Localization,ion channel regulation,and genetic interactions during abscisic acid signaling of the nuclear mRNA cap-binding protein,ABH1

Abscisic acid (ABA) regulates developmental processes and abiotic stress responses in plants. We recently characterized a new Arabidopsis mutant, abh1, which shows ABA-hypersensitive regulation of seed germination, stomatal closing, and cytosolic calcium increases in guard cells (V. Hugouvieux, J.M. Kwak, J.I. Schroeder [2001] Cell 106: 477-487). ABH1 encodes the large subunit of a dimeric Arabidopsis mRNA cap-binding complex and in expression profiling experiments was shown to affect mRNA levels of a subset of genes. Here, we show that the dimeric ABH1 and AtCBP20 subunits are ubiquitously expressed. Whole-plant growth phenotypes of abh1 are described and properties of ABH1 in guard cells are further analyzed. Complemented abh1 lines expressing a green fluorescent protein-ABH1 fusion protein demonstrate that ABH1 mainly localizes in guard cell nuclei. Stomatal apertures were smaller in abh1 compared with wild type (WT) when plants were grown at 40% humidity, and similar at 95% humidity. Correlated with stomatal apertures from plants grown at 40% humidity, slow anion channel currents were enhanced and inward potassium channel currents were decreased in abh1 guard cells compared with WT. Gas exchange measurements showed similar primary humidity responses in abh1 and WT, which together with results from abh1/abi1-1 double-mutant analyses suggest that abh1 shows enhanced sensitivity to endogenous ABA. Double-mutant analyses of the ABA-hypersensitive signaling mutants, era1-2 and abh1, showed complex genetic interactions, suggesting that ABH1 and ERA1 do not modulate the same negative regulator in ABA signaling. Mutations in the RNA-binding protein sad1 showed hypersensitive ABA-induced stomatal closing, whereas hyl1 did not affect this response. These data provide evidence for the model that the mRNA-processing proteins ABH1 and SAD1 function as negative regulators in guard cell ABA signaling.     

NUCLEAR PORE ANCHOR, the Arabidopsis homolog of Tpr/Mlp1/Mlp2/megator, is involved in mRNA export and SUMO homeostasis and affects diverse aspects of plant development

Vertebrate Tpr and its yeast homologs Mlp1/Mlp2, long coiled-coil proteins of nuclear pore inner basket filaments, are involved in mRNA export, telomere organization, spindle pole assembly, and unspliced RNA retention. We identified Arabidopsis thaliana NUCLEAR PORE ANCHOR (NUA) encoding a 237-kD protein with similarity to Tpr. NUA is located at the inner surface of the nuclear envelope in interphase and in the vicinity of the spindle in prometaphase. Four T-DNA insertion lines were characterized, which comprise an allelic series of increasing severity for several correlating phenotypes, such as early flowering under short days and long days, increased abundance of SUMO conjugates, altered expression of several flowering regulators, and nuclear accumulation of poly(A)+ RNA. nua mutants phenocopy mutants of EARLY IN SHORT DAYS4 (ESD4), an Arabidopsis SUMO protease concentrated at the nuclear periphery. nua esd4 double mutants resemble nua and esd4 single mutants, suggesting that the two proteins act in the same pathway or complex, supported by yeast two-hybrid interaction. Our data indicate that NUA is a component of nuclear pore-associated steps of sumoylation and mRNA export in plants and that defects in these processes affect the signaling events of flowering time regulation and additional developmental processes.     

Flowering time and elevated atmospheric CO2

Flowering is a critical milestone in the life cycle of plants, and changes in the timing of flowering may alter processes at the species, community and ecosystem levels. Therefore understanding flowering-time responses to global change drivers, such as elevated atmospheric carbon dioxide concentrations, [CO(2)], is necessary to predict the impacts of global change on natural and agricultural ecosystems. Here we summarize the results of 60 studies reporting flowering-time responses (defined as the time to first visible flower) of both crop and wild species at elevated [CO(2)]. These studies suggest that elevated [CO(2)] will influence flowering time in the future. In addition, interactions between elevated [CO(2)] and other global change factors may further complicate our ability to predict changes in flowering time. One approach to overcoming this problem is to elucidate the primary mechanisms that control flowering-time responses to elevated [CO(2)]. Unfortunately, the mechanisms controlling these responses are not known. However, past work has indicated that carbon metabolism exerts partial control on flowering time, and therefore may be involved in elevated [CO(2)]-induced changes in flowering time. This review also indicates the need for more studies addressing the effects of global change drivers on developmental processes in plants.     

Mutagenesis of plants overexpressing CONSTANS demonstrates novel interactions among Arabidopsis flowering-time genes

CONSTANS (CO) promotes flowering of Arabidopsis in response to long photoperiods. Transgenic plants carrying CO fused with the cauliflower mosaic virus 35S promoter (35S::CO) flowered earlier than did the wild type and were almost completely insensitive to length of day. Genes required for CO to promote flowering were identified by screening for mutations that suppress the effect of 35S::CO. Four mutations were identified that partially suppressed the early-flowering phenotype caused by 35S::CO. One of these mutations, suppressor of overexpression of CO 1 (soc1), defines a new locus, demonstrating that the mutagenesis approach is effective in identifying novel flowering-time mutations. The other three suppressor mutations are allelic with previously described mutations that cause late flowering. Two of them are alleles of ft, indicating that FT is required for CO to promote early flowering and most likely acts after CO in the hierarchy of flowering-time genes. The fourth suppressor mutation is an allele of fwa, and fwa soc1 35S::CO plants flowered at approximately the same time as co mutants, suggesting that a combination of fwa and soc1 abolishes the promotion of flowering by CO. Besides delaying flowering, fwa acted synergistically with 35S::CO to repress floral development after bolting. The latter phenotype was not shown by any of the progenitors and was most probably caused by a reduction in the function of LEAFY. These genetic interactions suggest models for how CO, FWA, FT, and SOC1 interact during the transition to flowering.     

