Peptide ligand-mediated trade-off between plant growth and stress response.

Deciding whether to grow or to divert energy to stress responses is a major physiological trade-off for plants surviving in fluctuating environments. We show that three leucine-rich repeat receptor kinases (LRR-RKs) act as direct ligand-perceiving receptors for PLANT PEPTIDE CONTAINING SULFATED TYROSINE (PSY)-family peptides and mediate switching between two opposing pathways. By contrast to known LRR-RKs, which activate signaling upon ligand binding, PSY receptors (PSYRs) activate the expression of various genes encoding stress response transcription factors upon depletion of the ligands. Loss of PSYRs results in defects in plant tolerance to both biotic and abiotic stresses. This ligand-deprivation-dependent activation system potentially enables plants to exert tuned regulation of stress responses in the tissues proximal to metabolically dysfunctional damaged sites where ligand production is impaired.

LRR-protein RNH1 dampens the inflammasome activation and is associated with COVID-19 severity.

Inflammasomes are cytosolic innate immune sensors of pathogen infection and cellular damage that induce caspase-1-mediated inflammation upon activation. Although inflammation is protective, uncontrolled excessive inflammation can cause inflammatory diseases and can be detrimental, such as in coronavirus disease (COVID-19). However, the underlying mechanisms that control inflammasome activation are incompletely understood. Here we report that the leucine-rich repeat (LRR) protein ribonuclease inhibitor (RNH1), which shares homology with LRRs of NLRP (nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain containing) proteins, attenuates inflammasome activation. Deletion of RNH1 in macrophages increases interleukin (IL)-1ß production and caspase-1 activation in response to inflammasome stimulation. Mechanistically, RNH1 decreases pro-IL-1ß expression and induces proteasome-mediated caspase-1 degradation. Corroborating this, mouse models of monosodium urate (MSU)-induced peritonitis and lipopolysaccharide (LPS)-induced endotoxemia, which are dependent on caspase-1, respectively, show increased neutrophil infiltration and lethality in Rnh1 -/- mice compared with wild-type mice. Furthermore, RNH1 protein levels were negatively related with disease severity and inflammation in hospitalized COVID-19 patients. We propose that RNH1 is a new inflammasome regulator with relevance to COVID-19 severity.

Leucine-rich repeat receptor-like protein kinase AtORPK1 promotes oxidative stress resistance in an AtORPK1-AtKAPP mediated module in Arabidopsis.

Signal perception and transduction by the cell surface receptors are essential for cell-cell communication and plant response to abiotic stress. In this work, a previously uncharacterized leucine-rich repeat receptor-like kinase (LRR-RLK), Oxidative-stress Related Protein Kinase 1 (AtORPK1), was isolated from Arabidopsis thaliana, and its biological function was investigated in protoplasts, BY-2 cells and transgenic Arabidopsis plants. AtORPK1 is ubiquitously expressed in various tissues and organs of Arabidopsis at different developmental stages. Loss-of-function of AtORPK1 reduced, whereas overexpression of AtORPK1 increased, the oxidative stress resistance and oxidative stress responsive gene expression in orpk1 mutant and AtORPK1 transgenic Arabidopsis. Sub-cellular localization analyses revealed that AtORPK1 is localized to plasma membrane and endosomes, and the specific localization was significantly affected by hydrogen peroxide (H2O2) treatment. Further GFP, CFP, YFP and RFP fusion protein co-localization and FRET analyses demonstrated that AtORPK1 interacted and co-localized with AtKAPP, a common downstream phosphatase, in the enlarged endosomes such as prevacuolar compartments. Our results indicate that AtORPK1 functions as a positive molecular link between the oxidative stress signaling and antioxidant stress in plants.

Closer vein spacing by ectopic expression of nucleotide-binding and leucine-rich repeat proteins in rice leaves.

KEY MESSAGE: Elevated expression of nucleotide-binding and leucine-rich repeat proteins led to closer vein spacing and higher vein density in rice leaves. To feed the growing global population and mitigate the negative effects of climate change, there is a need to improve the photosynthetic capacity and efficiency of major crops such as rice to enhance grain yield potential. Alterations in internal leaf morphology and cellular architecture are needed to underpin some of these improvements. One of the targets is to generate a "Kranz-like" anatomy in leaves that includes decreased interveinal spacing close to that in C4 plant species. As C4 photosynthesis has evolved from C3 photosynthesis independently in multiple lineages, the genes required to facilitate C4 may already be present in the rice genome. The Taiwan Rice Insertional Mutants (TRIM) population offers the advantage of gain-of-function phenotype trapping, which accelerates the identification of rice gene function. In the present study, we screened the TRIM population to determine the extent to which genetic plasticity can alter vein density (VD) in rice. Close vein spacing mutant 1 (CVS1), identified from a VD screening of approximately 17,000 TRIM lines, conferred heritable high leaf VD. Increased vein number in CVS1 was confirmed to be associated with activated expression of two nucleotide-binding and leucine-rich repeat (NB-LRR) proteins. Overexpression of the two NB-LRR genes individually in rice recapitulates the high VD phenotype, due mainly to reduced interveinal mesophyll cell (M cell) number, length, bulliform cell size and thus interveinal distance. Our studies demonstrate that the trait of high VD in rice can be achieved by elevated expression of NB-LRR proteins limited to no yield penalty.

Crosstalk between E-Cadherin/ß-Catenin and NF-κB Signaling Pathways: The Regulation of Host-Pathogen Interaction during Leptospirosis.

