Ethylene inhibits rice root elongation in compacted soil via ABA- and auxin-mediated mechanisms.

Soil compaction represents a major agronomic challenge, inhibiting root elongation and impacting crop yields. Roots use ethylene to sense soil compaction as the restricted air space causes this gaseous signal to accumulate around root tips. Ethylene inhibits root elongation and promotes radial expansion in compacted soil, but its mechanistic basis remains unclear. Here, we report that ethylene promotes abscisic acid (ABA) biosynthesis and cortical cell radial expansion. Rice mutants of ABA biosynthetic genes had attenuated cortical cell radial expansion in compacted soil, leading to better penetration. Soil compaction-induced ethylene also up-regulates the auxin biosynthesis gene OsYUC8. Mutants lacking OsYUC8 are better able to penetrate compacted soil. The auxin influx transporter OsAUX1 is also required to mobilize auxin from the root tip to the elongation zone during a root compaction response. Moreover, osaux1 mutants penetrate compacted soil better than the wild-type roots and do not exhibit cortical cell radial expansion. We conclude that ethylene uses auxin and ABA as downstream signals to modify rice root cell elongation and radial expansion, causing root tips to swell and reducing their ability to penetrate compacted soil.

Root-Expressed Rice PAP3b Enhances Secreted APase Activity and Helps Utilize Organic Phosphate.

Phosphate (Pi) deficiency leads to the induction of purple acid phosphatases (PAPs) in plants, which dephosphorylate organic phosphorus (P) complexes in the rhizosphere and intracellular compartments to release Pi. In this study, we demonstrate that OsPAP3b belongs to group III low-molecular weight PAP and is low Pi-responsive, preferentially in roots. The expression of OsPAP3b is negatively regulated with Pi resupply. Interestingly, OsPAP3b was found to be dual localized to the nucleus and secretome. Furthermore, OsPAP3b is transcriptionally regulated by OsPHR2 as substantiated by DNA-protein binding assay. Through in vitro biochemical assays, we further demonstrate that OsPAP3b is a functional acid phosphatase (APase) with broad substrate specificity. The overexpression (OE) of OsPAP3b in rice led to increased secreted APase activity and improved mineralization of organic P sources, which resulted in better growth of transgenics compared to the wild type when grown on organic P as an exogenous P substrate. Under Pi deprivation, OsPAP3b knock-down and knock-out lines showed no significant changes in total P content and dry biomass. However, the expression of other phosphate starvation-induced genes and the levels of metabolites were found to be altered in the OE and knock-down lines. In addition, in vitro pull-down assay revealed multiple putative interacting proteins of OsPAP3b. Our data collectively suggest that OsPAP3b can aid in organic P utilization in rice. The APase isoform behavior and nuclear localization indicate its additional role, possibly in stress signaling. Considering its important roles, OsPAP3b could be a potential target for improving low Pi adaptation in rice.

OsJAZ11 regulates phosphate starvation responses in rice.

MAIN CONCLUSION: OsJAZ11 regulates phosphate homeostasis by suppressing jasmonic acid signaling and biosynthesis in rice roots. Jasmonic Acid (JA) is a key plant signaling molecule which negatively regulates growth processes including root elongation. JAZ (JASMONATE ZIM-DOMAIN) proteins function as transcriptional repressors of JA signaling. Therefore, targeting JA signaling by deploying JAZ repressors may enhance root length in crops. In this study, we overexpressed JAZ repressor OsJAZ11 in rice to alleviate the root growth inhibitory action of JA. OsJAZ11 is a low phosphate (Pi) responsive gene which is transcriptionally regulated by OsPHR2. We report that OsJAZ11 overexpression promoted primary and seminal root elongation which enhanced Pi foraging. Expression studies revealed that overexpression of OsJAZ11 also reduced Pi starvation response (PSR) under Pi limiting conditions. Moreover, OsJAZ11 overexpression also suppressed JA signaling and biosynthesis as compared to wild type (WT). We further demonstrated that the C-terminal region of OsJAZ11 was crucial for stimulating root elongation in overexpression lines. Rice transgenics overexpressing truncated OsJAZ11ΔC transgene (i.e., missing C-terminal region) exhibited reduced root length and Pi uptake. Interestingly, OsJAZ11 also regulates Pi homeostasis via physical interaction with a key Pi sensing protein, OsSPX1. Our study highlights the functional connections between JA and Pi signaling and reveals JAZ repressors as a promising candidate for improving low Pi tolerance of elite rice genotypes.

