Arbuscular Mycorrhiza in Highly Fertilized Maize Cultures Alleviates Short-Term Drought Effects but Does Not Improve Fodder Yield and Quality

Under fertilization levels specific to intensive farming, the impact of compensation of soil nutritional value by arbuscular mycorrhiza (AM) might be limited. Therefore, the question arises whether modern crop varieties, selected for high NPK assimilation rate, are able to gain symbiotic benefits under other challenging field conditions, such as drought. Accordingly, in this study we aimed to evaluate the contribution of Rhizophagus irregularis to the drought response of a stay-green corn hybrid in pot cultures equally fertilized until silking, compared to non-mycorrhizal (NM) counterparts. The highest tested fertilization regime not detrimental to the long-term vitality of intraradical hyphae reached the levels recommended for field cultivation of silage corn, except phosphorus application restricted to 60%. Under normal watering, mycorrhiza increased leaf nitrogen and phosphorus acquisition but only in cultures supplied with low NPK levels. At high fertilization levels, only the older leaves retained AM dependency, whereas for other leaf positions the AM-NM differences were leveled out. The similar size and nutritional status of highly fertilized AM and NM cultures, used in this study, eliminated fungal benefits before and during the 2-week drought progression. Nevertheless, mycorrhizal contribution became evident at the time of renewed watering, when AM plants showed much faster reversal of drought-induced leaf senescence symptoms: impaired photosynthesis and nitrogen management. Our results suggest that mycorrhiza can alter drought-induced senescence even in stay-green mutants. Moreover, this effect was apparently not mediated by AM-improved growth but triggered by activation of fungal transport at the time of recovery. Interestingly, the fungal protective potential was shown to be preserved at the expense of lowering AM vesicle number. It can be interpreted as engagement of hyphal nutritional resources targeted to maintain the symbiotic relationship despite the reduced vitality of the host. Finally, we compared the productivity of AM and NM cultures subjected to short-term drought at silking time and further fertilized with moderate or high NPK doses until the grain-filling stage. The yield and nutritive value of green forage showed that alleviation of drought-induced senescence by AM was not sufficient to have a significant positive effect on the final productivity compared to NM plants

Identification of plasma membrane aquaporin isoforms of maize associated with arbuscular mycorrhiza and their participation in symbiotic tolerance to water deficiency

Wydział BiologiiPrzedmiotem badań była analiza akumulacji akwaporyn PIP oraz jej kontekstu transkrypcyjnego u kukurydzy poddanej mikoryzacji (AM) i progresywnie rozwijającej się suszy, a następnie rehydratacji. Badania te były prowadzone w odniesieniu do wybranych parametrów homeostazy roślinnej, a także w oparciu o ocenę żywotności grzyba Rhizophagus irregularis. Analizy proteomiczne wykazały przeciwstawny wzór akumulacji PIP2 w korzeniach i liściach w odpowiedzi na niedobór wody, niezależnie od stanu symbiotycznego. U roślin AM utrzymywany był wysoki poziom PIP1 i PIP2 w liściach oraz PIP1 w korzeniach podczas rehydratacji. Regulacja akwaporyn, pozytywnie korelowało z odtworzeniem fizjologii korzeni i liści do stanu wyjściowego. Identyfikacja proteomiczna, pozwoliła rozpoznać i oszacować proporcję ilościową izoform PIP1 i PIP2 w 13 kompleksach białkowych. Sugerujemy, że izoformy PIP1 i PIP2 łączą się w heterokompleksy o kompozycji zależnej od obecności mikoryzy, poziomu nawodnienia, a także lokalizacji organowej. Ekspresja genów akwaporyn PIP1 i PIP2 wykazywała niską wrażliwość na suszę w liściu wierzchołkowym niezależnie od stanu symbiotycznego, natomiast w liściu przykolbowym była niewrażliwa jedynie u roślin AM. Wydaje się zatem, że jedną z przyczyn mniejszej tolerancji na suszę roślin NM są różnice w poziomie ekspresji akwaporyn w liściach niżej położonych bliżej kolby nasiennej. W zestawieniu wyników analizowanych pięter liściowych możemy zaobserwować korelację akumulacji ABA, SA i parametrów wymiany gazowej oraz ekspresji akwaporyn, które wydają się być elementami utrzymania homeostazy w warunkach suszy.The object of the study was to analyze the accumulation of cell PIP aquaporins and its transcriptional context in mycorrhized (AM) maize and under progressively developing drought, and then rehydration. The study was conducted in relation to selected parameters of plant homeostasis as well as based on the assessment of viability of the fungus Rhizophagus irregularis. Proteomic analyzes showed an opposite accumulation pattern of PIP2 in roots and leaves in response to water deficiency, regardless of the symbiotic state. The AM plants maintained a high level of PIP1 and PIP2 in the leaves and PIP1 in the roots during rehydration. Regulation of aquaporins, positively correlated with restoration of root and leaf physiology to the initial state. Proteomic identification allowed to recognize and estimate the quantitative proportion of PIP1 and PIP2 isoforms in 13 protein complexes. We suggest that PIP1 and PIP2 isoforms combine into heterocomplexes with a composition depending on the presence of mycorrhiza, the level of hydration, as well as the organ localization. Gene expression of selected PIP1 and PIP2 aquaporins showed low sensitivity to drought in top leaf regardless of the symbiotic state, whereas in ear leaf it was insensitive only in mycorrhized plants. It therefore seems, that one of the reasons for the lower tolerance to drought of NM plants are the differences in the level of aquaporin expression in the lower leaves closer to the seed flask. In the results of the analyzed leaf floors, we can observe the correlation of accumulation of ABA, SA and gas exchange parameters as well as aquaporin expression, which seem to be the elements of maintaining homeostasis under drought conditions.Projekt współfinansowany ze środków Unii Europejskiej w ramach Europejskiego Funduszu Społecznego „Stypendia naukowe dla doktorantów kształcących się na kierunkach uznanych za szczególnie istotne z punktu widzenia rozwoju Województwa Lubuskiego”, Poddziałanie 8.2.2 „Regionalne Strategie Innowacji”, Stypendium Fundacji Uniwersytetu im. Adama Mickiewicza w Poznaniu na rok akademicki 2017/2018. Specjalne stypendium naukowe (KNOW) z dotacji podmiotowej w ramach Krajowego Naukowego Ośrodka Wiodącego „Poznańskie Konsorcjum RNA” dla uczestników stacjonarnych studiów doktoranckich na Wydziale Biologii Uniwersytetu im. Adama Mickiewicza w Poznaniu w latach 2016/2017.Praca finansowana ze środków Narodowego Centrum Nauki w ramach projektu nr 2011/01/B/NZ9/00362 realizowanego w latach 2012-2015

