Biochemical basis of activation and inhibition of an NLR immune receptor network

The plant immune system employs intracellular nucleotide-binding domain and leucine-rich repeat (NLR) receptors to identify and respond to pathogen-derived virulence proteins, termed effectors. NLRs function individually or in configurations such as pairs or networks. In solanaceous plants, the NLR required for cell death (NRC) network mediates resistance against various pathogens. The molecular mechanisms by which the NRC network activates are not understood. Moreover, some pathogen effectors can suppress NLRs to promote virulence, and the mechanisms of this suppression are largely unknown. In this thesis, I characterized a cell death-inducing truncation of the helper NRC4, leading to the identification of a conserved N-terminal motif in CC-NLRs. Using mutated NRC variants, I established a method for monitoring NRC activation and investigated sensor-helper communication in the NRC network. My findings support an activation and release model in which NRC-dependent sensors mediate oligomerization of helper NRCs without joining the helper oligomer. The NB domain can encode the minimal signal for resistosome formation in many NRC-dependent sensors. I also elucidated the suppression mechanisms of AVRcap1b and SS15, two effectors that inhibit NRC2 and NRC3. AVRcap1b connects activated NRCs to host TOL proteins, suppressing immunity, while SS15 binds inactive NRCs, preventing helper activation and resistosome formation. Understanding the suppression mechanism enabled me to bioengineer NRC variants that evade inhibition. This work presents a model for sensor-helper activation of NLRs, provides insights into pathogen manipulation of NLR signaling, and demonstrates a novel strategy for bioengineering of disease resistance

Darboux coordinates for (first order) tetrad gravity

The Hamiltonian form of the Hilbert action in the first order tetrad formalism is examined. We perform a non-linear field redefinition of the canonical variables isolating the part of the spin connection which is canonically conjugate to the tetrad. The geometrical meaning of the constraints written in these new variables is examined.Comment: 12 pages, Latex. Minor presentation changes and some references added. Version to appear in Classical and Quantum Gravit

Conserved Charges for Even Dimensional Asymptotically AdS Gravity Theories

Mass and other conserved Noether charges are discussed for solutions of gravity theories with locally Anti-de Sitter asymptotics in 2n dimensions. The action is supplemented with a boundary term whose purpose is to guarantee that it reaches an extremum on the classical solutions, provided the spacetime is locally AdS at the boundary. It is also shown that if spacetime is locally AdS at spatial infinity, the conserved charges are finite and properly normalized without requiring subtraction of a reference background. In this approach, Noether charges associated to Lorentz and diffeomorphism invariance vanish identically for constant curvature spacetimes. The case of zero cosmological constant is obtained as a limit of AdS, where $\Lambda$ plays the role of a regulator.Comment: 8 pages, RevTeX, no figures, two columns, references added and minor typos corrected, final version for Phys. Rev.

The helper NLR immune protein NRC3 mediates the hypersensitive cell death caused by the cell-surface receptor Cf-4

Cell surface pattern recognition receptors (PRRs) activate immune responses that can include the hypersensitive cell death. However, the pathways that link PRRs to the cell death response are poorly understood. Here, we show that the cell surface receptor-like protein Cf-4 requires the intracellular nucleotide-binding domain leucine-rich repeat containing receptor (NLR) NRC3 to trigger a confluent cell death response upon detection of the fungal effector Avr4 in leaves of Nicotiana benthamiana. This NRC3 activity requires an intact N-terminal MADA motif, a conserved signature of coiled-coil (CC)-type plant NLRs that is required for resistosome-mediated immune responses. A chimeric protein with the N-terminal α1 helix of Arabidopsis ZAR1 swapped into NRC3 retains the capacity to mediate Cf-4 hypersensitive cell death. Pathogen effectors acting as suppressors of NRC3 can suppress Cf-4-triggered hypersensitive cell-death. Our findings link the NLR resistosome model to the hypersensitive cell death caused by a cell surface PRR

Integral analysis of environmental and economic performance of combined agricultural intensification & bioenergy production in the Orinoquia region

Agricultural intensification is a key strategy to help meet increasing demand for food and bioenergy. It has the potential to reduce direct and indirect land use change (LUC) and associated environmental impacts while contributing to a favorable economic performance of the agriculture sector. We conduct an integral analysis of environmental and economic impacts of LUC from projected agricultural intensification and bioenergy production in the Orinoquia region in 2030. We compare three agricultural intensification scenarios (low, medium, high) and a reference scenario, which assumes a business-as-usual development of agricultural production. The results show that with current inefficient management or with only very little intensification between 26% and 93% of the existing natural vegetation areas will be converted to agricultural land to meet increasing food demand. This results in the loss of biodiversity by 53% and increased water consumption by 111%. In the medium and high scenarios, the intensification allows meeting increased food demand within current agricultural lands and even generating surplus land which can be used to produce bioenergy crops. This results in the reduction of biodiversity loss by 8-13% with medium and high levels of intensification compared to the situation in 2018. Also, a positive economic performance is observed, stemming primarily from intensification of cattle production and additional energy crop production. Despite increasing irrigation efficiency in more intensive production systems, the water demand for perennial crops and cattle production over the dry season increases significantly, thus sustainable management practices that target efficient water use are needed. Agricultural productivity improvements, particularly for cattle production, are crucial for reducing the pressure on natural areas from increasing demand for both food products and bioenergy. This implies targeted investments in the agricultural sector and integrated planning of land use. Our results showed that production intensification in the Orinoquia region is a mechanism that could reduce the pressure on natural land and its associated environmental and economic impacts