OsJAZ9 overexpression modulates jasmonic acid biosynthesis and potassium deficiency responses in rice.

KEY MESSAGE: Enhanced bioactive JA (JA-Ile) accumulation in OsJAZ9 overexpressing rice helps plants tolerate K deficiency. Potassium (K) represents up to 10% of the plant's total dry biomass, and its deficiency makes plants highly susceptible to both abiotic and biotic stresses. K shortage results in the inhibition of root and shoots growth, but the underlying mechanism of this response is unclear. Our RNA-Seq and qPCR analysis suggested leading roles for JA pathway genes under K deficiency in rice. Notably, K deficiency and JA application produced similar phenotypic and transcriptional responses. Here, we integrated molecular, physiological and morphological studies to analyze the role of OsJAZ9 in JA homeostasis and K deficiency responses. We raised OsJAZ9 over-expression, knockdown, transcriptional reporter, translational reporter and C-terminal deleted translational reporter lines in rice to establish the role of JA signaling in K ion homeostasis. JA profiling revealed significantly increased JA-Ile levels in OsJAZ9 OE lines under K deficiency. Furthermore, we established that OsJAZ9 overexpression and knockdown result in K deficiency tolerance and sensitivity, respectively, by modulating various K transporters and root system architecture. Our data provide evidence on the crucial roles of OsJAZ9 for improving K deficiency tolerance in rice by altering JA levels and JA responses.

A novel glycerophosphodiester phosphodiesterase improves phosphate deficiency tolerance in rice.

Soil phosphate (Pi) deficiency is major constraint for rice cultivation worldwide. Cellular membranes account for one third of cellular organic phosphorus (P) in the form of phospholipids. Therefore, remobilization of Pi from membrane phospholipids under Pi deficiency can be an important strategy to improve phosphorus use efficiency. Glycerophosphodiester phosphodiesterases (GDPDs) hydrolyse intermediate product of phospholipid catabolism, glycerophosphodiesters to glycerol-3-phosphate, a precursor for P and non P-lipid biosynthesis. Here, we show that OsGDPD2 is a Pi deficiency responsive gene, which is transcriptionally regulated by OsPHR2. In silico analysis of active site residues and enzymatic assays confirmed phosphodiesterase activity of OsGDPD2. All overexpression lines showed higher GDPD activity, Pi content, root growth, and biomass accumulation as compared with wild type. Conversely, silencing of OsGDPD2 led to decreased GDPD activity and Pi content. Notably, most of the P-containing metabolites and fatty acids were elevated in transgenic lines. Further, quantitative analysis of polar lipids revealed higher accumulation of several classes of phospholipids and galactolipids in overexpression lines indicating a potential role of OsGDPD2 in de novo glycerolipid biosynthesis. Thus, present study provides insights into novel physiological roles of OsGDPD2 in low Pi acclimation in rice.

OsHAD1, a Haloacid Dehalogenase-Like APase, Enhances Phosphate Accumulation.

