Plant Root Vol. 8(2014)

For systematic functional analysis of genes, we attempted the application of the FOX-hunting system to super growing-roots (SR) of legume species Lotus corniculatus which was previously reported by Himuro et al. (2011). In this study, we investigated the functional analysis of FSL#35, which was expressed by the rolB gene derived from the Agrobacterium rhizogenes Ri plasmid. In monoculture roots grown in liquid media, the FSL#35 showed specific phenotypes that increased root length, lateral root number and root surface area compared with SR. These enhanced phenotypes of FSL#35 were caused by cell profile alteration, while increased total root length and increased lateral root number were caused by the expansion of cortex cells and increased pericycle cells, respectively. In addition, the FSL#35 root showed high and specific respiration activity compared with SR. These results suggest that distinct cell profiles of FSL#35 were induced by the alteration of respiration activity in root tissue. The enhanced root growth in the FSL#35 root might be induced by alteration of ROS metabolisms. Investigating the details of the rolB gene function, for example by phytohormone analysis, will elucidate the novel benefits of the rolB gene for agriculture.

Plants including tomato produce several kinds of chelator proteins such as metallothioneins (MTs) for protection against Hg²⁺ toxicity. However, the mechanism of protection from Hg²⁺ is not perfectly clear. Hg²⁺ content subsequently was plateaued from days 1 to 7. Cell death and DNA digestion were not observed in the primary root in the presence of Hg²⁺ over the 7 days. The predicted protein sequences of 5 tomato type 2 MT-like (MT2-like) proteins were compared. The coding sequences of accession number Z68185 had no Cys-Cys motif in the N-terminal. However, the Z68185 cDNA genetic recombinant showed high resistance to Hg²⁺ in bacteria. In tomato, the expression was observed in the roots, but not in the leaves or stems. mRNA of the MT2-like protein was measured in tomato seedlings exposed to 1 μM Hg²⁺. The expression level did not increase until day 3, but increased expression was observed after day 5. These results suggest that new Metallothionein2-like protein express in root specific and it may trap mercury. Our results indicate that functional identification of an MT2-like protein will be useful for molecular breeding designed to improve plant tolerance to Hg²⁺.

Water stress is a major limiting factor for plant growth and development. In this study, we investigated the effects of L-β-phenyllactic acid (LPA) on growth of rice (Oryza sativa L.) seedlings under polyethylene glycol (PEG)-induced water deficit conditions. Seedlings were culture at 30 °C for 14 days in growth pouches supplemented with 1/100-strength Murashige and Skoog (MS) medium and PEG in the presence or absence of 100 mg L^-1 LPA. As evidenced by plant height, LPA application enhanced seedling growth under PEG-induced water deficit by 13%. The shoot dry weight was slightly increased, whereas that of roots was markedly enhanced during LPA treatment by 26% under water-deficit conditions. No difference was observed among treatments in the number of roots per seedling. The ratio of shoot dry weight to shoot length (RWL) was constant regardless of treatment, indicating that LPA does not cause spindly shoot growth. The total length, surface area, and volume of fine roots were increased by LPA under PEG-induced water deficit conditions. Plant height was significantly correlated with total root surface area and volume. The results imply that PEG-induced water deficit in rice seedlings can be alleviated by LPA application. This alleviative effect is partially attributable to alterations in root system developmental patterns, with increases in fine root total length, surface, and volume accelerating water and nutrient acquisition from the culture medium.

Soil sickness is a widespread problem in replanted apple orchards with a complex symptomatology and etiology influenced by soil and climate conditions. Consequently, a conclusive technical solution is still lacking for intensive apple orchards. The present work aims to analyze the morphological and functional changes occurring in the M9 apple root systems growing in pot filled with soil derived from five different European growing areas. For each growing area, the soil was collected from the apple orchard and used directly in the pot or gamma-ray sterilized before potting. Soil from a neighborhood fallow was also used as control for each growing area. In the non-sterilized replant soils plants developed poor root systems due to a limited biomass allocation. Fibrous roots production was particularly compromised. The roots had a smaller diameter and a lower ramification index. The root cell membrane integrity was also lower. Gamma-ray sterilized replant soils increased root growth, branching and cell integrity, while nearby fallow soil induced an intermediate root behavior. The magnitude of the symptoms showed a significant interaction between soil treatment and sampling site and root growth was correlated with the organic matter content in the soils.

We investigated the short-term response of poplar roots to low and high nitrogen availability in order to elucidate the mechanisms involved in nutrient acquisition. After 28 days of fertilization with low versus high ammonium nitrate, an increase in aboveground biomass was observed accompanied by a decrease in root biomass, reducing the root: shoot ratio after 28 days. These changes in biomass allocation were accompanied by changes in root architecture and altered gene expression. The gene expression response was evaluated after 7 days using a custom cDNA micrarray following transfer to low and high nitrogen supply. We found that 56 sequences were differentially expressed in poplar roots. Many of these 56 genes could be associated with putative roles in development or response to biotic and abiotic stress. A time course analysis of selected cell wall-related genes by RT-qPCR confirmed the expression patterns obtained by microarray and also showed the timing of this differential response. Our results show that patterns of transcript accumulation in roots of poplars are influenced by nitrogen supply, providing evidence of unique nitrogen-adaptative mechanisms.

