A set of Arabidopsis thaliana miRNAs involve shoot regeneration in vitro

Plant miRNAs, the critical regulator of gene expression, involve many development processes in vivo. However, the roles of miRNAs in plant cell proliferation and redifferntiation in vitro remain unknown. To determine better the molecular mechanism of these processes, we have recently reported that a set of miRNAs with different expression patterns between cells of totipotent and non-totipotent Arabidopsis calli. Some of these were specifically up- or downregulated during callus formation or shoot regeneration, and other development. Among them, miR160, and one of its target genes, ARF10, regulated Arabidopsis in vitro shoot regeneration via WUS, CLV3 and CUC1/2. The miR160-overexpressing, 35S transgenic lines, exhibited reduced shoot regeneration efficiency. The mARF10, a miR160-resistant form of ARF10, showed a high level of shoot regeneration ability. In the transgenic, expression of the above shoot meristem-specific genes was elevated, which is consistent with the improved shoot regeneration. In contrast, the ARF10 deficient knockout mutant produced fewer regenerated shoot. However, overexpressors of ARF10 were only marginally more efficient than the wild type with the respect to shoot regeneration. Our observation strongly supports that proper shoot regeneration from in vitro cultured cells requires the miR160-directed negative influence of ARF10. The enhanced expression of ARF10 is likely to have contributed to the improved regeneration ability.     

Recovery of dicer-like 1-late flowering phenotype by miR172 expressed by the noncanonical DCL4-dependent biogenesis pathway

MicroRNAs (miRNAs) act as down-regulators of gene expression, and play a dominant role in eukaryote development. In Arabidopsis thaliana, DICER-LIKE 1 (DCL1) is the main processor in miRNA biogenesis, and dcl1 mutants show various developmental defects at the early stage of embryogenesis or at gamete formation. However, miRNAs responsible for the respective developmental stages of the dcl1 defects have not been identified. Here, we developed a DCL1-independent miRNA expression system using the unique DCL4-dependent miRNA, miR839. By replacing the mature sequence in the miR839 precursor sequence with that of miR172, one of the most widely conserved miRNAs in angiosperms, we succeeded in expressing miR172 from a chimeric miR839 precursor in dcl1-7 plants and observed the repression of miR172 target gene expression. In parallel, the DCL4-dependent miR172 expression rescued the late flowering phenotype of dcl1-7 by acceleration of flowering. We established the DCL1-independent miRNA expression system, and revealed that the reduction of miR172 expression is responsible for the dcl1-7 late flowering phenotype.     

The I2 resistance gene homologues in Solanum have complex evolutionary patterns and are targeted by miRNAs

Background

Several resistance traits, including the I2 resistance against tomato fusarium wilt, were mapped to the long arm of chromosome 11 of Solanum. However, the structure and evolution of this locus remain poorly understood.

Results

Comparative analysis showed that the structure and evolutionary patterns of the I2 locus vary considerably between potato and tomato. The I2 homologues from different Solanaceae species usually do not have orthologous relationship, due to duplication, deletion and frequent sequence exchanges. At least 154 sequence exchanges were detected among 76 tomato I2 homologues, but sequence exchanges between I2 homologues in potato is less frequent. Previous study showed that I2 homologues in potato were targeted by miR482. However, our data showed that I2 homologues in tomato were targeted by miR6024 rather than miR482. Furthermore, miR6024 triggers phasiRNAs from I2 homologues in tomato. Sequence analysis showed that miR6024 was originated after the divergence of Solanaceae. We hypothesized that miR6024 and miR482 might have facilitated the expansion of the I2 family in Solanaceae species, since they can minimize their potential toxic effects by down-regulating their expression.

Conclusions

The I2 locus represents a most divergent resistance gene cluster in Solanum. Its high divergence was partly due to frequent sequence exchanges between homologues. We propose that the successful expansion of I2 homologues in Solanum was at least partially attributed to miRNA mediated regulation.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-743) contains supplementary material, which is available to authorized users.     

Stictocardia tiliifolia (Convolvulaceae) re-evaluated

Stictocardia tiliifolia (Desr.) H. Hall. is accepted as the correct name for the type species ofStictocardia. An earlier name,Ipomoea campanulata L., regarded as a synonym of S.tiliifolia for the last 50 years, is found to be synonymous withIpomoea illustris (C. B. Clarke) Prain, a species occurring in coastal/tidal riverine habitats of South Asia. Material from interior habitats, calledI. campanulata by authors of Indian floras, is believed to represent a variety of trueI. campanulata or a closely related species.     

Anisakis pegreffii Larvae in Sea Eels (Astroconger myriaster) from the South Sea,Republic of Korea

Anisakis simplex sensu stricto (s.s.), Anisakis pegreffii, Anisakis berlandi (=A. simplex sp. C), and Anisakis typica are the 4 major species of Anisakis type I larvae. In the Republic of Korea (Korea), A. pegreffii, A. berlandi, and A. typica larvae in fish hosts has seldom been documented. In this study, molecular analysis was performed on Anisakis larvae from the sea eels (Astroconger myriaster), the major source of human anisakiasis in Korea, collected from Tongyeong City, a southern coastal area of Korea. All 20 sea eels examined were infected with Anisakis type I larvae (160 larvae; 8 per fish). Their species were analyzed using PCR-RFLP patterns and nucleotide sequences of internal transcribed spacers (ITS1, 5.8 subunit gene, and ITS2) and mitochondrial cytochrome c oxidase 2 (cox2). Most (86.8%; 112/129) of the Anisakis type I larvae were A. pegreffii, and 7.8% (10/129) were A. typica. The remaining 5.4% (7/129) was not identified. Thus, A. pegreffii is the major species of anisakid larvae in sea eels of the southern coast of Korea.     

