Interspecies transfer of syntenic RAMOSA1 orthologs and promoter cis sequences impacts maize inflorescence architecture

Grass inflorescences support floral structures that each bear a single grain, where variation in branch architecture directly impacts yield. The maize RAMOSA1 (ZmRA1) transcription factor acts as a key regulator of inflorescence development by imposing branch meristem determinacy. Here, we show RA1 transcripts accumulate in boundary domains adjacent to spikelet meristems in Sorghum bicolor (Sb) and Setaria viridis (Sv) inflorescences similar as in the developing maize tassel and ear. To evaluate functional conservation of syntenic RA1 orthologs and promoter cis sequences in maize, sorghum and setaria, we utilized interspecies gene transfer and assayed genetic complementation in a common inbred background by quantifying recovery of normal branching in highly ramified ra1-R mutants. A ZmRA1 transgene that includes endogenous upstream and downstream flanking sequences recovered normal tassel and ear branching in ra1-R. Interspecies expression of two transgene variants of the SbRA1 locus, modeled as the entire endogenous tandem duplication or just the non-frameshifted downstream copy, complemented ra1-R branching defects and induced novel fasciation and branch patterns. The SvRA1 locus lacks conserved, upstream noncoding cis sequences found in maize and sorghum; interspecies expression of an SvRA1 transgene did not or only partially recovered normal inflorescence forms. Driving expression of the SvRA1 coding region by the ZmRA1 upstream region, however, recovered normal inflorescence morphology in ra1-R. These data leveraging interspecies gene transfer suggest that cis-encoded temporal regulation of RA1 expression is a key factor in modulating branch meristem determinacy that ultimately impacts grass inflorescence architecture.

Pepper Fruit Elongation Is Controlled by Capsicum annuum Ovate Family Protein 20

Fruit shape is one of the most important quality traits of pepper (Capsicum spp.) and is used as a major attribute for the classification of fruit types. Wide natural variation in fruit shape exists among the major cultivated species Capsicum annuum, allowing the identification of several QTLs controlling the trait. However, to date, no genes underlying fruit shape QTLs have been conclusively identified, nor has their function been verified in pepper. We constructed a mapping population from a cross of round- and elongated-fruited C. annuum parents and identified a single major QTL on chromosome 10, termed fs10, explaining 68 and 70% of the phenotypic variation for fruit shape index and for distal fruit end angle, respectively. The QTL was mapped in several generations and was localized to a 5 Mbp region containing the ortholog of SlOFP20 that suppresses fruit elongation in tomato. Virus-induced gene silencing of the pepper ortholog CaOFP20 resulted in increased fruit elongation on two independent backgrounds. Furthermore, CaOFP20 exhibited differential expression in fs10 near-isogenic lines, as well as in an association panel of elongated- and round-fruited accessions. A 42-bp deletion in the upstream region of CaOFP20 was most strongly associated with fruit shape variation within the locus. Histological observations in ovaries and fruit pericarps indicated that fs10 exerts its effect on fruit elongation by controlling cell expansion and replication. Our results indicate that CaOFP20 functions as a suppressor of fruit elongation in C. annuum and is the most likely candidate gene underlying fs10.

A Cysteine-Rich Protein, SpDIR1L, Implicated in S-RNase-Independent Pollen Rejection in the Tomato (Solanum Section Lycopersicon) Clade

Tomato clade species (Solanum sect. Lycopersicon) display multiple interspecific reproductive barriers (IRBs). Some IRBs conform to the SI x SC rule, which describes unilateral incompatibility (UI) where pollen from SC species is rejected on SI species’ pistils, but reciprocal pollinations are successful. However, SC x SC UI also exists, offering opportunities to identify factors that contribute to S-RNase-independent IRBs. For instance, SC Solanum pennellii LA0716 pistils only permit SC Solanum lycopersicum pollen tubes to penetrate to the top third of the pistil, while S. pennellii pollen penetrates to S. lycopersicum ovaries. We identified candidate S. pennellii LA0716 pistil barrier genes based on expression profiles and published results. CRISPR/Cas9 mutants were created in eight candidate genes, and mutants were assessed for changes in S. lycopersicum pollen tube growth. Mutants in a gene designated Defective in Induced Resistance 1-like (SpDIR1L), which encodes a small cysteine-rich protein, permitted S. lycopersicum pollen tubes to grow to the bottom third of the style. We show that SpDIR1L protein accumulation correlates with IRB strength and that species with weak or no IRBs toward S. lycopersicum pollen share a 150 bp deletion in the upstream region of SpDIR1L. These results suggest that SpDIR1L contributes to an S-RNase-independent IRB.

