Presence Of Auxin Research Articles

Antioxidant and Anticancer Activity of Tannins Isolated from Callus Cultures of Achyranthes aspera L.

In the current study, the impact of oxidative stress induced by free radicals generated during metabolic processes was investigated. It was found to be linked to a variety of diseases due to their detrimental effects on nucleic acids and proteins. The primary focus remained on investigating the free radical-scavenging properties and potential anticancer activities of tannins found in callus cultures of A. aspera. It was observed that the induction of callus formation was notably successful when using leaf and root explants, resulting in a callus index as high as 160, as compared to stem explants in the presence of auxins. The extraction yield of callus tannins was the most abundant in chloroform extracts, although the overall antioxidant activity was comparable among chloroform, methanol, and petroleum ether callus extracts. Among these extracts, the chloroform extract from stem callus cultures grown on the MSDN medium showed the highest total antioxidant activity. Notably, tannins extracted from A. aspera leaf callus extracts displayed significant anticancer effects against the Jurket cell line. These effects were evaluated through measures such as cell viability and colony formation. The anticancer activity was notably higher in callus culture extracts as compared to the control. In summary, the results showed that in vitro biomass production can be a valuable approach for augmenting the production of bioactive compounds, such as tannins, from selected medicinal plants including A. aspera. Tannins extracted from A. aspera hold promise for their potential role in the development of new medicines.

Abolishing ARF8A activity promotes disease resistance in tomato

Auxin response factors (ARFs) are a family of transcription factors that regulate auxin-dependent developmental processes. Class A ARFs function as activators of auxin-responsive gene expression in the presence of auxin, while acting as transcriptional repressors in its absence. Despite extensive research on the functions of ARF transcription factors in plant growth and development, the extent, and mechanisms of their involvement in plant resistance, remain unknown. We have previously reported that mutations in the tomato AUXIN RESPONSE FACTOR8 (ARF8) genes SlARF8A and SlARF8B result in the decoupling of fruit development from pollination and fertilization, leading to partial or full parthenocarpy and increased yield under extreme temperatures. Here, we report that fine-tuning of SlARF8 activity results in increased resistance to fungal and bacterial pathogens. This resistance is mostly preserved under fluctuating temperatures. Thus, fine-tuning SlARF8 activity may be a potent strategy for increasing overall growth and yield.

Cyanobacterial elicitor enhances the biomass of Mentha piperita L. and improves the production of high-value rosmarinic acid under in vitro culture of apical meristem

BackgroundRosmarinic acid (RA), like other phenolic compounds, is sources of antioxidants and anti-inflammatory agents in medicinal plants. In vitro culture of plants can improve the medicinal plants’ metabolite profile and phenolic compound quantity. To date, various methods have been proposed to increase this medicinal metabolite in plants, among which the use of bioelicitors can be mentioned. In the present study, a native isolate of heterocystous cyanobacteria, Nostoc spongiaeforme var. tenue ISB65, was used to stimulate the production of biomass and content of RA in Mentha piperita L. (peppermint) grown in vitro from apical meristem. Mentha piperita L. explants were inoculated in half strength Murashige and Skoog (1/2 MS) medium containing cyanobacterial lysate (CL). After 50 days of culturing, the growth indices, the content of photosynthetic pigments, and RA in control and treated plants were measured.ResultsCL inoculation resulted in a significant enhancement in the vegetative growth indices of peppermint, including root and shoot length, plant biomass and leaf number. The content of photosynthetic pigments also increased in cyanobacteria-treated plants. Inoculation with CL increased the RA content by 2.3-fold, meaning that the plants treated with CL had the highest RA content (7.68 mg. g− 1 dry weight) compared to the control (3.42 mg. g− 1 dry weight). Additionally, HPLC analysis revealed the presence of several auxins in CL.ConclusionsThe presence of auxins and the chemical content of CL such as K+ and Ca2+, as regulators of metabolic pathways and molecular activities of cells, may be responsible for the enhanced growth and phenolic compounds of plants under tissue culture conditions. An improvement in RA content in the tissue culture of medicinal plants treated with CL was reported for the first time in this investigation.

