LIKE HETEROCHROMATIN PROTEIN1 (LHP1) is involved in epigenetic silencing of the target genes such as flowering genes. We isolated and characterized twin apple LHP1 homolog genes, MdLHP1a and MdLHP1b. These genes may have been generated as a result of ancient genome duplication. Although the putative MdLHP1 proteins showed lower similarity to any other known plant LHP1 homologs, a chromo domain, a chromo shadow domain, and the nuclear localization signal motifs were highly conserved among them. RT-PCR analysis showed that MdLHP1a and MdLHP1b were expressed constantly in developing shoot apices of apple trees throughout the growing season. Constitutive expression of MdLHP1a or MdLHP1b could compensate for the pleiotropic phenotype of lhp1/tfl2 mutant, suggesting that apple LHP1 homolog genes are involved in the regulation of flowering time and whole-plant growth. Based on these results, LHP1 homolog genes might have rapidly evolved among plant species, but the protein functions were conserved, at least between Arabidopsis and apple. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

phoR1, a gene encoding a new histidine protein kinase Myxococcus xanthus

A soil bacterium able to undergo multicellular development and a coordinated gliding in swarms, requires an accurate regulatory network of phosphorelay proteins. Inorganic phosphate is a limiting nutrient in soil and its importance in regulation is critical. As a step towards studying phosphate regulation and its influence in the developmental process in this bacterium, we screened a Myxococcus xanthus library for clones with phosphatase activity, and found four different ones. The deduced sequence of one of the cloned inserts is similar to that of the classic transmembrane histidine protein kinase of the sensor family of the two-component signal transduction systems with a high sequence similarity to the sensor kinase in the Pho regulon of Bacillus subtilis PhoR. This gene has been named phoR1 and its deduced amino acid sequence consists of 455 residues with a predicted molecular mass of 48.5 kDa. The M. xanthus PhoR1 deduced sequence contains all the characteristic histidine protein kinase motifs in the same order and with the same spacing. A hydropathy profile indicates two membrane-spanning segments located at the extreme N-terminus, according to the putative sensor role of this domain. A gene-disrupted mutant is unable to produce normal mature fruiting bodies and produces fewer spores. This revised version was published online in June 2006 with corrections to the Cover Date.     

Characterization of a Sorghum bicolor gene family encoding putative protein kinases with a high similarity to the yeast SNF1 protein kinase

C₄ photosynthesis is functionally dependent on metabolic interactions between mesophyll and bundle-sheath cells. Although the C₄ cycle is biochemically well understood many aspects of the regulation of enzyme activities, gene expression and cell differentiation are elusive.Protein kinases are likely involved in these regulatory processes providing links to hormonal, metabolic and developmental signal transduction pathways. We have identified several protein kinases that are differentially expressed in mesophyll and bundle-sheath cells of the C₄ plant Sorghum bicolor. Here we describe the characterization of two putative protein kinases that show high similarity to the SNF1/AMPK family of protein serine/threonine kinases. The mRNA of both kinases accumulates to much higher levels in mesophyll cells than in the bundle-sheath and can also be detected in root tissue. Complementation experiments with a snf1 mutant of Saccharomyces cerevisiae indicate that the S. bicolor protein kinase SNFL1 does not represent a functional homologue of the yeast SNF1 protein kinase.     

The yeast RNA1 protein, necessary for RNA processing, is homologous to the human ribonuclease/angiogenin inhibitor (RAI)

Summary Mutations in theRNA1 gene ofSaccharomyces cerevisiae, which encodes an essential cytosolic protein, affect the production and processing of all major classes of RNA. The mechanisms underlying these effects are not at all understood. Detailed comparative sequence analyses revealed that the RNA1 protein belongs to a superfamily, the members of which contain repetitive leucine-rich motifs (LRM). Within this superfamily RNA1 is most closely related to the ribonuclease/angiogenin inhibitor (RAI), which is a tightly binding inhibitor of ribonucleolytic activities in mammals. These results not only provide important clues to the structure, function and evolution of the RNAI protein, but also have intriguing implications for possible novel functions of RAI.     

ITN1, a novel gene encoding an ankyrin-repeat protein that affects the ABA-mediated production of reactive oxygen species and is involved in salt-stress tolerance in Arabidopsis thaliana

Salt stress and abscisic acid (ABA) induce accumulation of reactive oxygen species (ROS) in plant cells. ROS not only act as second messengers for the activation of salt-stress responses, but also have deleterious effects on plant growth due to their cytotoxicity. Therefore, the timing and degree of activation of ROS-producing or ROS-scavenging enzymes must be tightly regulated under salt-stress conditions. We identified a novel locus of Arabidopsis, designated itn1 (increased tolerance to NaCl1), whose disruption leads to increased salt-stress tolerance in vegetative tissues. ITN1 encodes a transmembrane protein with an ankyrin-repeat motif that has been implicated in diverse cellular processes such as signal transduction. Comparative microarray analysis between wild-type and the itn1 mutant revealed that induction of genes encoding the ROS-producing NADPH oxidases (RBOHC and RBOHD) under salt-stress conditions was suppressed in the mutant. This suppression was accompanied by a corresponding reduction in ROS accumulation. The ABA-induced expression of RBOHC and RBOHD was also suppressed in the mutant, as was the case for RD29A, an ABA-inducible marker gene. However, the ABA-induced expression of another marker gene, RD22, was not impaired in the mutant. These results suggest that the itn1 mutation partially impairs ABA signaling pathways, possibly leading to the reduction in ROS accumulation under salt-stress conditions. We discuss the possible mechanisms underlying the salt-tolerant phenotype of the itn1 mutant.     