Expression patterns of maize PIP aquaporins in middle or upper leaves correlate with their different physiological responses to drought and mycorrhiza

Here we report the effect of Rhizophagus irregularis on maize leaf expression of six plasma membrane aquaporin isoforms from PIP1 and PIP2 subfamilies under severe drought development and recovery. The novelty of our study is the finding that leaf-specific mycorrhizal regulation of aquaporins is dependent on the position of the leaf on the shoot and changes in parallel with the rate of photosynthesis and the stomatal response to drought. The transcripts were isolated from the upper third (L3) or ear (L5) leaf, which differed greatly in physiological response to stress within each symbiotic variant. Aquaporins expression in upper L3 leaves appeared to be largely not sensitive to drought, regardless of symbiotic status. In contrast, L5 leaf of non-mycorrhizal plants, showed strong down-regulation of all PIPs. Mycorrhiza, however, protected L5 leaf from such limitation, which under maximal stress was manifested by 6-fold and circa 4-fold higher transcripts level for PIP1s and PIP2s, respectively. Distinct expression patterns of L3 and L5 leaves corresponded to differences in key parameters of leaf homeostasis - stomatal conductance, photosynthetic rates, and accumulation of ABA and SA as phytohormonal indicators of drought stress. In result symbiotic plants showed faster restoration of photosynthetic capability, regardless of leaf position, which we recognize as the hallmark of better stress tolerance. In summary, arbuscular mycorrhiza alleviates short-term drought effects on maize by preventing the down-regulation of plasma membrane aquaporins within middle leaves, thereby affecting stomatal conductance

DataSheet_1_Expression patterns of maize PIP aquaporins in middle or upper leaves correlate with their different physiological responses to drought and mycorrhiza.docx

Here we report the effect of Rhizophagus irregularis on maize leaf expression of six plasma membrane aquaporin isoforms from PIP1 and PIP2 subfamilies under severe drought development and recovery. The novelty of our study is the finding that leaf-specific mycorrhizal regulation of aquaporins is dependent on the position of the leaf on the shoot and changes in parallel with the rate of photosynthesis and the stomatal response to drought. The transcripts were isolated from the upper third (L3) or ear (L5) leaf, which differed greatly in physiological response to stress within each symbiotic variant. Aquaporins expression in upper L3 leaves appeared to be largely not sensitive to drought, regardless of symbiotic status. In contrast, L5 leaf of non-mycorrhizal plants, showed strong down-regulation of all PIPs. Mycorrhiza, however, protected L5 leaf from such limitation, which under maximal stress was manifested by 6-fold and circa 4-fold higher transcripts level for PIP1s and PIP2s, respectively. Distinct expression patterns of L3 and L5 leaves corresponded to differences in key parameters of leaf homeostasis - stomatal conductance, photosynthetic rates, and accumulation of ABA and SA as phytohormonal indicators of drought stress. In result symbiotic plants showed faster restoration of photosynthetic capability, regardless of leaf position, which we recognize as the hallmark of better stress tolerance. In summary, arbuscular mycorrhiza alleviates short-term drought effects on maize by preventing the down-regulation of plasma membrane aquaporins within middle leaves, thereby affecting stomatal conductance.</p

Genome-wide identification and characterisation of ammonium transporter gene family in barley

