Silvia Quaggiotti

Humic substances biological activity at the plant-soil interface: From environmental aspects to molecular factors

Humic substances (HS) represent the organic material mainly widespread in nature. HS have positive effects on plant physiology by improving soil structure and fertility and by influencing nutrient uptake and root architecture. The biochemical and molecular mechanisms underlying these events are only partially known. HS have been shown to contain auxin and an “auxin-like” activity of humic substances has been proposed, but support to this hypothesis is fragmentary. In this review article, we are giving an overview of available data concerning molecular structures and biological activities of humic substances, with special emphasis on their hormone-like activities.

NO homeostasis is a key regulator of early nitrate perception and root elongation in maize

Nitrate reductase produces nitric oxide (NO) as an early response to nitrate, and the coordinated induction of ns-haemoglobins finely modulates NO level. The control of NO homeostasis regulates root elongation and represents a novel key component of nitrate signaling in maize

Transcriptome analysis reveals coordinated spatiotemporal regulation of hemoglobin and nitrate reductase in response to nitrate in maize roots

Given the importance of nitrogen for plant growth and the environmental costs of intense fertilization, an understanding of the molecular mechanisms underlying the root adaptation to nitrogen fluctuations is a primary goal for the development of biotechnological tools for sustainable agriculture. This research aimed to identify the molecular factors involved in the response of maize roots to nitrate. cDNA-amplified fragment length polymorphism was exploited for comprehensive transcript profiling of maize (Zea mays) seedling roots grown with varied nitrate availabilities; 336 primer combinations were tested and 661 differentially regulated transcripts were identified. The expression of selected genes was studied in depth through quantitative real-time polymerase chain reaction and in situ hybridization. Over 50% of the genes identified responded to prolonged nitrate starvation and a few were identified as putatively involved in the early nitrate signaling mechanisms. Real-time results and in situ localization analyses demonstrated co-regulated transcriptional patterns in root epidermal cells for genes putatively involved in nitric oxide synthesis/scavenging. Our findings, in addition to strengthening already known mechanisms, revealed the existence of a new complex signaling framework in which brassinosteroids (BRI1), the module MKK2-MAPK6 and the fine regulation of nitric oxide homeostasis via the co-expression of synthetic (nitrate reductase) and scavenging (hemoglobin) components may play key functions in maize responses to nitrate.

Transcriptome profiling of genes differentially modulated by sulfur and chromium identifies potential targets for phytoremediation and reveals a complex S-Cr interplay on sulfate transport regulation in B. juncea

A differential display cDNA-AFLP derived technique was used to identify gene transcripts regulated by chromium (Cr) in relation to sulfur (S) nutrition in Brassica juncea. Twelve-day old plants were grown with 200 μM sulfate (+S), without sulfate (-S), with 200 μM sulfate plus 200 μM chromate (+S+Cr), or without sulfate plus 200 μM chromate (-S+Cr). Forty-four combinations of degenerate primers were assayed, which allowed the detection of 346 Transcript-Derived Fragments (TDFs) differentially regulated by Cr and S at times 0, 10 min, 1 h, 24 h. Eight sulfate transporters were identified, whose transcript abundance was dependent on the levels of plant S-compounds. For some of these transporters, a tight coordinated regulation of gene expression was observed in response to Cr. The MapMan analysis revealed a differential pattern of gene expression between +S+Cr and -S+Cr plants for several other transcripts and highlighted an overlap among responses to metals, defence against pathogens and senescence, hence suggesting the existence of common mechanisms of gene regulation. Among the identified gene transcripts, those involved in S metabolism and proteolitic processes may represent potential targets of genetic engineering in efforts to increase Cr accumulation and tolerance in plant species employed in phytoremediation techniques.

Nitrate sensing by the maize root apex transition zone: a merged transcriptomic and proteomic survey

Highlight A combined untargeted approach was adopted to achieve a picture of the transcriptional and proteomic profiles typifying the maize root transition zone in response to nitrate.

NO signaling is a key component of the root growth response to nitrate in Zea mays L.

Roots are considered to be a vital organ system of plants due to their involvement in water and nutrient uptake, anchorage, propagation, storage functions, secondary metabolite (including hormones) biosynthesis, and accumulation. Crops are strongly dependent on the availability of nitrogen in soil and on the efficiency of nitrogen utilization for biomass production and yield. However, knowledge about molecular responses to nitrogen fluctuations mainly derives from the study of model species. Nitric oxide (NO) has been proposed to be implicated in plant adaptation to environment, but its exact role in the response of plants to nutritional stress is still under evaluation. Recently a novel role for NO production and scavenging, thanks to the coordinate spatio-temporal expression of nitrate reductase and non-symbiotic hemoglobins, in the maize root response to nitrate, has been postulated. This control of NO homeostasis is preferentially accomplished by the cells of the root transition zone (TZ) which seems to represent the most nitrate responsive portion of maize root. The TZ is already known to function as a sensory center able to gather information from the external environment and to re-elaborate them in an adequate response. These results indicate that it could play a central role also for nitrate sensing by roots. A lot of work is still needed to identify and characterize other upstream and downstream signals involved in the “nitrate-NO” pathway, leading to root architecture adjustments and finally to stress adaptation.

