\u3cem\u3eFRIGIDA LIKE 2\u3c/em\u3e Is a Functional Allele in Landsberg \u3cem\u3eerecta\u3c/em\u3e and Compensates for a Nonsense Allele of \u3cem\u3eFRIGIDA LIKE 1\u3c/em\u3e

The Landsberg erecta (Ler) accession of Arabidopsis (Arabidopsis thaliana) has a weak allele of the floral inhibitor FLOWERING LOCUS C (FLC). FLC-Ler is weakly up-regulated by the active San Feliu-2 (Sf2) allele of FRIGIDA (FRI-Sf2), resulting in a moderately late-flowering phenotype. By contrast, the Columbia (Col) allele of FLC is strongly up-regulated by FRI-Sf2, resulting in a very late-flowering phenotype. In Col, the FRI-related gene FRI LIKE 1 (FRL1) is required for FRI-mediated upregulation of FLC. It is shown here that in Ler, the FRL1-related gene FRI LIKE 2 (FRL2), but not FRL1, is required for FRI-mediated up-regulation of FLC. FRL1-Ler is shown to be a nonsense allele of FRL1 due to a naturally occurring premature stop codon in the middle of the conceptual protein sequence, suggesting that FRL1-Ler is nonfunctional. Compared to FRL2-Col, FRL2-Ler has two amino acid changes in the conceptual protein sequence. Plants homozygous for FRI-Sf2, FLC-Ler, FRL1-Ler, and FRL2-Col have no detectable FLC expression, resulting in an extremely early flowering phenotype. Transformation of a genomic fragment of FRL2-Ler, but not of FRL2-Col, into a recombinant inbred line derived from these plants restores both FRI-mediated up-regulation of FLC expression and a late-flowering phenotype, indicating that FRL2-Ler is the functional allele of FRL2. Taken together, these results suggest that in the two different Arabidopsis accessions Col and Ler, either FRL1 or FRL2, but not both, is functional and required for FRI-mediated up-regulation of FLC

RNA Levels and Activity of \u3cem\u3eFLOWERING LOCUS C\u3c/em\u3e are Modified in Mixed Genetic Backgrounds of \u3cem\u3eArabidopsis Thaliana\u3c/em\u3e

Flowering time and FLOWERING LOCUS C (FLC) RNA levels were analyzed in different accessions of Arabidopsis thaliana and in mixed genetic backgrounds resulting from crosses between accessions. Dominant alleles of FRIGIDA (FRI) promote accumulation of FLC RNA, which in turn promotes late flowering. Although the coding regions of sequenced FLC alleles are identical, some accessions have genetically weak alleles that do not promote late flowering in the presence of FRI. In this study, a new weak allele of FLC with open reading frame identity to previously sequenced alleles was isolated from a Niederzenz (Nd) accession. The FLC‐Nd allele accumulated less RNA in the presence of FRI than did the strong Columbia (Col) allele. The weak FLC‐Nd allele was semidominant in the mixed Nd/Col genetic background containing FRI, and a linear correlation between the level of FLC RNA and lateness of flowering was observed. However, late‐flowering transgressions with elevated levels of FLC RNA in the absence of FRI were also obtained from crosses between early‐flowering accessions Col and Nd. Moreover, compared to Nd, the weak Landsberg erecta (Ler) allele of FLC was recessive and not semidominant in the mixed Ler/Col genetic background. However, very early‐flowering transgressions lacking detectable FLC RNA were also obtained from crosses between FRI containing Col and Ler. The results indicate that modifier genes other than FRI influence the level and genetic activity of FLC RNA in different genetic backgrounds resulting from crosses between naturally occurring accessions of A. thaliana

Biennialism and Vernalization-Promoted Flowering in \u3cem\u3eHyoscyamus niger\u3c/em\u3e: a Comparison with \u3cem\u3eArabidopsis\u3c/em\u3e

There are genetic similarities between the biennial growth habit of Hyoscyamus niger (H. niger) and winter annual ecotypes of Arabidopsis thaliana, including the response to demethylating agents. One focus of our research group at Marquette is to determine whether FLOWERING LOCUS C (FLC) homologs or FLC-related MADS-box genes are involved in biennialism of H. niger. This review also summarizes our initial characterization of expression profiles of 4 groups of H. niger MADS-box genes. Our results suggest that B-class floral homeotic gene homologs of H. niger are differentially expressed in flowers of annual and vernalized, biennial plants