Nitrogen (N) is an essential macronutrient for the growth and development of plants, but excessive use of nitrogen fertiliser in agriculture can result in environmental pollution. As a preferred nitrogen form, ammonium (NH4+) is absorbed from the soil by the plants through ammonium transporters (AMTs). Therefore, it is important to explore AMTs to improve the efficiency of plant N utilisation. Here, we performed a comprehensive genome-wide analysis to identify and characterise the AMT genes in barley (HvAMTs), which is a very important cereal crop. A total of seven AMT genes were identified in barley and further divided into two subfamilies (AMT1 and AMT2) based on phylogenetic analysis. All HvAMT genes were distributed on five chromosomes with only one tandem duplication. HvAMTs might play an important role in plant growth, development, and various stress responses, as indicated by cis-regulatory elements, miRNAs, and protein interaction analysis. Further, we analysed the expression pattern of HvAMTs in various developmental plant tissues, which indicated that AMT1 subfamily members might play a major role in the uptake of NH4+ from the soil through the roots in barley. Altogether, these findings might be helpful to improve the barley crop with improved nitrogen use efficiency, which is not only of great significance to the crop but also for land and water as it will reduce N fertiliser pollution in the surrounding ecosystem

The physiological role of fat body and muscle tissues in response to cold stress in the tropical cockroach <i>Gromphadorhina coquereliana</i>

<div><p>Protective mechanisms against cold stress are well studied in terrestrial and polar insects; however, little is known about these mechanisms in tropical insects. In our study, we tested if a tropical cockroach <i>Gromphadorhina coquereliana</i>, possesses any protective mechanisms against cold stress. Based on the results of earlier studies, we examined how short-term (3 h) cold (4°C) influences biochemical parameters, mitochondrial respiration activity, and the level of HSPs and aquaporins expression in the fat body and leg muscles of <i>G</i>. <i>coquereliana</i>. Following cold exposure, we found that the level of carbohydrates, lipids and proteins did not change significantly. Nevertheless, we observed significant changes in mitochondrial respiration activity. The oxygen consumption of resting (state 4) and phosphorylating (state 3) mitochondria was altered following cold exposure. The increase in respiratory rate in state 4 respiration was observed in both tissues. In state 3, oxygen consumption by mitochondria in fat body was significantly lower compared to control insects, whereas there were no changes observed for mitochondria in muscle tissue. Moreover, there were cold-induced changes in UCP protein activity, but the changes in activity differed in fat body and in muscles. Additionally, we detected changes in the level of HSP70 and aquaporins expression. Insects treated with cold had significantly higher levels of HSP70 in fat body and muscles. On the other hand, there were lower levels of aquaporins in both tissues following exposure to cold. These results suggest that fat body play an important role in protecting tropical insects from cold stress.</p></div

Bioenergetic parameters of mitochondria isolated from the fat body tissue of <i>G</i>. <i>coquereliana</i>.

<p>(A-B) Respiration was measured in the presence of 10 mM succinate and in the absence (state 4 respiration) or presence of 400 μM ADP (state 3 respiration). C) RCR refers to respiratory control ratio. D) Changes in the UCP activity of the fat body of <i>G</i>. <i>coquereliana</i> cockroaches following cold exposure. UCP activity was measured in isolated mitochondria in the presence of succinate palmitic acid (PA, an activator of UCP) and GTP (an inhibitor of UCP). Data represent mean value ±SD. Statistical significance is indicated by either <i>p</i>≤0.05 (*) or <i>p</i>≤0.01 (**), or <i>p</i>≤0.001 (***). Student's <i>t</i>-test.</p

The physiological role of fat body and muscle tissues in response to cold stress in the tropical cockroach <i>Gromphadorhina coquereliana</i>

<div><p>Protective mechanisms against cold stress are well studied in terrestrial and polar insects; however, little is known about these mechanisms in tropical insects. In our study, we tested if a tropical cockroach <i>Gromphadorhina coquereliana</i>, possesses any protective mechanisms against cold stress. Based on the results of earlier studies, we examined how short-term (3 h) cold (4°C) influences biochemical parameters, mitochondrial respiration activity, and the level of HSPs and aquaporins expression in the fat body and leg muscles of <i>G</i>. <i>coquereliana</i>. Following cold exposure, we found that the level of carbohydrates, lipids and proteins did not change significantly. Nevertheless, we observed significant changes in mitochondrial respiration activity. The oxygen consumption of resting (state 4) and phosphorylating (state 3) mitochondria was altered following cold exposure. The increase in respiratory rate in state 4 respiration was observed in both tissues. In state 3, oxygen consumption by mitochondria in fat body was significantly lower compared to control insects, whereas there were no changes observed for mitochondria in muscle tissue. Moreover, there were cold-induced changes in UCP protein activity, but the changes in activity differed in fat body and in muscles. Additionally, we detected changes in the level of HSP70 and aquaporins expression. Insects treated with cold had significantly higher levels of HSP70 in fat body and muscles. On the other hand, there were lower levels of aquaporins in both tissues following exposure to cold. These results suggest that fat body play an important role in protecting tropical insects from cold stress.</p></div

Protein profiles of the fat body (A) and leg muscles (B) of <i>G</i>. <i>coquereliana cockroaches</i>.

<p>The frames indicate bands of proteins that show significant changes in protein levels.</p