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Differentially expressed genes obtained from RNA-sequencing during different phases of adventitious root formation in A. euchroma
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Background
Adventitious root formation is considered a major developmental step during the propagation of difficult to root plants, especially in horticultural crops. Recently, adventitious roots induced through plant tissue culture methods have also been used for production of phytochemicals such as flavonoids, anthocyanins and anthraquinones. It...
Citations
... Because of the largely unknown, complex and unexplored genome of high-altitude medicinal plants, deciphering the underlying mechanism is crucial. Sequencing of mRNA through a highthroughput transcriptomics approach provides efficient profiling of differentially expressed genes (DEGs), cellular, biological and molecular processes, metabolic pathways, transcription factors (TFs) and evolutionary studies (Barros et al., 2021;Devi et al., 2021). In our previous experiment with micro-propagated plantlets of N. jatamansi treated with two differentially coated AuNPs, namely citrate (Àve charge) and CTAB (+ve charge), we observed improved total biomass, antioxidant potential and hormonal profiling (Joshi et al., 2022a). ...
Nardostachys jatamansi is a Himalayan medicinal plant known for its excellent antioxidant and metabolic properties. Differentially charged gold nanoparticles were reported earlier to play immense role in improvement of organogenesis and cellular homeostasis in different plant species. In this study, efforts were made to unveil the molecular programming underpinning the elicitation response of charged gold nanoparticles in N. jatamansi under in vitro conditions. RNA sequencing of in vitro developed shoots subjected to control, 60 mM citrate-AuNPs and 40 mM CTAB-AuNPs, generated 217.1 million sequenced reads that identified 104, 1255 and 660 significantly differentially expressed genes in non-treated control vs 60 mM citrate-AuNPs, non-treated control vs 40 mM CTAB-AuNPs and 60 mM citrate-AuNPs vs 40 mM CTAB-AuNPs libraries, respectively. Gene ontology (GO) analysis revealed that major biological, cellular and metabolic processes were affected more by CTAB-AuNP treatment than by citrate-AuNP treatment. Several DEGs were identified having key role in growth, antioxidant potential, hormonal regulation, metabolite production, and stress response. KEGG analysis revealed differentially expressed genes associated with photosynthesis, antioxidant activity, hormone metabolism, cellular internalisation, secondary metabolites biosynthesis and defence response. Transcription factors regulating these pathways such as Myb, WRKY, NAC, ERF, bHLH and dof were reported to be upregulated. In conclusion, positively charged CATB-AuNPs were found to be more effective than negatively charged Citrate-AuNPs for in vitro propagation. Thus, besides providing insights into molecular dynamics underlying in vitro nono-elicitation affecting proliferation of N. jatamansi, our study also provides ample genomic resources for the in vitro improvement of medicinal plants.
... The adventitious roots in T. cordifolia were found to be evolved due to the presence of signatures of adaptive evolution in these genes involved in adventitious root formation (Supplementary Table S17). These genes with evolutionary signatures were also reported in other plant species and play key roles in adventitious root formation [92][93][94] . Additionally, phytohormones also play an important role in regulating adventitious root formation in plants 95 . ...
Tinospora cordifolia (Willd.) Hook.f. & Thomson, also known as Giloy, is among the most important medicinal plants that have numerous therapeutic applications in human health due to the production of a diverse array of secondary metabolites. To gain genomic insights into the medicinal properties of T. cordifolia, the genome sequencing was carried out using 10× Genomics linked read and Nanopore long-read technologies. The draft genome assembly of T. cordifolia was comprised of 1.01 Gbp, which is the genome sequenced from the plant family Menispermaceae. We also performed the genome size estimation for T. cordifolia, which was found to be 1.13 Gbp. The deep sequencing of transcriptome from the leaf tissue was also performed. The genome and transcriptome assemblies were used to construct the gene set, resulting in 17,245 coding gene sequences. Further, the phylogenetic position of T. cordifolia was also positioned as basal eudicot by constructing a genome-wide phylogenetic tree using multiple species. Further, a comprehensive comparative evolutionary analysis of gene families contraction/expansion and multiple signatures of adaptive evolution was performed. The genes involved in benzyl iso-quinoline alkaloid, terpenoid, lignin and flavonoid biosynthesis pathways were found with signatures of adaptive evolution. These evolutionary adaptations in genes provide genomic insights into the presence of diverse medicinal properties of this plant. The genes involved in the common symbiosis signalling pathway associated with endosymbiosis (Arbuscular Mycorrhiza) were found to be adaptively evolved. The genes involved in adventitious root formation, peroxisome biogenesis, biosynthesis of phytohormones, and tolerance against abiotic and biotic stresses were also found to be adaptively evolved in T. cordifolia.