Intrinsic disorder and conformational coexistence in auxin coreceptors

AUXIN/INDOLE 3-ACETIC ACID (Aux/IAA) transcriptional repressor proteins and the TRANSPORT INHIBITOR RESISTANT 1/AUXIN SIGNALING F-BOX (TIR1/AFB) proteins to which they bind act as auxin coreceptors. While the structure of TIR1 has been solved, structural characterization of the regions of the Aux/IAA protein responsible for auxin perception has been complicated by their predicted disorder. Here, we use NMR, CD and molecular dynamics simulation to investigate the N-terminal domains of the Aux/IAA protein IAA17/AXR3. We show that despite the conformational flexibility of the region, a critical W-P bond in the core of the Aux/IAA degron motif occurs at a strikingly high (1:1) ratio of cis to trans isomers, consistent with the requirement of the cis conformer for the formation of the fully-docked receptor complex. We show that the N-terminal half of AXR3 is a mixture of multiple transiently structured conformations with a propensity for two predominant and distinct conformational subpopulations within the overall ensemble. These two states were modeled together with the C-terminal PB1 domain to provide the first complete simulation of an Aux/IAA. Using MD to recreate the assembly of each complex in the presence of auxin, both structural arrangements were shown to engage with the TIR1 receptor, and contact maps from the simulations match closely observations of NMR signal-decreases. Together, our results and approach provide a platform for exploring the functional significance of variation in the Aux/IAA coreceptor family and for understanding the role of intrinsic disorder in auxin signal transduction and other signaling systems.

The changes in the maize root cell walls after exogenous application of auxin in the presence of cadmium.

Cadmium (Cd) is a transition metal and hazardous pollutant that has many toxic effects on plants. This heavy metal poses a health risk for both humans and animals. The cell wall is the first structure of a plant cell that is in contact with Cd; therefore, it can change its composition and/or ratio of wall components accordingly. This paper investigates the changes in the anatomy and cell wall architecture of maize (Zea mays L.) roots grown for 10 days in the presence of auxin indole-3-butyric acid (IBA) and Cd. The application of IBA in the concentration 10-9 M delayed the development of apoplastic barriers, decreased the content of lignin in the cell wall, increased the content of Ca2+ and phenols, and influenced the composition of monosaccharides in polysaccharide fractions when compared to the Cd treatment. Application of IBA improved the Cd2+ fixation to the cell wall and increased the endogenous concentration of auxin depleted by Cd treatment. The proposed scheme from obtained results may explain the possible mechanisms of the exogenously applied IBA and its effects on the changes in the binding of Cd2+ within the cell wall, and on the stimulation of growth that resulted in the amelioration of Cd stress.

FEATURES OF GROWTH AND INULIN CONTENT IN CALLUS CULTURES Cichorium intybus L. in vitro

In vitro callus cultures of common chicory (Cichorium intybus L.) were obtained and their growth and biochemical characteristics depending on the hormonal composition of the MS medium and the spectral composition of light were studied. The study of the effect of light culture on callus tissue formation and inulin accumulation in it was carried out in opaque grow tents with radiation aligned with the flux density of pho- tosynthetic photons and different ratios of its levels in the region of 660 nm (R, red) and 730 nm (FR, far red). The control variant was placed under white linear fluorescent lamps. The resulting cultures were character- ized by high proliferative activity and the capability for morphogenesis. It has been established that the inter- action of two factors—the presence of auxins in the nutrient medium (IAA or NAA at a concentration of 7.5 mg/L in combination with BAP 0.5 mg/L) and cultivation under light culture conditions (FR R, FR = R, FR R)—had a significant impact on the biosynthetic potential of cell cultures. In the obtained cultures, a study of the quantitative content of inulin was carried out. It has been shown that the high content of inulin (7.55–7.95%) in callus cultures was on the MS medium in combination with IAA at FR R illumination. This is probably due to the fact that well proliferating and highly morphogenic callus tissue was formed under these conditions. The obtained results confirm the hypothesis about the specificity of cultured cells to in vitro synthesize and accumulate secondary metabolites in dedifferentiated cells and the dependence of this process on factors of chemical and physical nature.