Cold Responsive \u3cem\u3eEARLI1\u3c/em\u3e Type HyPRPs Improve Freezing Survival of Yeast Cells and Form Higher Order Complexes in Plants

Plants have large families of proteins sharing a conserved eight-cysteine-motif (8CM) domain. The biological functions of these proteins are largely unknown. EARLI1 is a cold responsive Arabidopsis gene that encodes a hybrid proline-rich protein (HyPRP) with a three-domain architecture: a putative signal peptide at the N-terminus, a proline-rich domain (PRD) in the middle, and an 8CM domain at the C-terminus. We report here that yeast cells expressing different EARLI1 genes had significantly higher rates of freezing survival than empty-vector transformed controls. Arabidopsis plants with knocked down EARLI1 genes had an increased tendency for freezing-induced cellular damage. EARLI1-GFP Fluorescence in transgenic plants and immunoblot analyses using protoplasts suggested cell wall localization for EARLI1 proteins. Immunoblot analyses showed that EARLI1 proteins form higher order complexes in plants, and that the PRD is a soluble and the 8CM an insoluble protein domain. We propose that EARLI1 proteins have a bimodular architecture in which the PRD may interact with the cell wall and the 8CM domain with the plasma membrane to protect the cells during freezing stress

A Gene Expression Screen Identifies \u3cem\u3eEARLI1\u3c/em\u3e as a Novel Vernalization-Responsive Gene in \u3cem\u3eArabidopsis Thaliana\u3c/em\u3e

Vernalization promotes early flowering in late ecotypes of Arabidopsis thaliana. The mechanisms of vernalization are poorly understood. A subtractive hybridization approach was used to isolate vernalization-responsive genes from a late-flowering ecotype of Arabidopsis thaliana based on the premise that transcript levels of such genes would increase with cold treatment and remain high even after removal of the vernalization stimulus. EARLI1 is the first Arabidopsis gene shown to be stably activated by vernalization. The abundance of its RNA is progressively elevated by vernalization and remains high for at least 20 days at room temperature. The basal level of EARLI1 RNA is higher in early-flowering ecotypes, but is increased also after vernalization. Vernalization and subsequent growth in long-day photoperiods have an additive or synergistic effect on EARLI1 activation. EARLI1 RNA levels are also transiently induced by brief exposures to cold, but not to abscisic acid. EARLI1 is thus a novel vernalization-responsive gene in Arabidopsis thaliana that can be used to investigate vernalization-specific transcriptional regulation

A Highly Sensitive Plant Hybrid Protein Assay System Based on the \u3cem\u3eSpm\u3c/em\u3e Promoter and TnpA Protein for Detection and Analysis of Transcription Activation Domains

TnpA is a multifunctional DNA binding protein encoded by the maize Suppressor-mutator (Spm) transposable element. TnpA is required for transposition and is a repressor of the unmethylated Spm promoter. While analyzing protein domains using a yeast GAL4-based hybrid system in transiently transformed tobacco cells, we found that TnpA represses the \u3e10-fold transcriptional activation observed when the GAL4 DNA-binding domain is used alone. By contrast, compared to the backgroundless TnpA DNA-binding domain alone, 33- to 45-fold activation of the Spm promoter was observed when the VP16 activation domain was fused to it. TnpA-binding sites, but no TATA box, were required for transcription activation. Among the TnpA deletion derivatives tested, those retaining the coding sequences for the DNA-binding and protein dimerization domains gave the highest level of transcription activation when fused with the VP16 activation domain. The TnpA gene and TnpA-binding sites in the short Spm promoter therefore provide a novel, highly sensitive single-hybrid system for identifying and studying plant transcription activation domains in plant cells

Low Temperature Antioxidant Activity QTL Associate with Genomic Regions Involved in Physiological Cold Stress Tolerance Responses in Rice (\u3cem\u3eOryza sativa\u3c/em\u3e L.)