... This balance of high levels of IAA and low levels of BR promoted the formation and development of AR. Studies of AR formation in purple grass, sugarcane and mulberry cuttings showed significant differences in the expression of several genes involved in BR and IAA signaling pathways, similar to the results of the present study (Devi et al. 2021;Li et al. 2020;Cao et al. 2018). ...
The generation of adventitious roots (ARs) is the key to the success of cuttings. The appropriate environment for AR differentiation in tea plants is acidic. However, the mechanism is unclear. In this study, pH 4.5 was suitable condition for the differentiation of AR in tea plants. At the base of cuttings, the root primordia differentiated ARs more rapidly at pH 4.5 than pH 7.0, and nine AR differentiation-related genes were found to be differentially expressed in 30 days, the result was also validated by qRT-PCR. The promoter regions of these genes contained auxin and brassinosteroid response elements. The expression levels of several genes which were involved in auxin and brassinosteroid synthesis as well as signaling at pH 4.5 compared to pH 7.0 occurred differential expression. Brassinolide (BL) and indole-3-acetic acid (IAA) could affect the differentiation of ARs under pH 4.5 and pH 7.0. By qRT-PCR analysis of genes during ARs generation, BL and IAA inhibited and promoted the expression of CsIAA14 gene, respectively, to regulate auxin signal transduction. Meanwhile, the expression levels of CsKNAT4, CsNAC2, CsNAC100, CsWRKY30 and CsLBD18 genes were up-regulated upon auxin treatment and were positively correlated with ARs differentiation.This study showed that pH 4.5 was the most suitable environment for the root primordia differentiation of AR in tea plant. Proper acidic pH conditions promoted auxin synthesis and signal transduction. The auxin initiated the expression of AR differentiation-related genes, and promoted its differentiated. BL was involved in ARs formation and elongation by regulating auxin signal transduction.
... Auxin is considered a key factor and the most common PGR for in vitro root induction [23]. Roots grown under in vitro conditions are technically considered adventitious roots because they do not develop from primary roots [32]. Adventitious root growth occurs in three stages: induction, initiation, and expression, and several genes involved in these phases are regulated by auxin [33]. ...
... During the induction phase, NAA induces root founder cells (AUX/IAA) family proteins. Te degradation allows the AUXIN RESPONSE FACTOR (ARF) gene family to upregulate GRETCHEN3 (GH3) genes and promote adventitious root formation [32,33]. During adventitious root initiation, the application of NAA led to the activation of WUSCHEL-RELATED HOMEOBOX 11 and 12 (WOX11/12) genes. ...
Pokeweed, Phytolacca americana L., is considered a widely spreading invasive plant, while saponin contents accumulated in the roots have pharmaceutical uses, such as rheumatism treatments and anti-inflammation. Adventitious root cultures are an important source of diverse secondary metabolites, which have significant applications in various fields. This study focused on the optimization of parameters for root induction using different medium states and treatments with auxins on a pokeweed leaf. Semisolid and liquid MS (Murashige and Skoog, 1962) media were supplemented with indole-3-butyric acid (IBA) and 1-naphthylacetic acid (NAA) at 0.5, 1, 2, and 4 mg/L. Root growth parameters, e.g., induction percentage, root numbers, length, and weight, were measured to determine the adventitious root induction efficiency. Total phenolic content, total flavonoid content, total saponin content, and antioxidant activity were recorded. Results showed that adventitious roots induced in semisolid MS medium supplemented with 0.5 mg/L NAA exhibited a high density of lateral roots. Appropriate medium state and auxin for adventitious root induction in pokeweed were determined as semisolid medium supplemented with 2 mg/L NAA. Considering phytochemicals, adventitious roots induced in liquid medium containing 0.5–1 mg/L NAA had the highest yield extract percentage. Additionally, adventitious roots cultivated in a liquid medium enriched with 1 mg/L NAA exhibited the highest phenolic and saponin contents. A principal component analysis (PCA) biplot and hierarchical cluster analysis (HCA) heatmap demonstrated different response patterns between semisolid and liquid media applied with NAA. The results of the semisolid media were grouped together due to high expression levels of the root induction parameters, while elevated phytochemical values were observed in the liquid media treatments. The results suggested two different media that provide the highest adventitious root induction efficiency and the greatest phytochemical contents: semisolid medium with 2 mg/L NAA and liquid medium with 1 mg/L NAA, respectively. These culture media can be applied to optimize adventitious root culture of pokeweed and in vitro phytochemical production.