Auxin-inducible degronsystem: an efficient protein degradation tool to study protein function.

Targeted protein degradation, with its rapid protein depletion kinetics, allows the measurement of acute changes in the cell. The auxin-inducible degron (AID) system, rapidly degrades AID-tagged proteins only in the presence of auxin. TheAID system being inducible makes the study of essential genes and dynamic processes like cell differentiation, cell cycleand genome organization feasible. The AID degradation system has been adapted to yeast, protozoans, C. elegans, Drosophila, zebrafish, mouseand mammalian cell lines. Using the AID system, researchers have unveiled novel functions for essential proteins at developmental stages that werepreviously difficult to investigate due to early lethality. This comprehensive review discusses the development, advancements, applicationsand drawbacks of the AID system and compares it with other available protein degradation systems.

Каллусогенез у растений баклажана Solanum melongena L. в in vitro культурах

Eggplant (Solanum melongena L.) is an important vegetable crop from the Solanaceae family and one of the most popular vegetable crops in many countries around the world. Transformation of eggplant plants remains one of the main problems of plant genetic engineering due to the relatively low efficiency of callus genesis in different varieties of this crop. Our study analyzed the callus formation of two white-fruited eggplant varieties on a standard regeneration medium with the addition of different concentrations and combinations of hormones in 5 variants. The phytohormones used in the work were 1-naphthaleneacetic acid (NAA), kinetin, which is a cytokinin - a plant hormone that promotes cell division in the presence of auxin and is widely used for callus formation or to induce shoot growth, 6-benzylaminopurine (6-BAP), which is used to accelerate plant growth. Option I with 0.5 mg/l 6-BAP and 2.0 mg/l NAA, option II with 5.0 mg/l 6-BAP and 0.1 mg/l NAA, option III with 2.0 mg/l l 6-BAP and 0.1 mg/l NAA, option IV with 2.0 mg/l 6-BAP and 2.0 mg/l kinetin, option V and 10.0 mg/l 6-BAP and 0.2 mg/l NAA. The most suitable were option 2, where some calli formed growths in the form of roots, and in others the rudiments of microplants appeared, and option 3, where a very loose callus of white or light brown color was formed, on which the rudiments of microplants also appeared. But later, in both variants, all calli died. In variants 1, 4 and 5, the cotyledon explants turned brown, first at the edges, and then completely, and then died. Unfortunately, our study did not select optimal combinations of hormones that would be suitable for effective callusogenesis of different eggplant varieties and the work will continue.

Fodder radish aqueous extracts and associated rhizobacteria as bio-stimulants on tomato growth

ABSTRACT Use of bio-stimulants can reduce amounts of synthetic chemicals for improving tomato (Solanum lycopersicum L.) growth and production. The investigation evaluated the rhizobacterial isolates B45, B46, B48, B54 and B55 associated with fodder radish (Raphanus sativus L.) and its aqueous extracts for their ability to promote growth of tomato plants under greenhouse conditions. Rhizobacteria were applied by root dipping followed by seedling drenching at planting. The isolates were characterized, based on macro-morphological and biochemical traits, and mechanisms involved in Plant Growth-Promoting (PGP) features were explored. For direct PGP traits, isolates were able to fix nitrogen on N-free agar medium. Isolates B45, B46, B54 and B55 were positive for indole-3-acetic acid production; isolates B46, B48 and B55 were positive for phosphate solubilization with the highest activity exhibited by B46. For indirect PGP traits, isolates were chitinase-, lipase-, and amylase-producing agents. Based on phylogenetic analysis of the 16S rDNA, the most bioactive isolate B46 was affiliated to Bacillus vallismortis (OK083726). All isolates displayed antifungal activity against soil-borne fungi Aspergillus niger, Penicillium digitatum, Fusarium oxysporum, F.sambucinum, and Trichoderma harzianum. Fodder radish aqueous extracts were tested at 5, 7.5, 10 or 15% (w:v). Chemical characterization by spectrophotometer of compounds indicated presence of auxin and polyphenols. Fodder radish aqueous extract displayed antifungal activity against all fungal species. Fodder radish appears to be a natural source for isolation of potent PGP rhizobacterial isolates and/or secondary metabolites bioactive for promotion of tomato growth and with an antifungal potential.