A PMR2 tandem repeat with a modified C-terminus is located downstream from the KRS1 gene encoding lysyl-tRNA synthetase in Saccharomyces cerevisiae

Summary TheKRS1 gene encodes the cytoplasmic form ofSaccharomyces cerevisiae lysyl-tRNA synthetase. TheKRS1 locus has been characterized. The lysyl-tRNA synthetase gene is unique in the yeast genome. The gene is located on the right arm of chromosome IV and disruption of the open reading frame leads to lethality. These results contrast with the situation encountered inEscherichia coli where lysyl-tRNA synthetase is coded by two distinct genes,lysS andlysU, and further address the possible biological significance of this gene duplication. The nucleotide sequence of the 3′-flanking region has been established. It encodes a long open reading frame whose nucleotide and amino acid structures are almost identical toPMR2, a cluster of tandemly repeated genes coding for P-type ion pumps. The sequence alterations relative toPMR2 are mainly located at the C-terminus of the protein.     

The gene family encoding the Arabidopsis thaliana translation elongation factor EF-1 alpha: molecular cloning, characterization and expression

Summary The gene family encoding the Arabidopsis thaliana translation elongation factor (EF-1) was analysed. This family contains four genes (A1-A4) organized in a similar manner in different varieties of Arabidopsis. Based upon both their physical separation and a comparison of their sequences, it is suggested that the A4 gene and the A1, A2, and A3 genes constitute two distinct subfamilies within the genome. By introducing chimaeric gene constructs into Arabidopsis cells, we showed that the Al gene promoter mediates a transient expression about twofold higher than that obtained using the CaMV 35 S promoter. This expression depends on a 348 by DNA fragment extending from –982 to –634 by upstream of the initiation codon. This element contains a characteristic telomeric sequence (AACCCTAA) which is also found in the promoters of the A2 and A4 genes as well as in the promoters of the Drosophila EF-1 F1 gene and of several highly expressed plant genes.     

SOT1, a pentatricopeptide repeat protein with a small MutS‐related domain,is required for correct processing of plastid 23S–4.5S rRNA precursors in Arabidopsis thaliana

Ribosomal RNA processing is essential for plastid ribosome biogenesis, but is still poorly understood in higher plants. Here, we show that SUPPRESSOR OF THYLAKOID FORMATION1 (SOT1), a plastid‐localized pentatricopeptide repeat (PPR) protein with a small MutS‐related domain, is required for maturation of the 23S–4.5S rRNA dicistron. Loss of SOT1 function leads to slower chloroplast development, suppression of leaf variegation, and abnormal 23S and 4.5S processing. Predictions based on the PPR motif sequences identified the 5′ end of the 23S–4.5S rRNA dicistronic precursor as a putative SOT1 binding site. This was confirmed by electrophoretic mobility shift assay, and by loss of the abundant small RNA ‘footprint’ associated with this site in sot1 mutants. We found that more than half of the 23S–4.5S rRNA dicistrons in sot1 mutants contain eroded and/or unprocessed 5′ and 3′ ends, and that the endonucleolytic cleavage product normally released from the 5′ end of the precursor is absent in a sot1 null mutant. We postulate that SOT1 binding protects the 5′ extremity of the 23S–4.5S rRNA dicistron from exonucleolytic attack, and favours formation of the RNA structure that allows endonucleolytic processing of its 5′ and 3′ ends.     

In Arabidopsis thaliana, 1% of the genome codes for a novel protein family unique to plants

In the sequences released by the Arabidopsis Genome Initiative (AGI), we discovered a new and unexpectedly large family of orphan genes (127 genes by 01.08.99), named AtPCMP. The distribution of the AtPCMP genes on the five chromosomes suggests that the genome of Arabidopsis thaliana contains more than 200 genes of this family (1% of the whole genome). The deduced AtPCMP proteins are characterized by a surprising combinatorial organization of sequence motifs. The amino-terminal domain is made of a succession of three conserved motifs which generate an important diversity. These proteins are classified into three subfamilies based on the length and nature of their carboxy-terminal domain constituted by 1–6 motifs. All the motifs characterized have an important level of conservation in both sequence and spacing. A specific signature of this large family is defined. The presence of ESTs in databases and the detection of clones in A. thaliana cDNA libraries indicate that most of the genes of this family are expressed. The absence of similar sequences outside the plant kingdom strongly suggests that this unusually large orphan family is unique to plants. Features, the genesis, the potential function and the evolution of this plant combinatorial and modular protein family are discussed.