Description of Globodera ellingtonae n. sp. (Nematoda: Heteroderidae) from Oregon

A new species of cyst nematode, Globodera ellingtonae, is described from soil collected from a field in Oregon. Second-stage juveniles (J2) of the species are characterized by body length of 365-515 μm, stylet length of 19-22.5 μm, basal knobs rounded posteriorly and pointed anteriorly, tail 39-55 μm, hyaline tail terminus 20-32.5 μm, and tail tapering uniformly but abruptly narrowing and constricted near the posterior third of the hyaline portion, ending with a peg-like, finely rounded to pointed terminus. Cysts are spherical to sub-spherical, dark to light brown and circumfenestrate and cyst wall pattern is ridge-like with heavy punctations. Males have a stylet length of 21-25 μm and spicule length of 30-37 μm with a pointed thorn-like tip. Females have a stylet length of 20-22.5 μm, one head annule and labial disc, heavy punctations on the cuticle, and short vulval slit 7.5-8 μm long. Morphologically this new, round-cyst species differs from the related species G. pallida, G. rostochiensis, G. tabacum complex and G. mexicana by its distinctive J2 tail, and by one or another of the following: shorter mean stylet length in J2, females and males; number of refractive bodies in the hyaline tail terminus of J2; cyst morphology including Granek’s ratio; number of cuticular ridges between the anus and vulva; and in the shape and length of spicules in males. Its relationship to these closely related species are discussed. Based upon analysis of ribosomal internal transcribed spacer (ITS) sequences, G. ellingtonae n. sp. is distinct from G. pallida, G. rostochiensis, G. tabacum and G. mexicana. Bayesian and Maximum Parsimony analysis of cloned ITS rRNA gene sequences indicated three clades, with intraspecific variability as high as 2.8%. In silico analysis revealed ITS restriction fragment length polymorphisms for enzymes Bsh 1236I, Hinf I, and Rsa I that overlap patterns for other Globodera species.     

Estimating contribution of anthocyanin pigments to osmotic adjustment during winter leaf reddening

The association between plant water stress and synthesis of red, anthocyanin pigments in leaves has led some plant biologists to propose an osmotic function of leaf reddening. According to this hypothesis, anthocyanins function as a solute in osmotic adjustment (OA), contributing to depression of osmotic potential (Ψ_π) and maintenance of turgor pressure during drought-stressed conditions. Here we calculate the percent contribution of anthocyanin to leaf Ψ_π during OA in two angiosperm evergreen species, Galax urceolata and Gaultheria procumbens. Both species exhibit dramatic leaf reddening under high light during winter, concomitant with declines in leaf water potential and accumulation of solutes. Data previously published by the authors on osmotic potential at full turgor (Ψ_π,100) of G. urceolata and G. procumbens leaves before and after leaf reddening were used to estimate OA. In vivo molar concentrations of anthocyanin, glucose, fructose, and sucrose measured from the same individuals were converted to pressure equivalents using the Ideal Gas Law, and percent contribution to OA was estimated. Estimated mean OA during winter was −0.7 MPa for G. urceolata and −0.8 MPa for G. procumbens. In vivo concentrations of anthocyanin (3–10 mM) were estimated to account for ∼2% of OA during winter, and comprised <0.7% of Ψ_π,100 in both species. Glucose, fructose, and sucrose combined accounted for roughly 50 and 80% of OA for G. urceolata and G. procumbens, respectively, and comprised ∼20% of Ψ_π,100. We observed that a co-occurring, acyanic species (Vinca minor) achieved similar OA without synthesizing anthocyanin. We conclude that anthocyanins represent a measurable, albeit meager, component of OA in red-leafed evergreen species during winter. However, due to their low concentrations, metabolic costliness relative to other osmolytes, and striking red color (unnecessary for an osmotic function), it is unlikely that they are synthesized solely for an osmoprotectant role.     

Profiling of MicroRNAs under Wound Treatment in Aquilaria sinensis to Identify Possible MicroRNAs Involved in Agarwood Formation

Agarwood, a kind of highly valued non-timber product across Asia, is formed only when its resource trees -- the endangered genus Aquilaria are wounded or infected by some microbes. To promote the efficiency of agarwood production and protect the wild resource of Aquilaria species, we urgently need to reveal the regulation mechanism of agarwood formation. MicroRNAs (miRNAs) are a group of gene expression regulators with overwhelming effects on a large spectrum of biological processes. However, their roles in agarwood formation remain unknown. This work aimed at identifying possible miRNAs involved in the wound induced agarwood formation. In this study, the high-throughput sequencing was adopted to identify miRNAs and monitor their expression under wound treatment in the stems of A. sinensis. The miR171, miR390, miR394, miR2111, and miR3954 families remained at the reduced level two days after the treatment. 131 homologous miRNAs in the 0.5 h library showed over three-fold variation of read number compared with the control library, of which 12 exhibiting strong expression alterations were further confirmed by real-time quantitative PCR. Target prediction and annotation of the miRNAs demonstrated that the binding, metabolic process, catalytic activity, and cellular process are the most common functions of the predicted targets of these newly identified miRNAs in A.sinensis. The cleaveage sites of three newly predicted targets were verified by 5''RACE.