Identification and characterization of a curly-leaf locus CL1 encoding an IAA2 protein in Brassica napus

The leaf is the main organ for rapeseed photosynthesis, and its morphology influences photosynthetic efficiency and supports increased planting density and yield. However, the molecular regulatory mechanism of leaf morphology in Brassica napus is poorly understood, restricting progress in breeding for the trait. We describe a novel dominant mutation, curly leaf 1 (cl1), which confers uneven dorsal–ventral axis development, irregular cellular structure and influenced gravitropic response in the seedling stage. The CL1 locus was mapped to a 1.573-Mb interval on chromosome A05 using simple sequence repeat (SSR) markers, and co-segregated with the phenotype of plants in the curly F2 population. A substitution (P62S) was identified in the highly conserved degron motif (GWSPV) of the IAA2 protein in the cl1 mutant, and the P62S substitution impaired the interaction between IAA2 and TIR1 in the presence of auxin, influencing auxin signaling. The P62S substitution-induced curly leaf phenotype was verified by ectopic expression of BnaA05.iaa2 in Arabidopsis and B. napus. Our findings explain the function of IAA2 in rapeseed, providing a foundation for future investigation of auxin signaling and the mechanisms underlying leaf development in B. napus.

Yeast Kinesin-5 Motor Protein CIN8 Promotes Accurate Chromosome Segregation.

Accurate chromosome segregation depends on bipolar chromosome–microtubule attachment and tension generation on chromosomes. Incorrect chromosome attachment results in chromosome missegregation, which contributes to genome instability. The kinetochore is a protein complex that localizes at the centromere region of a chromosome and mediates chromosome–microtubule interaction. Incorrect chromosome attachment leads to checkpoint activation to prevent anaphase onset. Kinetochore detachment activates the spindle assembly checkpoint (SAC), while tensionless kinetochore attachment relies on both the SAC and tension checkpoint. In budding yeast Saccharomyces cerevisiae, kinesin-5 motor proteins Cin8 and Kip1 are needed to separate spindle pole bodies for spindle assembly, and deletion of CIN8 causes lethality in the absence of SAC. To study the function of Cin8 and Kip1 in chromosome segregation, we constructed an auxin-inducible degron (AID) mutant, cin8-AID. With this conditional mutant, we first confirmed that cin8-AID kip1∆ double mutants were lethal when Cin8 is depleted in the presence of auxin. These cells arrested in metaphase with unseparated spindle pole bodies and kinetochores. We further showed that the absence of either the SAC or tension checkpoint was sufficient to abolish the cell-cycle delay in cin8-AID mutants, causing chromosome missegregation and viability loss. The tension checkpoint-dependent phenotype in cells with depleted Cin8 suggests the presence of tensionless chromosome attachment. We speculate that the failed spindle pole body separation in cin8 mutants could increase the chance of tensionless syntelic chromosome attachments, which depends on functional tension checkpoint for survival.

Dopamine, Chlorogenic Acid, and Quinones as Possible Cofactors of Increasing Adventitious Rooting Potential of In Vitro Krymsk 5 Cherry Rootstock Explants