Expression of two maize putative nitrate transporters in response to nitrate and sugar availability.

A full-length cDNA encoding a putative high-affinity nitrate transporter (ZmNrt2.2) from maize was isolated and characterised, together with another previously identified transporter (ZmNrt2.1), in terms of phylogenesis, protein structure prediction and regulation of transcript accumulation in response to nitrate and sugar availability. The expression of both genes was evaluated by quantitative and semi-quantitative RT-PCR in response to nitrate and sugar supply and the in planta localisation of mRNA was studied by in situ hybridisation. Data obtained suggested similar genetic evolution and identical transmembrane structure prediction between the two deduced proteins, and differences in both regulation of their expression and mRNA localisation in response to nitrate, leading us to hypothesise a principal role for ZmNRT2.1 in the influx activity and the major involvement of ZmNRT2.2 in the xylem loading process. Our data suggest opposing sugar regulation by ZmNrt2.1 and ZmNrt2.2 transcription in the presence or absence of nitrate and the existence of both hexokinase-dependent and hexokinase-independent transduction mechanisms for the regulation of ZmNrt2.1 and ZmNrt2.2 expression by sugars.

Physiological and molecular aspects of sulphate uptake in two maize hybrids in response to S-deprivation

Two maize hybrids (KW2 and KW7), chosen on the basis of their productivity with low nutritional input, showed different physiological strategies, with opposite sulphate uptake Vmax and Km values, in response to sulphate deprivation. In order to characterise the physiological differences between the two hybrids, sulphate influx rates were measured in different nutritional conditions. When grown for 7 days in S-deprived solution, significantly higher influx rates were measured in roots of KW7 than KW2. Withdrawal of sulphate, after 7 days in presence of S, induced the derepression of the sulphate transport system in both hybrids. However, maize hybrid KW2 seemed to have a more rapid activation than that of KW7, which, in turn, seemed to be more resistant to prolonged sulphur deprivation. The data obtained were not supported by the outcome of the analysis of mRNA abundance, performed with the homologue probe ZmST1 on roots of both hybrids. Root sulphate and glutathione contents were in all cases higher in KW7 than KW2. The discrepancy between the physiological and the molecular data suggests the possible existence of other root transporters involved in sulphate uptake, and/or of some post-transcriptional regulatory mechanisms.

Whole-mount in situ detection of microRNAs on Arabidopsis tissues using Zip Nucleic Acid probes

MicroRNAs (miRNAs) affect fundamental processes of development. In plants miRNAs regulate organ development, transition to flowering, and responses to abiotic/biotic stresses. To understand the biological role of miRNAs, in addition to identifying their targeted transcripts, it is necessary to characterize the spatiotemporal regulation of their expression. Many methods have been used to define the set of organ-specific miRNAs by tissue dissection and miRNA profiling but none of them can describe their tissue and cellular distribution at the high resolution provided by in situ hybridization (ISH). This article describes the setup and optimization of a whole-mount ISH protocol to target endogenous miRNAs on intact Arabidopsis seedlings using DIG-labeled Zip Nucleic Acid (ZNA) oligonucleotide probes. Automation of the main steps of the procedure by robotized liquid handling has also been implemented in the protocol for best reproducibility of results, enabling running of ISH experiments at high throughput.

The miRNA-mediated post-transcriptional regulation of maize response to nitrate

Stress responses depend on the correct regulation of gene expression. The discovery that abiotic as well as biotic stresses can regulate miRNA levels, coupled with the identification and functional analyses of stress-associated genes as miRNA targets, provided clues about the vital role that several miRNAs may play in modulating plant resistance to stresses. Nitrogen availability seriously affects crops productivity and environment and the understanding of the miRNA-guided stress regulatory networks should provide new tools for the genetic improvement of nitrogen use efficiency of crops. A recent study revealed the potential role of a number of nitrate-responsive miRNAs in the maize adaptation to nitrate fluctuations. In particular, results obtained suggested that a nitrate depletion might regulate the expression of genes involved in the starvation adaptive response, by affecting the spatio-temporal expression patterns of specific miRNAs.