Roots of pomegranate (Punica granatum L.), belonging to Punicaceae, were investigated anatomically to record changes in tissue development from growth to maturity. When roots start secondary growth, a protective tissue called polyderm composed of alternating suberized and non-suberized cell layers, develop beyond the endodermis in certain families of plants including Myrtaceae. Punicaceae family is not known to develop a polyderm. However, new layers formed beyond the endodermis during secondary growth and biopolymer deposition was observed in their cell walls. The present study aims to gather more knowledge on this tissue discovered in pomegranate roots and cross check whether it is a polyderm or a unique type of periderm. Root specimens were sectioned freehand or with an ultramicrotome after embedding in Technovit 7100 resin. After staining with berberine hemisulfate-aniline blue-safranin O, the root sections were observed under fluorescent or optical microscopes. Unlike the polyderm in Myrtaceae roots, in pomegranate roots, ligno-suberic material accumulated in every cell layer beyond the endodermis. The alternating suberized and non-suberized layers that define the polyderm were absent. Lignin accumulation in the cell wall was pronounced in every cell layer of this outermost tissue and suberin-like autofluorescence was also observed in the same layer. We considered this to be a unique feature typical in pomegranate periderm. It is possible that accumulating both lignin and suberin in the same cell layer instead of alternative layers is more efficient because metabolic energy is not spent in forming a separate cell layer. Further experiments are underway to acknowledge changes in such biopolymer accumulation in the outermost tissue in abiotic stress conditions.

Details on localized deposition of biopolymers in cell wall have rarely been studied. Anatomical observation is important to understand tissue-specific accumulation of biopolymers and such knowledge can further clarify tissue-function. Myrtaceae roots possess alternating suberized and non-suberized cell layers known as polyderm that exist beyond the endodermis. It is important to understand firstly, where biopolymers accumulate in this tissue and secondly what mechanisms control cell wall modification. This study aims to study areas of biopolymer deposition in Myrtaceae root tissues. Root specimens were sectioned freehand or with an ultramicrotome after embedding in Technovit 7100 resin. Root sections were stained with berberine hemisulfate-aniline blue-safranin O series or just phloroglucinol and observed under a fluorescent or optical microscope. Different biopolymers accumulated alternatively on opposite sides of the cell wall in polyderm. In non-suberized tissues, lignin accumulation was dominant and its accumulation appeared to be “closed” centripetally resembling the letter “w”. In suberized tissues, it resembled the letter “m”. This tissue-specific accumulation pattern was common in all five Myrtaceae species. What factors could control and regulate such patterned and tissue-specific accumulation of biopolymers? Could this accumulation pattern itself be a contributing factor to its protective role? Discovery of this pattern, specific to Myrtaceae root polyderm, triggers more investigations on the effect of biotic and abiotic stress on biopolymer accumulation in it.

Agrobacterium rhizogenes mediated hairy root culture of Plumbago rosea L. is an attractive alternative for the production of plumbagin which is the major bioactive compound in P. rosea tuberous roots. The traditional industries form the major consumer of the tuberous roots as these are used in many ayurvedic preparations. The present work investigates the prospects of utilizing hairy roots in the place of tuberous roots based on bacterial survival test of hairy roots and comparison through phytochemical analyses (TLC, Spectrophotometry, HPLC and LC-MS). Since the traditional system of medicine follows stringent curing procedure before incorporation of the roots in medicinal preparations, cured tuberous and hairy roots were also compared. The phytochemical profile of hairy roots was remarkably similar to that of tuberous roots. Curing caused no change in the phytotchemical composition of the roots but only a reduction in the amount of plumbagin and other molecules. Plumbagin was reduced to 0.372 ± 0.026% dry weight (DW) in cured tuberous roots (1.15 ± 0.08% DW in uncured) and 0.061± 0.0043% DW in cured hairy roots (1.32 ± 0.09% DW in uncured). An 11.3 fold increase in root biomass with 1.56% g DW plumbagin obtained in bioreactor as against 5.39 fold in shake-flasks (with 1% w/v inoculum over 3 weeks period), adds to the prospects of its applicability in traditional systems. The results suggest a refurbishment of conventional high quantity cured roots in traditional preparations with low quantity uncured roots, irrespective of root types.

Many crops are sensitive to waterlogging. A small, fast-growing grass, Brachypodium distachyon (Bd21), whose genome has been sequenced, is a new model for studying cereal crops such as wheat and barley, and for developing novel biomass grasses. However, its waterlogging tolerance and oxygen transport properties are not known. Here, we show that in stagnant deoxygenated nutrient solution, which mimics waterlogged soil, B. distachyon grows poorly and does not increase the number of newly formed roots. In both aerated and stagnant conditions, aerenchyma was hardly observed in roots, and root porosities were low. Suberin and lignin, which are thought to be constituents of Casparian strips and the barrier to radial oxygen loss, did not develop in the outer part of roots in either aerated or stagnant conditions. Our results suggest that the abilities of oxygen transport in B. distachyon are insufficient to grow and survive in stagnant deoxygenated conditions.