... However, we do not know how to promote adventitious rooting by regulating the expression of genes related to plant hormone signalling. Crosstalk with other hormones plays a significant role at different developmental stages of AR formation either in combination or individually [7,22]. Although time scales vary between species, auxin has been confirmed as a mediator of AR formation by interacting with other plant hormones, such as CTK, ET, JA, SA and BRs. ...
... Although time scales vary between species, auxin has been confirmed as a mediator of AR formation by interacting with other plant hormones, such as CTK, ET, JA, SA and BRs. However, ET, auxin, JA, GA, CTK and ABA affect AR formation at the induction stage [22][23][24]. Populus is an important commercial tree, and AR formation is a major setback to its asexual reproduction. ...
... Most tree species share a common trait that auxin is closely related to the start of AR formation [8,22,24,25]. Auxin signalling acts through three transcription factors, AUX/IAA, GH 3 and SAUR, from the ARF gene family in Arabidopsis thaliana, in which AtARF17 positively regulates the downstream GH3 family, while AtARF6/8 negatively regulates the downstream GH3 family [31]. This study shows the upregulation and downregulation of AUX, AUX/IAA, ARF, GH 3 and SAUR (Figures 3 and 4 and Tables S1 and S2). ...
Adventitious roots (ARs) occur naturally in many species and are important for plants to absorb nutrients and water. AR formation can also be induced from explants of trees, whose clonal propagation is needed. AR formation is gridlock for many woody plant mass propagations. Plant hormones have been regarded as playing a key role in AR formation, and the molecular regulatory mechanisms need to be elucidated. In this study, RNA-Seq was performed to reveal the molecular mechanisms in the different periods of AR formation from hybrid poplar clone 84K (Populus alba × P. glandulosa) and AUXIN SIGNALING F-BOX (PagFBL1-OE). To understand the importance of differentially expressed genes (DEGs), we found that many genes involved in signal transduction mechanisms were induced at 12, 24 and 48 h in 84K and PagFBL1-OE cells by NOG classification. We also found that many DEGs were enriched in hormone signal transduction only for the first 12 h in 84K and PagFBL1-OE by KEGG pathway enrichment. Notably, more DEGs appeared in indole-3-acetic acid (IAA), abscisic acid (ABA), ethylene (ETH), jasmonic acid (JA), brassinolide (BR), cytokinin (CTK) and gibberellin (GA) signal transduction for the first 12 h in PagFBL1-OE than in 84K. Moreover, ARF (Pop_G01G075686), IAA14 (Pop_A10G047257), SAURs (Pop_A03G019756, Pop_A12G067965, Pop_G03G055849 and Pop_G12G008821), JAR1s (Pop_A14G000375 and Pop_G14G044264), CTR1 (Pop_A17G052594 and Pop_G09G030293), CRE1s (Pop_G07G086605 and Pop_G07G086618), GID1 (Pop_A04G026477), BKI1 (Pop_A02G066155), PYR/PYLs (Pop_A03G050217 and Pop_G01G089222), and TGAs (Pop_A04G059310, Pop_G04G060065 and Pop_G05G008153) were only specifically expressed in PagFBL1-OE and could play an important role in AR formation, especially in the first 12 h under plant hormone signal transduction. These results show that the complex biological process of AR formation is primarily influenced by the hormone signalling pathway in Populus. This study reveals the initial regulation of AR formation in woody plant cuttings and thus contributes to further elucidating the molecular mechanism by which hormones interact.