Phosphorus (P) deficiency limits plant growth and yield. Since plants can absorb only the inorganic form of P (Pi), a large portion of soil P (organic and inorganic P complexes) remains unused. Here, we identified and characterized a PHR2-regulated, novel low-Pi-responsive haloacid dehalogenase (HAD)-like hydrolase, OsHAD1 While OsHAD1 is a functional HAD protein having both acid phosphatase and phytase activities, it showed little homology with other known low-Pi-responsive HAD superfamily members. Recombinant OsHAD1 is active at acidic pH and dephosphorylates a broad range of organic and inorganic P-containing substrates, including phosphorylated serine and sodium phytate. Exogenous application of recombinant OsHAD1 protein in growth medium supplemented with phytate led to marked increases in growth and total P content of Pi-deficient wild-type rice (Oryza sativa) seedlings. Furthermore, overexpression of OsHAD1 in rice resulted in enhanced phosphatase activity, biomass, and total and soluble P contents in Pi-deficient transgenic seedlings treated with phytate as a restricted Pi source. Gene expression and metabolite profiling revealed enhanced Pi starvation responses, such as up-regulation of multiple genes involved in Pi uptake and solubilization, accumulation of organic acids, enhanced secretory phosphatase activity, and depletion of ATP in overexpression lines as compared with the wild type. To elucidate the underlying regulatory mechanisms of OsHAD1, we performed in vitro pull-down assays, which revealed the association of OsHAD1 with protein kinases such as OsNDPKs. We conclude that, besides dephosphorylation of cellular organic P, OsHAD1 in coordination with kinases may regulate the phosphorylation status of downstream targets to accomplish Pi homeostasis under limited Pi supply.

Morpho-physiological and transcriptome profiling reveal novel zinc deficiency-responsive genes in rice.

Intensive farming has depleted the soil zinc (Zn) availability resulting in decreased crop productivity. Here, we attempt to understand the Zn deficiency response in rice through temporal transcriptome analysis. For this, rice seedlings were raised under Zn-deficient conditions up to 4 weeks followed by Zn re-supply for 3 days. Zn-deficient plants developed characteristic deficiency symptoms such as leaf bronzing, decrease in biomass, total chlorophyll, PSII efficiency, decreased carbonic anhydrase activity and increased ROS production. Interestingly, severe alterations in root system architecture were also observed. Comprehensive transcriptome analyses of rice seedlings were carried out after 2 (DEF2W) and 4 weeks (DEF4W) of Zn deficiency with respect to transcriptome profiles of corresponding Zn sufficient conditions (SUF2W, SUF4W). Additionally, to detect the potential Zn-responsive genes, transcriptome profile of Zn-recovered seedlings was compared with DEF4W. All differentially expressed Zn-responsive genes were categorized into early and late Zn deficiency response, and a set of 77 genes, induced and repressed on Zn deficiency and re-supply, respectively, was identified. These genes could be used as low Zn-responsive marker genes. Further, genes involved in membrane transport, phytosiderophore activity and organic acid biosynthesis showed high differential expression. Additionally, the present study unravelled several genes putatively associated with alterations in root system architecture under Zn deficiency and provides novel insights into the interpretation of morpho-physiological, biochemical and molecular regulation of zinc deficiency responses in rice.

Improvement in phosphate acquisition and utilization by a secretory purple acid phosphatase (OsPAP21b) in rice.

Phosphate (Pi) deficiency in soil system is a limiting factor for rice growth and yield. Majority of the soil phosphorus (P) is organic in nature, not readily available for root uptake. Low Pi-inducible purple acid phosphatases (PAPs) are hypothesized to enhance the availability of Pi in soil and cellular system. However, information on molecular and physiological roles of rice PAPs is very limited. Here, we demonstrate the role of a novel rice PAP, OsPAP21b in improving plant utilization of organic-P. OsPAP21b was found to be under the transcriptional control of OsPHR2 and strictly regulated by plant Pi status at both transcript and protein levels. Biochemically, OsPAP21b showed hydrolysis of several organophosphates at acidic pH and possessed sufficient thermostability befitting for high-temperature rice ecosystems with acidic soils. Interestingly, OsPAP21b was revealed to be a secretory PAP and encodes a distinguishable major APase (acid phosphatase) isoform under low Pi in roots. Further, OsPAP21b-overexpressing transgenics showed increased biomass, APase activity and P content in both hydroponics supplemented with organic-P sources and soil containing organic manure as sole P source. Additionally, overexpression lines depicted increased root length, biomass and lateral roots under low Pi while RNAi lines showed reduced root length and biomass as compared to WT. In the light of these evidences, present study strongly proposes OsPAP21b as a useful candidate for improving Pi acquisition and utilization in rice.