Fast Particle Acceleration in Three-dimensional Relativistic Reconnection

Magnetic reconnection is invoked as one of the primary mechanisms to produce energetic particles. We employ large-scale 3D particle-in-cell simulations of reconnection in magnetically dominated (σ = 10) pair plasmas to study the energization physics of high-energy particles. We identify an acceleration mechanism that only operates in 3D. For weak guide fields, 3D plasmoids/flux ropes extend along the z-direction of the electric current for a length comparable to their cross-sectional radius. Unlike in 2D simulations, where particles are buried in plasmoids, in 3D we find that a fraction of particles with γ ≳ 3σ can escape from plasmoids by moving along z, and so they can experience the large-scale fields in the upstream region. These “free” particles preferentially move in z along Speiser-like orbits sampling both sides of the layer and are accelerated linearly in time—their Lorentz factor scales as γ ∝ t, in contrast to γ ∝ t in 2D. The energy gain rate approaches ∼eE rec c, where E rec ≃ 0.1B 0 is the reconnection electric field and B 0 the upstream magnetic field. The spectrum of free particles is hard, dN free / d γ ∝ γ − 1.5 , contains ∼20% of the dissipated magnetic energy independently of domain size, and extends up to a cutoff energy scaling linearly with box size. Our results demonstrate that relativistic reconnection in GRB and AGN jets may be a promising mechanism for generating ultra-high-energy cosmic rays.

Dioxin Disrupts Thyroid Hormone and Glucocorticoid Induction of klf9, a Master Regulator of Frog Metamorphosis

Frog metamorphosis, the development of an air-breathing froglet from an aquatic tadpole, is under endocrine control by thyroid hormone (TH) and glucocorticoids (GC). Metamorphosis is susceptible to disruption by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), an aryl hydrocarbon receptor (AHR) agonist. Krüppel-Like Factor 9 (klf9), an immediate early gene in the endocrine-controlled cascade of expression changes that govern metamorphosis, can be synergistically induced by both hormones. This process is mediated by an upstream enhancer cluster, the klf9 synergy module (KSM). klf9 is also a target of the AHR. We measured klf9 mRNA expression following combined exposures to triiodothyronine (T3), corticosterone (CORT), and TCDD in the Xenopus laevis cell line XLK-WG. klf9 was induced 6-fold by 50 nM T3, 4-fold by 100 nM CORT, and 3-fold by 175 nM TCDD. Co-treatments of CORT and TCDD or T3 and TCDD induced klf9 mRNA 7- and 11-fold, respectively, while treatment with all 3 agents induced a 15-fold increase. Transactivation assays examined regulatory sequences from the Xenopus tropicalisklf9 upstream region. KSM-containing segments mediated a strong T3 response and a larger T3/CORT response, while induction by TCDD was mediated by a region ~1 kb farther upstream containing 5 AHR response elements. Unexpectedly, this region also supported a CORT response in the absence of readily- identifiable glucocorticoid responsive elements, suggesting mediation by protein-protein interactions. A similar AHRE cluster is positionally conserved in the human genome, and klf9 was induced by TCDD and TH in HepG2 cells. These results indicate that AHR binding to an upstream AHRE cluster represents an initiating event in TCDD disruption of klf9 expression and metamorphosis.

Use of High Resolution Spatiotemporal Gene Expression Data to Uncover Novel Tissue-Specific Promoters in Tomato

Genetic modification can be an effective strategy for improving the agronomic traits of tomato (Solanum lycopersicum) to meet demands for yield, quality, functional components, and stress tolerance. However, limited numbers of available tissue-specific promoters represent a bottleneck for the design and production of transgenic plants. In the current study, a total of 25 unigenes were collected from an RNA-sequence dataset based on their annotation as being exclusively expressed in five type of tissues of tomato pericarp (outer and inner epidermis, collenchyma, parenchyma, and vascular tissues), and every five unigenes, was respectively selected from each tissue based on transcription expression. The 3-kb 5′ upstream region of each unigene was identified from the tomato genome sequence (SL2.50) using annotated unigene sequences, and the promoter sequences were further analyzed. The results showed an enrichment in T/A (T/A > 70%) in the promoter regions. A total of 15 putative tissue-/organ-specific promoters were identified and analyzed by real-time (RT) quantitative (q) PCR analysis, of which six demonstrated stronger activity than widely used tissue-specific tomato promoters. These results demonstrate how high spatiotemporal and high throughput gene expression data can provide a powerful means of identifying spatially targeted promoters in plants.

Shock Structures Using the OBurnett Equations in Combination with the Holian Conjecture

In the present work, we study the normal shock wave flow problem using a combination of the OBurnett equations and the Holian conjecture. The numerical results of the OBurnett equations for normal shocks established several fundamental aspects of the equations such as the thermodynamic consistency of the equations, and the existence of the heteroclinic trajectory and smooth shock structures at all Mach numbers. The shock profiles for the hydrodynamic field variables were found to be in quantitative agreement with the direct simulation Monte Carlo (DSMC) results in the upstream region, whereas further improvement was desirable in the downstream region of the shock. For the discrepancy in the downstream region, we conjecture that the viscosity–temperature relation (μ∝Tφ) needs to be modified in order to achieve increased dissipation and thereby achieve better agreement with the benchmark results in the downstream region. In this respect, we examine the Holian conjecture (HC), wherein transport coefficients (absolute viscosity and thermal conductivity) are evaluated using the temperature in the direction of shock propagation rather than the average temperature. The results of the modified theory (OBurnett + HC) are compared against the benchmark results and we find that the modified theory improves upon the OBurnett results, especially in the case of the heat flux shock profile. We find that the accuracy gain is marginal at lower Mach numbers, while the shock profiles are described better using the modified theory for the case of strong shocks.