... A lack of nutrient elements such as N, P, boron, calcium, iron, etc. impairs cell wall formation and cell growth (Fang et al., 2019;Qin et al., 2019;Zhu et al., 2019;Liu et al., 2021b;Rivai et al., 2021;Chen et al., 2022a;Li et al., 2023;Nezamivand-Chegini et al., 2023). The integrity of plant cell wall also affected the root formation process (Devi et al., 2021). Expansins are cell-wall-loosening proteins, they interact with the primary cell wall, which is composed of cellulose, hemicellulose, pectin, and xyloglucan. ...
Nitrogen (N) and phosphorus (P) are essential phytomacronutrients, and deficiencies in these two elements limit growth and yield in apple (Malus domestica Borkh.). The rootstock plays a key role in the nutrient uptake and environmental adaptation of apple. The objective of this study was to investigate the effects of N and/or P deficiency on hydroponically-grown dwarfing rootstock ‘M9-T337’ seedlings, particularly the roots, by performing an integrated physiological, transcriptomics-, and metabolomics-based analyses. Compared to N and P sufficiency, N and/or P deficiency inhibited aboveground growth, increased the partitioning of total N and total P in roots, enhanced the total number of tips, length, volume, and surface area of roots, and improved the root-to-shoot ratio. P and/or N deficiency inhibited NO 3 − influx into roots, and H⁺ pumps played a important role in the response to P and/or N deficiency. Conjoint analysis of differentially expressed genes and differentially accumulated metabolites in roots revealed that N and/or P deficiency altered the biosynthesis of cell wall components such as cellulose, hemicellulose, lignin, and pectin. The expression of MdEXPA4 and MdEXLB1, two cell wall expansin genes, were shown to be induced by N and/or P deficiency. Overexpression of MdEXPA4 enhanced root development and improved tolerance to N and/or P deficiency in transgenic Arabidopsis thaliana plants. In addition, overexpression of MdEXLB1 in transgenic Solanum lycopersicum seedlings increased the root surface area and promoted acquisition of N and P, thereby facilitating plant growth and adaptation to N and/or P deficiency. Collectively, these results provided a reference for improving root architecture in dwarfing rootstock and furthering our understanding of integration between N and P signaling pathways.
... Several genes have been reported during the AR emergence to be showing a participation in the cell wall remodeling (Rigal et al., 2012). PINFORMED1 (PIN1) and its analogs act as auxin efflux carriers and generate local auxin maxima by coordinating with PIN proteins that play a central role in adventitious root development (Devi et al., 2021). ...
The formation of adventitious root primordia (ARP) is an ecological precious developmental event in the plant. The ARP evolution is a crucial step for the plant to fulfill two important phases of the life cycle, viz., vegetative and reproduction. The identification of several candidate genes has been known to provoke the induction, initiation, and maintenance of ARP-associated signaling cascading network. The expression of various genes, including WOUND INDUCED DEDIFFERENTIATION (WIND) genes induction such as APETALA2/ETHYLENE RESPONSE FACTORs (AP2/ERFs), CLAVATA3/EMBRYO SURROUNDING REGION-RELATED, ABERRANT LATERAL ROOT FORMATION4 (ALF4), was observed to be enhanced during ARP formation. This chapter highlights the histological documentation of adventitious root development, the role of phytohormones in adventitious root (AR) development, the role of sugars in AR development, the role of signaling peptides in AR development, epigenetic control in AR development, and cell wall modification during AR emergence. Additionally, the role of phytohormones and their molecular network is also explained during ARP initiation, elongation, and maturation in different plant species. Furthermore, the genetic signals that play an important role in the crosstalk generate complex network that regulates the generation of ARP.
... An increasing number of studies have shown that signaling regulation by endogenous and exogenous hormones is closely related to AR formation. In Arnebia euchroma and sugarcane, both endogenous and exogenous auxin have been reported to regulate AR formation through similar signaling pathways (Devi et al. 2021;Li et al. 2020). In the current work, we speculated that exogenous IBA application might trigger multiple plant hormone signaling pathways and affect the development of AR. ...