OsMED14_2, a tail module subunit of Mediator complex, controls rice development and involves jasmonic acid.

The Mediator complex is essential for eukaryotic transcription, yet its role and the function of its individual subunits in plants, especially in rice, remain poorly understood. Here, we investigate the function of OsMED14_2, a subunit of the Mediator tail module, in rice development. Overexpression and knockout of OsMED14_2 resulted in notable changes in panicle morphology and grain size. Microscopic analysis revealed impact of overexpression on pollen maturation, reflected by reduced viability, irregular shapes, and aberrant intine development. OsMED14_2 was found to interact with proteins involved in pollen development, namely, OsMADS62, OsMADS63 and OsMADS68, and its overexpression negatively affected the expression of OsMADS68 and the expression of other genes involved in intine development, including OsCAP1, OsGCD1, OsRIP1, and OsCPK29. Additionally, we found that OsMED14_2 overexpression influences jasmonic acid (JA) homeostasis, affecting bioactive JA levels, and expression of OsJAZ genes. Our data suggest OsMED14_2 may act as a regulator of JA-responsive genes through its interactions with OsHDAC6 and OsJAZ repressors. These findings contribute to better understanding of the Mediator complex's role in plant traits regulation.

Turning up the volume: How root branching adaptive responses aid water foraging.

Access to water is critical for all forms of life. Plants primarily access water through their roots. Root traits such as branching are highly sensitive to water availability, enabling plants to adapt their root architecture to match soil moisture distribution. Lateral root adaptive responses hydropatterning and xerobranching ensure new branches only form when roots are in direct contact with moist soil. Root traits are also strongly influenced by atmospheric humidity, where a rapid drop leads to a promotion of root growth and branching. The plant hormones auxin and/or abscisic acid (ABA) play key roles in regulating these adaptive responses. We discuss how these signals are part of a novel "water-sensing" mechanism that couples hormone movement with hydrodynamics to orchestrate root branching responses.

A novel Ca2+ sensor switch for elevated salt tolerance in plants.

Calcium signaling is vital for sensing and alleviating salt stress in plants. In this issue of Developmental Cell, Steinhorst et al. show that salt stress quantitatively translates into an increasing Ca2+ signaling output that activates the CBL8-CIPK24-SOS1 module, which, functioning with CBL4-CIPK24-SOS1, confers enhanced salt tolerance under severe salinity stress.

OsJAZ11 regulates spikelet and seed development in rice.

Seed size is one of the major determinants of seed weight and eventually, crop yield. As the global population is increasing beyond the capacity of current food production, enhancing seed size is a key target for crop breeders. Despite the identification of several genes and QTLs, current understanding about the molecular regulation of seed size/weight remains fragmentary. In the present study, we report novel role of a jasmonic acid (JA) signaling repressor, OsJAZ11 controlling rice seed width and weight. Transgenic rice lines overexpressing OsJAZ11 exhibited up to a 14% increase in seed width and ~30% increase in seed weight compared to wild type (WT). Constitutive expression of OsJAZ11 dramatically influenced spikelet morphogenesis leading to extra glume-like structures, open hull, and abnormal numbers of floral organs. Furthermore, overexpression lines accumulated higher JA levels in spikelets and developing seeds. Expression studies uncovered altered expression of JA biosynthesis/signaling and MADS box genes in overexpression lines compared to WT. Yeast two-hybrid and pull-down assays revealed that OsJAZ11 interacts with OsMADS29 and OsMADS68. Remarkably, expression of OsGW7, a key negative regulator of grain size, was significantly reduced in overexpression lines. We propose that OsJAZ11 participates in the regulation of seed size and spikelet development by coordinating the expression of JA-related, OsGW7 and MADS genes.