In the present study, the effect of some not commonly used phenolic compounds was evaluated during the in vitro rooting stage of the cherry rootstock ‘Krymsk 5′ (P. fruticosa × P. lannesiana), in the absence or presence of auxin. Two sets of experiments were conducted. In the first set, the following substances were tested: the o-diphenol chlorogenic acid, in five concentrations (0 μΜ, 0.5 μΜ, 1 μΜ, 5 μΜ, and 50 μΜ) in the presence of a suboptimal indolebutyric acid (IBA) concentration (5 μΜ), the catecholamine dopamine in five concentrations (0 μΜ, 0.5 μΜ, 1 μΜ, 5 μΜ, and 50 μΜ), and the quinone 5-hydroxy-1,4-naphthoquinone in four concentrations (0 μΜ, 0.25 μΜ, 1 μΜ, and 5 μΜ) in the absence or presence of 5 μΜ IBA. In the second experiment, the quinones p-benzoquinone; 1,4-napthoquinone; and 2-hydroxy-1,4-naphthoquinone were tested in four concentrations (0 μΜ, 5 μΜ, 50 μΜ, and 100 μΜ) in the presence of 5 μΜ IBA. An application of 5 μΜ of 5-hydroxy-1,4-naphthoquinone in the auxin-free medium increased rooting potential almost 1.7 times. Rooting percentage was also enhanced up to 4.2 times by dopamine; chlorogenic acid; 5-hydroxy-1,4-naphthoquinone; p-benzoquinone; and 1,4 napthoquinone in the presence of IBA. The present results indicate a possible promotive role of quinones and dopamine during in vitro rooting, at least for Prunus species, and their potential use as rooting cofactors. Moreover, a possible mode of action of the compounds studied related to IAA-oxidase is discussed.

Influence of plant growth regulators and salicylic acid on the production of some secondary metabolites in callus and cell suspension culture of Satureja sahendica Bornm.

The impact of combinations of plant growth regulators (PGRs) on callus culture of Satureja sahendica Bornm. was investigated. In nodal explants, the response of secondary metabolite production to different concentrations of PGRs was analyzed regarding the presence and absence of polyvinylpyrrolidone (PVP). The explants were cultured on MS media in presence of auxins (2,4-dichlorophenoxyacetic acid and naphthylacetic acid) and cytokinins (thidiazuron and kinetin); which were used in equal concentrations of 0.5, 1, and 2 mg l-1. The treatment of 2 mg l-1 2,4-D + 2 mg l-1 Kin (MD3) led to the highest production of total phenolics (4.303 ± 0.449 mg GAE g-1) and flavonoids (24.903 ± 7.016 mg QE g-1). Moreover, the effect of salicylic acid (SA) on the production of secondary metabolites in cell suspension culture of Satureja sahendica was evaluated. The cell suspension culture was established by culturing the nodal-derived friable callus in the liquid medium containing different concentrations of SA (0, 100, 150, 200 µM). An inverse relationship exists between the fresh mass and secondary metabolites contents. In addition, there was a significant difference among concentrations of SA in the production of total phenolics and flavonoid compounds. SA enhances secondary metabolites production and decreases cell fresh mass.

SUMO orchestrates multiple alternative DNA-protein crosslink repair pathways.

Endogenous metabolites, environmental agents, and therapeutic drugs promote formation of covalent DNA-protein crosslinks (DPCs). Persistent DPCs compromise genome integrity and are eliminated by multiple repair pathways. Aberrant Top1-DNA crosslinks, or Top1ccs, are processed by Tdp1 and Wss1 functioning in parallel pathways in Saccharomyces cerevisiae. It remains obscure how cells choose between diverse mechanisms of DPC repair. Here, we show that several SUMO biogenesis factors (Ulp1, Siz2, Slx5, and Slx8) control repair of Top1cc or an analogous DPC lesion. Genetic analysis reveals that SUMO promotes Top1cc processing in the absence of Tdp1 but has an inhibitory role if cells additionally lack Wss1. In the tdp1Δ wss1Δ mutant, the E3 SUMO ligase Siz2 stimulates sumoylation in the vicinity of the DPC, but not SUMO conjugation to Top1. This Siz2-dependent sumoylation inhibits alternative DPC repair mechanisms, including Ddi1. Our findings suggest that SUMO tunes available repair pathways to facilitate faithful DPC repair.