Taxus cuspidata S. et Z is a perennial tree with significant economic and medicinal values. T. cuspidata propagation using cuttings is one of the most efficient approaches to solve its propagation material production on a larger scale. Its adventitious roots play a crucial role in the hardwood cutting propagation. To understand the molecular mechanisms involved in T. cuspidata adventitious root development and thus improve the effectiveness of propagation techniques, we investigated the optimal exogenous hormone applications that can rapidly induce adventitious roots in T. cuspidata hardwood cuttings. The best rooting responses were observed in cuttings treated with indole butyric acid (IBA). Moreover, biochemical and molecular profiling analyses of cuttings treated with different IBA concentrations were carried out. Peroxidase and polyphenol oxidase activities consistently increased with IBA concentration increase, except for a decrease observed at IBA150. Indole-3-acetic acid, gibberellin, and ethylene concentrations were significantly higher in all IBA-treated samples compared with the control group. Comparative transcriptome analysis revealed thousands of differentially expressed genes among the four samples (IBA0, IBA50, IBA100, and IBA150) evaluated. Most differentially expressed genes were assigned to phytohormone signaling pathways and sugar metabolism. The AP2/ERF, bHLH, and MYB transcription factor families and genes involved in root development and cell division were also overrepresented during adventitious root formation. This study provides insights into establishing improved asexual propagation protocols and elucidating into the molecular mechanisms underlying root development in T. cuspidata hardwood cuttings.
... A. euchroma plants were collected from Kibber (32.33 • N, 78.00 • E), Spiti valley, Himachal Pradesh, India and maintained in the polyhouse (20-30 • C annual temperature and 60-70% relative humidity) at CSIR-Institute of Himalayan Bioresource Technology, Palampur, Himachal Pradesh, India. The taxonomically identified plant specimen (Voucher number: PLP18571) was used to establish in vitro shoot cultures from rhizome buds in MS medium as per earlier reported protocol (Devi et al., 2021b). ...
... A. euchroma plants were collected from Kibber (32.33 • N, 78.00 • E), Spiti valley, Himachal Pradesh, India and maintained in the polyhouse (20-30 • C annual temperature and 60-70% relative humidity) at CSIR-Institute of Himalayan Bioresource Technology, Palampur, Himachal Pradesh, India. The taxonomically identified plant specimen (Voucher number: PLP18571) was used to establish in vitro shoot cultures from rhizome buds in MS medium as per earlier reported protocol (Devi et al., 2021b). ...
Arnebia euchroma is a valuable Himalayan herb, whose roots are savagely harvested from the wild to extract red
naphthoquinone pigments with applications in food, cosmetic and pharmaceutical industries. The primary
objective of this study was to induce A. euchroma adventitious roots and use the same for the production of
naphthoquinone pigments. A number of media and auxin types were tried for the formation of adventitious roots
from leaves of A. euchroma under in vitro conditions. The data showed 100% root induction on semi-solid Schenk
& Hildebrandt (SH) medium fortified with 2.5 mg/L indole-3-butyric acid (IBA) after 4-weeks of culture.
Thereafter, factors like inoculum density and sucrose concentration were optimized to maximize adventitious
root growth and pigment production in liquid shake flask cultures. The results revealed a significantly high root
biomass (7.00 ± 0.25 g/L dry weight (DW)) and pigment (192.92 ± 1.61 mg/g DW) yield in SH medium having
10 g/L inoculum density, 0.5 mg/L IBA and 5% sucrose concentration. In addition, the cultivation of adventitious
roots in Balloon-Type Bubble Bioreactor (5 L cap.) was also done, which showed comparably lower root biomass
(4.33 ± 0.09 g/L DW) yield to shake flask culturing, however, resulted in higher pigments accumulation (195.36
± 1.95 mg/g DW). The chemical profile of in vitro induced adventitious roots cultivated in shake flask and
bioreactor was also compared with parent plant rhizome using Ultra Performance Liquid Chromatography. As
per the results, deoxyshikonin and β, β-dimethylacrylshikonin were found to be the key pigments in adventitious
roots, although shikonin was not detectable which otherwise also present in parent plant rhizomes. Considering
the shorter cultivation cycle (4-weeks) and regenerative potential of induced adventitious roots along with
comparable metabolic profile, the developed process seems to be a feasible alternative for the production of
natural pigments.