Hydraulic flux-responsive hormone redistribution determines root branching.

Plant roots exhibit plasticity in their branching patterns to forage efficiently for heterogeneously distributed resources, such as soil water. The xerobranching response represses lateral root formation when roots lose contact with water. Here, we show that xerobranching is regulated by radial movement of the phloem-derived hormone abscisic acid, which disrupts intercellular communication between inner and outer cell layers through plasmodesmata. Closure of these intercellular pores disrupts the inward movement of the hormone signal auxin, blocking lateral root branching. Once root tips regain contact with moisture, the abscisic acid response rapidly attenuates. Our study reveals how roots adapt their branching pattern to heterogeneous soil water conditions by linking changes in hydraulic flux with dynamic hormone redistribution.

Identification of Purple Acid Phosphatases in Chickpea and Potential Roles of CaPAP7 in Seed Phytate Accumulation.

Purple acid phosphatases (PAPs) play important roles in phosphate (Pi) acquisition and utilization. These PAPs hydrolyze organic Phosphorus (P) containing compounds in rhizosphere as well as inside the plant cell. However, roles of PAPs in one of the most widely cultivated legumes, chickpea (Cicer arietnum L.), have not been unraveled so far. In the present study, we identified 25 putative PAPs in chickpea (CaPAPs) which possess functional PAP motifs and domains. Differential regulation of CaPAPs under different nutrient deficiencies revealed their roles under multiple nutrient stresses including Pi deficiency. Interestingly, most of the CaPAPs were prominently expressed in flowers and young pods indicating their roles in flower and seed development. Association mapping of SNPs underlying CaPAPs with seed traits revealed significant association of low Pi inducible CaPAP7 with seed weight and phytate content. Biochemical characterization of recombinant CaPAP7 established it to be a functional acid phosphatase with highest activity on most abundant organic-P substrate, phytate. Exogenous application of recombinant CaPAP7 enhanced biomass and Pi content of Arabidopsis seedlings supplemented with phytate as sole P source. Taken together, our results uncover the PAPs in chickpea and potential roles of CaPAP7 in seed phytate accumulation.

Genome-wide DNA polymorphisms in low Phosphate tolerant and sensitive rice genotypes.

Soil Phosphorus (P) deficiency is one of the major challenges to rice crop world-wide. Modern rice genotypes are highly P-responsive and rely on high input of P fertilizers. However, low P tolerant traditional cultivars and landraces have genetic potential to sustain well under low P. Identification of high resolution DNA polymorphisms (SNPs and InDels) in such contrasting genotypes is largely missing for low P response at gene levels. Here, we report high quality DNA polymorphisms in low P sensitive genotype, PB1 and tolerant traditional genotype, Dular. We performed whole genome resequencing using Illumina NGS platform and identified a total of 5,157,939 sequence variants in PB1 and Dular with reference to Nipponbare genome. We have identified approximately 2.3 million and 2.9 million high quality polymorphisms in PB1 and Dular, respectively, with an average read depth of ≥24X. We further mapped several DNA polymorphisms (non-synonymous and regulatory variants) having potential functional significance to key Phosphate Starvation Responsive (PSR) and root architecture genes in Dular and Kasalath using a compiled list of low P responsive genes. These identified variants can serve as a useful source of genetic variability for improving low P tolerance and root architecture of high yielding modern genotypes.

Comparative Morphophysiological Analyses and Molecular Profiling Reveal Pi-Efficient Strategies of a Traditional Rice Genotype.