Two NLR immune receptors acquired high-affinity binding to a fungal effector through convergent evolution of their integrated domain

A subset of plant NLR immune receptors carry unconventional integrated domains in addition to their canonical domain architecture. One example is rice Pik-1 that comprises an integrated heavy metal-associated (HMA) domain. Here, we reconstructed the evolutionary history of Pik-1 and its NLR partner, Pik-2, and tested hypotheses about adaptive evolution of the HMA domain. Phylogenetic analyses revealed that the HMA domain integrated into Pik-1 before Oryzinae speciation over 15 million years ago and has been under diversifying selection. Ancestral sequence reconstruction coupled with functional studies showed that two Pik-1 allelic variants independently evolved from a weakly binding ancestral state to high-affinity binding of the blast fungus effector AVR-PikD. We conclude that for most of its evolutionary history the Pik-1 HMA domain did not sense AVR-PikD, and that different Pik-1 receptors have recently evolved through distinct biochemical paths to produce similar phenotypic outcomes. These findings highlight the dynamic nature of the evolutionary mechanisms underpinning NLR adaptation to plant pathogens

Plant pathogens convergently evolved to counteract redundant nodes of an NLR immune receptor network

In plants, nucleotide-binding domain and leucine-rich repeat (NLR)-containing proteins can form receptor networks to confer hypersensitive cell death and innate immunity. One class of NLRs, known as NLR required for cell death (NRCs), are central nodes in a complex network that protects against multiple pathogens and comprises up to half of the NLRome of solanaceous plants. Given the prevalence of this NLR network, we hypothesised that pathogens convergently evolved to secrete effectors that target NRC activities. To test this, we screened a library of 165 bacterial, oomycete, nematode, and aphid effectors for their capacity to suppress the cell death response triggered by the NRC-dependent disease resistance proteins Prf and Rpi-blb2. Among 5 of the identified suppressors, 1 cyst nematode protein and 1 oomycete protein suppress the activity of autoimmune mutants of NRC2 and NRC3, but not NRC4, indicating that they specifically counteract a subset of NRC proteins independently of their sensor NLR partners. Whereas the cyst nematode effector SPRYSEC15 binds the nucleotide-binding domain of NRC2 and NRC3, the oomycete effector AVRcap1b suppresses the response of these NRCs via the membrane trafficking-associated protein NbTOL9a (Target of Myb 1-like protein 9a). We conclude that plant pathogens have evolved to counteract central nodes of the NRC immune receptor network through different mechanisms. Coevolution with pathogen effectors may have driven NRC diversification into functionally redundant nodes in a massively expanded NLR network

Energy Estimation of Cosmic Rays with the Engineering Radio Array of the Pierre Auger Observatory

The Auger Engineering Radio Array (AERA) is part of the Pierre Auger Observatory and is used to detect the radio emission of cosmic-ray air showers. These observations are compared to the data of the surface detector stations of the Observatory, which provide well-calibrated information on the cosmic-ray energies and arrival directions. The response of the radio stations in the 30 to 80 MHz regime has been thoroughly calibrated to enable the reconstruction of the incoming electric field. For the latter, the energy deposit per area is determined from the radio pulses at each observer position and is interpolated using a two-dimensional function that takes into account signal asymmetries due to interference between the geomagnetic and charge-excess emission components. The spatial integral over the signal distribution gives a direct measurement of the energy transferred from the primary cosmic ray into radio emission in the AERA frequency range. We measure 15.8 MeV of radiation energy for a 1 EeV air shower arriving perpendicularly to the geomagnetic field. This radiation energy -- corrected for geometrical effects -- is used as a cosmic-ray energy estimator. Performing an absolute energy calibration against the surface-detector information, we observe that this radio-energy estimator scales quadratically with the cosmic-ray energy as expected for coherent emission. We find an energy resolution of the radio reconstruction of 22% for the data set and 17% for a high-quality subset containing only events with at least five radio stations with signal.Comment: Replaced with published version. Added journal reference and DO

Measurement of the Radiation Energy in the Radio Signal of Extensive Air Showers as a Universal Estimator of Cosmic-Ray Energy

We measure the energy emitted by extensive air showers in the form of radio emission in the frequency range from 30 to 80 MHz. Exploiting the accurate energy scale of the Pierre Auger Observatory, we obtain a radiation energy of 15.8 \pm 0.7 (stat) \pm 6.7 (sys) MeV for cosmic rays with an energy of 1 EeV arriving perpendicularly to a geomagnetic field of 0.24 G, scaling quadratically with the cosmic-ray energy. A comparison with predictions from state-of-the-art first-principle calculations shows agreement with our measurement. The radiation energy provides direct access to the calorimetric energy in the electromagnetic cascade of extensive air showers. Comparison with our result thus allows the direct calibration of any cosmic-ray radio detector against the well-established energy scale of the Pierre Auger Observatory.Comment: Replaced with published version. Added journal reference and DOI. Supplemental material in the ancillary file