Boosting cold stress tolerance in crop plants can minimize stress-mediated yield losses. Asian rice (Oryza sativa L.), one of the most consumed cereal crops, originated from subtropical regions and is generally sensitive to low temperature environments. Within the two subspecies of rice, JAPONICA, and INDICA, the cold tolerance potential of its accessions is highly variable and depends on their genetic background. Yet, cold stress tolerance response mechanisms are complex and not well understood. This study utilized 370 accessions from the Rice Diversity Panel 1 (RDP1) to investigate and correlate four cold stress tolerance response phenotypes: membrane damage, seedling survivability, and catalase and anthocyanin antioxidative activity. Most JAPONICA accessions, and admixed accessions within JAPONICA, had lower membrane damage, higher antioxidative activity, and overall, higher seedling survivability compared to INDICA accessions. Genome-wide association study (GWAS) mapping was done using the four traits to find novel quantitative trait loci (QTL), and to validate and fine-map previously identified QTL. A total of 20 QTL associated to two or more traits were uncovered by our study. Gene Ontology (GO) term enrichment analyses satisfying four layers of filtering retrieved three potential pathways: signal transduction, maintenance of plasma membrane and cell wall integrity, and nucleic acids metabolism as general mechanisms of cold stress tolerance responses involving antioxidant activity

Concerted Formation of Macromolecular \u3cem\u3eSuppressor-mutator\u3c/em\u3e Transposition Complexes

Transposition of the maize Suppressor-mutator (Spm) transposon requires two element-encoded proteins, TnpA and TnpD. Although there are multiple TnpA binding sites near each element end, binding of TnpA to DNA is not cooperative, and the binding affinity is not markedly affected by the number of binding sites per DNA fragment. However, intermolecular complexes form cooperatively between DNA fragments with three or more TnpA binding sites. TnpD, itself not a sequence-specific DNA-binding protein, binds to TnpA and stabilizes the TnpA-DNA complex. The high redundancy of TnpA binding sites at both element ends and the protein-protein interactions between DNA-bound TnpA complexes and between these and TnpD imply a concerted transition of the element from a linear to a protein crosslinked transposition complex within a very narrow protein concentration range

\u3cem\u3eArabidopsis\u3c/em\u3e AZI1 Family Proteins Mediate Signal Mobilization for Systemic Defence Priming

Priming is a major mechanism behind the immunological \u27memory\u27 observed during two key plant systemic defences: systemic acquired resistance (SAR) and induced systemic resistance (ISR). Lipid-derived azelaic acid (AZA) is a mobile priming signal. Here, we show that the lipid transfer protein (LTP)-like AZI1 and its closest paralog EARLI1 are necessary for SAR, ISR and the systemic movement and uptake of AZA in Arabidopsis. Imaging and fractionation studies indicate that AZI1 and EARLI1 localize to expected places for lipid exchange/movement to occur. These are the ER/plasmodesmata, chloroplast outer envelopes and membrane contact sites between them. Furthermore, these LTP-like proteins form complexes and act at the site of SAR establishment. The plastid targeting of AZI1 and AZI1 paralogs occurs through a mechanism that may enable/facilitate their roles in signal mobilization

Plant Growth-Promoting Activity of Bacteria Isolated from Asian Rice (\u3cem\u3eOryza sativa\u3c/em\u3e L.) Depends on Rice Genotype

Asian rice is one of the most important crops because it is a staple food for almost half of the world’s population. To have production of rice keep pace with a growing world population, it is anticipated that the use of fertilizers will also need to increase, which may cause environmental damage through runoff impacts. An alternative strategy to increase crop yield is the use of plant growth-promoting bacteria. Thousands of microbial species can exist in association with plant roots and shoots, and some are critical to the plant’s survival. We isolated 140 bacteria from two distantly related rice accessions and investigated whether their impact on the growth of four different rice accessions. The bacterial isolates were screened for their ability to solubilize phosphate, a known plant growth-promoting characteristic, and 25 isolates were selected for further analysis. These 25 phosphate-solubilizing isolates were also able to produce other potentially growth-promoting factors. Five of the most promising bacterial isolates were chosen for whole-genome sequencing. Four of these bacteria, isolates related to Pseudomonas mosselii, a Microvirga sp., Paenibacillus rigui, and Paenibacillus graminis, improved root and shoot growth in a rice genotype-dependent manner. This indicates that while bacteria have several known plant growth-promoting functions, their effects on growth parameters are rice genotype dependent and suggest a close relationship between plants and their microbial partners