Phosphate (Pi) deficiency severely affects crop yield. Modern high yielding rice genotypes are sensitive to Pi deficiency whereas traditional rice genotypes are naturally compatible with low Pi ecosystems. However, the underlying molecular mechanisms for low Pi tolerance in traditional genotypes remain largely elusive. To delineate the molecular mechanisms for low Pi tolerance, two contrasting rice genotypes, Dular (low Pi tolerant), and PB1 (low Pi sensitive), have been selected. Comparative morphophysiological, global transcriptome and lipidome analyses of root and shoot tissues of both genotypes grown under Pi deficient and sufficient conditions revealed potential low Pi tolerance mechanisms of the traditional genotype. Most of the genes associated with enhanced internal Pi utilization (phospholipid remobilization) and modulation of root system architecture (RSA) were highly induced in the traditional rice genotype, Dular. Higher reserves of phospholipids and greater accumulation of galactolipids under low Pi in Dular indicated it has more efficient Pi utilization. Furthermore, Dular also maintained greater root growth than PB1 under low Pi, resulting in larger root surface area due to increased lateral root density and root hair length. Genes involved in enhanced low Pi tolerance of the traditional genotype can be exploited to improve the low Pi tolerance of modern high yielding rice cultivars.

Physical localization and probable transcriptional activity of 18S-5.8S-26S rRNA gene loci in some Asiatic Cymbidiums (Orchidaceae) from north-east India.

Fluorescence in situ hybridization based physical localization of 45S ribosomal DNA in eight horticulturally important species of Cymbidium (Orchidaceae) from north-east India (South-East Asia) has been carried for the first time. Observations revealed only one pair of chromosomes had NOR loci. Three, out of eight Cymbidiums showed decondensed, dispersed, extended form of hybridization signals of rDNA as dots of fluorescence (transcriptionally active), where as the rest of the Cymbidiums revealed condensed (non-active) forms, hence demonstrated the heteromorphism in size, intensities and their appurtenance which may be under epigenetic control. Except for the ribosomal genes, no other active genes have been reported to reside within the nucleoli. Such observations provide useful chromosome landmarks and provide valuable evidence about the genome evolution, speciation and ploidy both at molecular and chromosomal levels which is more or less highly ambiguous in family Orchidaceae.

Auditory neuropathy spectrum disorder: its prevalence and audiological characteristics in an Indian tertiary care hospital.

OBJECTIVE: Auditory neuropathy spectrum disorder (ANSD) has become a well accepted clinical entity. ANSD has now become easy to recognize with advent of test battery approach for hearing loss evaluation. However, little is known about its epidemiology and aetiology, especially its prevalence and incidence in India. This study was conducted to determine the prevalence and audiological characteristics of ANSD in our tertiary care hospital at New Delhi, India. METHODS: We prospectively enrolled all the paediatric cases i.e. less than 12years of age referred to our outpatient department for hearing evaluation. All the cases were evaluated using complete audiological test battery consisting of pure tone audiometry (PTA), impedance audiometry, acoustic reflexes, otoacoustic emissions (OAE), and brain stem evoked response audiometry (BERA). The instrument used was GSI Audera. BERA with both condensation and rarefaction polarity was performed in cases with absent wave V even at highest intensity levels. The criteria used for diagnosis of ANSD was normal or near normal cochlear hair cells (sensory) function (preservation of otoacoustic emissions and/or cochlear microphonics) and absent or abnormal auditory nerve function (absent or severely abnormal auditory brainstem potentials). A repeat test battery was performed after 3months on the diagnosed cases of auditory neuropathy. Based on their behavioural findings the appropriate management programme was formulated. RESULTS: A total of 487 paediatric cases were referred for hearing evaluation. 183 (37.6%) cases showed absent BERA and 26(5.3%) cases fulfilled the diagnostic criteria for ANSD. Repeat examination after 3months revealed presence of OAE's in 57.6% (15/26) cases while cochlear microphonics were present in all the 26 cases. CONCLUSION: The prevalence of ANSD in our study is 5.3% and in children diagnosed with severe to profound hearing loss is 14%. Presence of cochlear microphonics with absent BERA seems to be reliable criteria for diagnosing ANSD.