Biostimulants in the development of tomato and collard greens seedlings

The success of crop yields begins with the use of quality seedlings. Crops from vigorous plants respond better to phytosanitary treatments and management techniques, resulting in economic returns and better use of inputs. The objective of this study was to evaluate NPK fertilizer rates and rates of an NPK + auxin biostimulant on the effects over agronomic features of Collard Greens and salad tomato seedlings. The research consisted of two experiments in randomized block, with four replications each. Both experiments were in factorial scheme 2 x 4, consisting of the combination of two nutrient sources (NPK fertilizer 9-45-11 and biostimulant composed of NPK 9-45-11 + 400 mg kg -1 of auxin IAA), and four rates, being then 50; 100; 150 and 200% of the recommended rate of biostimulant for tomato and 50; 75; 100 and 125% of the recommended rate for Collard Greens. The biostimulant favored the development of tomato seedlings, since it provides greater root dry mass accumulation and didn’t promote seedling shedding, as it occurred in the application of NPK fertilizers. On the other hand, the Collard Greens seedlings didn’t distinguish by the presence of auxin in the biostimulant, developing greater seedlings heights with NPK application. Concentrations of 50-200% of the recommended biostimulant fertilizer didn’t interfere on root length, diameter and root dry mass of Collard Greens.

FLAVONOIDS OF IRIS SIBIRICA L. GROWN IN VITRO CULTURE

The aim of this study was to evaluate the effect of 6-benzylaminopurine (BAP) separately and in interaction with auxins on the change in the qualitative and quantitative composition of flavonoids in the raw materials of regenerating plants Iris sibirica L. Cambridge grade in comparison with aeroponic and intact raw materials using the method of high-performance liquid chromatography.
 Raw materials of I. sibirica Cambridge variety obtained in vitro culture had a richer qualitative composition of flavonoids than intact plants. The dependence of the accumulation of flavonoids on the concentration of 6-benzylaminopurine in nutrient media was noted. The presence of 13 compounds was observed in extracts of 70% ethyl alcohol from regenerating plants grown at the lowest concentration of BAP (1.0 µM) within the experiment. In quantitative terms, the flavonoid apigenin was maximally determined on a medium with BAP 1 µM, and kaempferol - on media with BAP 5.0 µM, supplemented with auxins. For a medium with 7.5 µM BAP, the lowest variety of compounds was observed (9) and the lowest kaempferol content. Auxins influenced the synthesis of flavonoids. The amount of flavonoids in all variants of the experiment increased by an average of 13% in the presence of auxins.
 The stages of the technological process of obtaining raw materials I. sibirica Cambridge variety on the basis of clonal micropropagation and cultivation in aeroponics conditions allowed to obtain raw materials that do not contain heavy and toxic metals, are not infected with pathogens and pests. With 1 m2 of useful area of aeroponics for 1 year, it is possible to collect 5 times more raw materials than with field cultivation. According to the qualitative composition of phenolic compounds, aeroponic raw materials are identical to intact plants.

Evidence of starch accumulation in tobacco Bright Yellow (TBY-2) cells in the presence of auxin

Tobacco Bright Yellow (TBY-2) cell suspension is known to produce starch when cultured in a medium supplemented with cytokinin or in a hormone-free medium. Unexpectedly, TBY-2 cells continuously cultivated in the presence of auxin alone were also able to accumulate starch at the beginning of a stationary growth phase, with a yield of 9.22% ± 0.68%. This starch production was strongly correlated with a 25-fold increase in starch synthase activity. Moreover, this TBY-2 line was able to produce an amylopectin-rich starch with a ratio of amylopectin over amylose of 2.7, which was also linked to typical small granules (1.4 µm). According to our preliminary results, this plant cell suspension could yield a low-cost, amylopectin-rich starch — which is needed in the food industry to produce edible film or bioplastic — without being impacted by climate or seasonal weather changes.

Effect of Plant Growth Regulators on Protease Activity in Forest Floor of Norway Spruce Stand

Soil proteases are involved in organic matter transformation processes and, thus, influence ecosystem nutrient turnovers. Phytohormones, similarly to proteases, are synthesized and secreted into soil by fungi and microorganisms, and regulate plant rhizosphere activity. The aim of this study was to determine the effect of auxins, cytokinins, ethephon, and chlorocholine chloride on spruce forest floor protease activity. It was concluded that the presence of auxins stimulated native proteolytic activity, specifically synthetic auxin 2-naphthoxyacetic acid (16% increase at added quantity of 5 μg) and naturally occurring indole-3-acetic acid (18%, 5 μg). On the contrary, cytokinins, ethephon and chlorocholine chloride inhibited native soil protease activity, where ethephon (36% decrease at 50 μg) and chlorocholine chloride (34%, 100 μg) showed the highest inhibitory effects. It was concluded that negative phytohormonal effects on native proteolytic activity may slow down organic matter decomposition rates and hence complicate plant nutrition. The study enhances the understanding of rhizosphere exudate effects on soil microbial activity and soil nitrogen cycle.