Reconstruction of a global gene regulatory network for Streptomyces coelicolor: Curation, inference, and assessment

Streptomyces coelicolor A3(2) is a model microorganism for the study of Streptomycetes, antibiotic production, and secondary metabolism in general. However, little effort to globally study its transcription has been made even though S. coelicolor has an outstanding variety of regulators among bacteria. We manually curated 29 years of literature and databases to assemble a meta-curated experimentally-validated gene regulatory network (GRN) with 5386 genes and 9707 regulatory interactions (~41% of the total expected interactions). This provides the most extensive and up-to-date reconstruction available for the regulatory circuitry of this organism. We found a low level of direct experimental validation for the regulatory interactions reported in the literature and curated in this work. Only ~6% (533/9687) are supported by experiments confirming the binding of the transcription factor to the upstream region of the target gene, the so-called "strong" evidence. To tackle network incompleteness, we performed network inference using several methods (including two proposed here) for motif detection in DNA sequences and GRN inference from transcriptomics. Further, we contrasted the structural properties and functional architecture of the networks to assess the predictions' reliability, finding the inference from DNA sequence data to be the most trustworthy. Finally, we show two possible applications of the inferred and the curated network. The inferred one allowed us to identify putative novel transcription factors for the key Streptomyces antibiotic regulatory proteins (SARPs). The curated one allows us to study the conservation of the system-level components between S. coelicolor and Corynebacterium glutamicum. There we identified the basal machinery as the common signature between the two organisms. The curated networks were deposited in Abasy Atlas (https://abasy.ccg.unam.mx/) while the inferences are available as Supplementary Material.

Hybrid de novo genome-reassembly reveals new insights on pathways and pathogenicity determinants in rice blast pathogen Magnaporthe oryzae RMg_Dl

Blast disease incited by Magnaporthe oryzae is a major threat to sustain rice production in all rice growing nations. The pathogen is widely distributed in all rice paddies and displays rapid aerial transmissions, and seed-borne latent infection. In order to understand the genetic variability, host specificity, and molecular basis of the pathogenicity-associated traits, the whole genome of rice infecting Magnaporthe oryzae (Strain RMg_Dl) was sequenced using the Illumina and PacBio (RSII compatible) platforms. The high-throughput hybrid assembly of short and long reads resulted in a total of 375 scaffolds with a genome size of 42.43 Mb. Furthermore, comparative genome analysis revealed 99% average nucleotide identity (ANI) with other oryzae genomes and 83% against M. grisea, and 73% against M. poe genomes. The gene calling identified 10,553 genes with 10,539 protein-coding sequences. Among the detected transposable elements, the LTR/Gypsy and Type LINE showed high occurrence. The InterProScan of predicted protein sequences revealed that 97% protein family (PFAM), 98% superfamily, and 95% CDD were shared among RMg_Dl and reference 70-15 genome, respectively. Additionally, 550 CAZymes with high GH family content/distribution and cell wall degrading enzymes (CWDE) such endoglucanase, beta-glucosidase, and pectate lyase were also deciphered in RMg_Dl. The prevalence of virulence factors determination revealed that 51 different VFs were found in the genome. The biochemical pathway such as starch and sucrose metabolism, mTOR signaling, cAMP signaling, MAPK signaling pathways related genes were identified in the genome. The 49,065 SNPs, 3267 insertions and 3611 deletions were detected, and majority of these varinats were located on downstream and upstream region. Taken together, the generated information will be useful to develop a specific marker for diagnosis, pathogen surveillance and tracking, molecular taxonomy, and species delineation which ultimately leads to device improved management strategies for blast disease.

MeX pipeline for analysis of mobile genetic elements in cancer genome

Background: Mobile genetic elements (MGEs) comprise a major portion of the human genome and are essential for genetic diversity. These elements are known to have the capability to induce mutations in the human genome. To date, there are several MGE insertions which have been reported to be associated with cancer. We aim to use genome next-generation sequencing data and appropriate bioinformatics tools to accurately identify the insertion sites of MGEs in the human genome.Results: Herein, we introduce the MeX pipeline for the localization and annotation of MGEs in paired-end sequencing data. It requires the reference genome sequence, MGE sequences and paired-end sequencing reads. We evaluated MeX on high depth (>75×) Illumina HiSeq data produced at the Broad Institute (NA12878) against human genome 38-built (including only chromosome 1, 2 and 3) and Alu elements. We could identify 78 reference and 1 non-reference Alu insertions in the NA12878 sample. Upon annotation, it was found that the non-reference Alu element was in the 3' UTR region of the RNF2 gene. Out of 78 reference insertions, 42 were in the intronic region, 7 in the upstream region, 5 in the downstream region, 1 in the 3’ UTR region and the rest were not associated with any gene. MeX showed high performance for the identification and annotation of MGEs in genome samples.Conclusion: This study showed that MeX is a robust and powerful tool for the identification and annotation of MGE insertions. It may also serve as a valuable tool to study the phenotypic changes resulting from transpositional events in cancer genomics.