EFO1 and EFO2, encoding putative WD-domain proteins, have overlapping and distinct roles in the regulation of vegetative development and flowering of Arabidopsis

From screening a population of Arabidopsis overexpression lines, two Arabidopsis genes were identified, EFO1 (early flowering by overexpression 1) and EFO2, that confer early flowering when overexpressed. The two genes encode putative WD-domain proteins which share high sequence similarity and constitute a small subfamily. Interestingly, the efo2-1 loss-of-function mutant also flowered earlier in short days and slightly earlier in long days than the wild type, while no flowering-time or morphological differences were observed in efo1-1 relative to the wild type. In addition, the efo2-1 mutation perturbed hypocotyl elongation, leaf expansion and formation, and stem elongation. EFO1 and EFO2 are both regulated by the circadian clock. Expression and genetic analyses revealed that EFO2 suppresses flowering largely through the action of CONSTANS (CO) and flowering locus T (FT), suggesting that EFO2 is a negative regulator of photoperiodic flowering. The growth defects in efo2-1 were augmented in efo1 efo2, but the induction of FT in the double mutant was comparable to that in efo2-1. Thus, while EFO2 acts as a floral repressor, EFO1 may not be directly involved in flowering, but the two genes do have overlapping roles in regulating other developmental processes. EFO1 and EFO2 may function collectively to serve as one of the converging points where the signals of growth and flowering intersect.     

FRIGIDA-independent variation in flowering time of natural Arabidopsis thaliana accessions

FRIGIDA (FRI) and FLOWERING LOCUS C (FLC) are two genes that, unless plants are vernalized, greatly delay flowering time in Arabidopsis thaliana. Natural loss-of-function mutations in FRI cause the early flowering growth habits of many A. thaliana accessions. To quantify the variation among wild accessions due to FRI, and to identify additional genetic loci in wild accessions that influence flowering time, we surveyed the flowering times of 145 accessions in long-day photoperiods, with and without a 30-day vernalization treatment, and genotyped them for two common natural lesions in FRI. FRI is disrupted in at least 84 of the accessions, accounting for only approximately 40% of the flowering-time variation in long days. During efforts to dissect the causes for variation that are independent of known dysfunctional FRI alleles, we found new loss-of-function alleles in FLC, as well as late-flowering alleles that do not map to FRI or FLC. An FLC nonsense mutation was found in the early flowering Van-0 accession, which has otherwise functional FRI. In contrast, Lz-0 flowers late because of high levels of FLC expression, even though it has a deletion in FRI. Finally, eXtreme array mapping identified genomic regions linked to the vernalization-independent, late-flowering habit of Bur-0, which has an alternatively spliced FLC allele that behaves as a null allele.     

亲环素AtCYP1 RNA干涉载体的构建及对拟南芥发育的影响

根据拟南芥AtCYP1基因序列设计特异引物,以拟南芥总DNA为模板,扩增AtCYP1基因中344 bp转录本,插入表达载体PTCK303,构建目的基因RNA干扰载体Ubi::AtCYP1i。利用改良的农杆菌浸染技术获得拟南芥RNAi转基因株系,RT-PCR分析结果表明转基因株系中AtCYP1基因的表达量低于野生型,表型观察结果表明RNAi转基因纯合株系抽苔时间比野生型晚3.32 d,抽苔叶片数较野生型多2.49片,其开花时间、结出第一个种荚的时间、株高等方面也与野生型存在明显差异。此结果说明AtCYP1可能参与了拟南芥的早花发育过程,为进一步研究其在植物生长发育中的功能奠定了基础。     

Expression of flowering-time genes in soybean E1 near-isogenic lines under short and long day conditions

Control of soybean flowering time is important for geographic adaptation and maximizing yield. Plant breeders have identified a series of genes (E genes) that condition time to flowering; however, the molecular basis in the control of flowering by these E genes, in conjunction with canonical flowering-time genes, has not been studied. Time to flowering in near-isogenic lines (NILs) at the E1 locus was tested using a reciprocal transfer experiment under short day (SD) and long day (LD) conditions. Beginning 8 days after planting, three plant samples were harvested every 3 h for a 48-h period. RNA was isolated from these plants, and RNA samples were pooled for each line and each time period for cDNA synthesis. RT-PCR analysis was performed using primers synthesized for a number of putative flowering-time genes based on homology of soybean EST and genomic sequences to Arabidopsis genes. The results of the reciprocal transfer experiment suggest that the pre-inductive photoperiod-sensitive phase of the E1 NILs responsible for inducing flowering is perceived as early as 5–7-day post-planting. No gene expression differences were found between the E1 and e1 NILs, suggesting that the E1 gene does not directly affect the flowering-time genes during the time period tested; however, differences were observed in gene expression between SD and LD treatments for the putative soybean TOC1, CO, and FT genes. The gene expression results in this study were similar to those of flowering-time genes found in other SD species, suggesting that the selected genes correspond to the soybean flowering-time orthologs.