Salivary Origins of Exogenous Cytokinins

Cytokinins are a class of phytohormones which play an important role in a variety of plant processes, such as cell growth and differentiation, formation of roots and shoots, and the reallocation of nutrients to form mobilizing sinks. Exogenous cytokinins found in plant‐manipulating insects are known to contribute to many effects on host plants. Perhaps the most apparent effect is that cytokinins, in the presence of auxin, leads to cell division and the proliferation of plant tissue; consequently, plant galls or tumors are formed. Cytokinins have also been implicated in the reallocation of sugars and nutrients for the oviposition site to act as a mobilizing sink, where the intention may be gall induction or sustenance for the organism. There are two known pathways for the biosynthesis of isoprenoid cytokinins. The first is the tRNA‐ipt pathway, in which an adenine residue is modified by the addition of an isoprenoid chain by the tRNA‐ipt gene. In plant plastids and bacteria, the methyl erythritol (MEP) pathway is the main source of the prenyl group, but in the cytosol of plants and insects, the prenyl group is synthesized via the mevalonate (MVA) pathway. Cytokinins are produced upon the degradation of the prenylated tRNA. To further investigate the role of cytokinins in gall induction by insects, I performed gene expression experiments using the gall‐inducing Eurosta solidaginis and its bacterial symbiont Wolbachia sp. I also performed targeted gene expression analyses in order to examine the expression of genes in the MEP, MVA, and tRNA‐ipt pathways. Up‐regulation of these pathways in the salivary gland implies that cytokinins are being synthesized in the salivary glands by either the insect and/or its bacterial symbiont. Along with the known widespread distribution of cytokinins in insects, these further imply that cytokinin production and secretion has been amplified and is used by the insects to manipulate their host plants. However, the genes involved in cytokinin production have also been found in other insect groups, so this plant manipulation may not just be exclusive to gall‐inducing insects, suggesting widespread hostplant manipulation among plant‐eating insects.Support or Funding InformationNIH MBRS‐RISE: R25‐GM059298

Detecting and quantifying oligomerization of ARF PB1 Domains

Auxin response factors (ARFs) are a family of transcription factors within plants that serve to regulate the plant’s response to auxin, a growth initiating hormone. ARFs can either serve as transcription activators or repressors to auxin regulation in the presence of auxin. The majority ARFs have 3 domains: a PB1 protein interaction domain, the low‐complexity middle region, and a DNA binding domain. Major differences between ARF activators and repressors lay in large sequence differences in the middle region, but sequence analysis of the PB1s domains show key differences with implications for their role in transcription regulation. PB1 domains are responsible the interactions necessary for auxin regulation. All ARF PB1 domains are type I/II meaning they contain both a positive and negative residue on opposite sides for electrostatic interactions with copies of themselves or other PB1 domains. The potential for bidentate interaction interfaces complicates the model of auxin response by requiring us to consider oligomerization through the PB1 domain including the potential for hetero‐oligomerizatin between ARF activators and repressors. The work examines the degree of oligomerization and quantifies dimerization affinity in both the wild‐type ARF5 PB1, a transcription activator, and the ARF1 PB1, a transcription repressor. More specifically, to answer these questions crosslinking, gel filtration, and fluorescence anisotropy were used to compare a broad range of concentrations for analysis as well as to improve quantification. Both wild‐type ARF5 and ARF1 self‐associated as expected under varying conditions, forming small oligomers, but ARF5 remained primarily in the dimer state. This work suggests a more nuanced model of transcriptional regulation depending on the specific population of regulatory PB1s present.