Approximately 1 million cases of leptospirosis, an emerging infectious zoonotic disease, are reported each year. Pathogenic Leptospira species express leucine-rich repeat (LRR) proteins that are rarely expressed in non-pathogenic Leptospira species. The LRR domain-containing protein family is vital for the virulence of pathogenic Leptospira species. In this study, the biological mechanisms of an essential LRR domain protein from pathogenic Leptospira were examined. The effects of Leptospira and recombinant LRR20 (rLRR20) on the expression levels of factors involved in signal transduction were examined using microarray, quantitative real-time polymerase chain reaction, and western blotting. The secreted biomarkers were measured using an enzyme-linked immunosorbent assay. rLRR20 colocalized with E-cadherin on the cell surface and activated the downstream transcription factor ß-catenin, which subsequently promoted the expression of MMP7, a kidney injury biomarker. Additionally, MMP7 inhibitors were used to demonstrate that the secreted MMP7 degrades surface E-cadherin. This feedback inhibition mechanism downregulated surface E-cadherin expression and inhibited the colonization of Leptospira. The degradation of surface E-cadherin activated the NF-κB signal transduction pathway. Leptospirosis-associated acute kidney injury is associated with the secretion of NGAL, a downstream upregulated biomarker of the NF-κB signal transduction pathway. A working model was proposed to illustrate the crosstalk between E-cadherin/ß-catenin and NF-κB signal transduction pathways during Leptospira infection. Thus, rLRR20 of Leptospira induces kidney injury in host cells and inhibits the adhesion and invasion of Leptospira through the upregulation of MMP7 and NGAL.

Evolution of variable lymphocyte receptor B antibody loci in jawless vertebrates.

Three types of variable lymphocyte receptor (VLR) genes, VLRA, VLRB, and VLRC, encode antigen recognition receptors in the extant jawless vertebrates, lampreys and hagfish. The somatically diversified repertoires of these VLRs are generated by serial stepwise copying of leucine-rich repeat (LRR) sequences into an incomplete germline VLR gene. Lymphocytes that express VLRA or VLRC are T cell-like, while VLRB-expressing cells are B cell-like. Here, we analyze the composition of the VLRB locus in different jawless vertebrates to elucidate its configuration and evolutionary modification. The incomplete germline VLRB genes of two hagfish species contain short noncoding intervening sequences, whereas germline VLRB genes in six representative lamprey species have much longer intervening sequences that exhibit notable genomic variation. Genomic clusters of potential LRR cassette donors, fragments of which are copied to complete VLRB gene assembly, are identified in Japanese lamprey and sea lamprey. In the sea lamprey, 428 LRR cassettes are located in five clusters spread over a total of 1.7 Mbp of chromosomal DNA. Preferential usage of the different donor cassettes for VLRB assemblage is characterized in our analysis, which reveals evolutionary modifications of the lamprey VLRB genes, elucidates the organization of the complex VLRB locus, and provides a comprehensive catalog of donor VLRB cassettes in sea lamprey and Japanese lamprey.

Suppression of breast cancer progression by FBXL16 via oxygen-independent regulation of HIF1α stability.

Triple-negative breast cancers (TNBCs) are characterized by high rates of recurrence and poor clinical outcomes. Deregulated E3 ligases are involved in breast cancer pathogenesis and progression, but the underlying mechanisms are unclear. Here, we find that F-box and leucine-rich repeat protein 16 (FBXL16) acts as a tumor suppressor in TNBCs. FBXL16 directly binds to HIF1α and induces its ubiquitination and degradation, regardless of the tumor microenvironment, resulting in blockade of the HIF1α-mediated epithelial-mesenchymal transition (EMT) and angiogenesis features of breast cancer. In TNBCs, FBXL16 expression is downregulated by the p38/miR-135b-3p axis, and loss of FBXL16 expression restores HIF1α-mediated metastatic features of breast cancer. Low expression of FBXL16 is associated with high-grade and lymph node-positive tumors and poor overall survival of breast cancer. Taken together, these findings demonstrate that modulation of FBXL16 expression may offer a favorable strategy for treatment of patients with metastatic breast cancer, including TNBCs.

NB-LRR-encoding genes conferring susceptibility to organophosphate pesticides in sorghum.

Organophosphate is the commonly used pesticide to control pest outbreak, such as those by aphids in many crops. Despite its wide use, however, necrotic lesion and/or cell death following the application of organophosphate pesticides has been reported to occur in several species. To understand this phenomenon, called organophosphate pesticide sensitivity (OPS) in sorghum, we conducted QTL analysis in a recombinant inbred line derived from the Japanese cultivar NOG, which exhibits OPS. Mapping OPS in this population identified a prominent QTL on chromosome 5, which corresponded to Organophosphate-Sensitive Reaction (OSR) reported previously in other mapping populations. The OSR locus included a cluster of three genes potentially encoding nucleotide-binding leucine-rich repeat (NB-LRR, NLR) proteins, among which NLR-C was considered to be responsible for OPS in a dominant fashion. NLR-C was functional in NOG, whereas the other resistant parent, BTx623, had a null mutation caused by the deletion of promoter sequences. Our finding of OSR as a dominant trait is important not only in understanding the diversified role of NB-LRR proteins in cereals but also in securing sorghum breeding free from OPS.

CCR -NB-LRR proteins MdRNL2 and MdRNL6 interact physically to confer broad-spectrum fungal resistance in apple (Malus × domestica).

Apple leaf spot, a disease caused by Alternaria alternata f. sp. mali and other fungal species, leads to severe defoliation and results in tremendous losses to the apple (Malus × domestica) industry in China. We previously identified three RPW8, nucleotide-binding, and leucine-rich repeat domain CCR -NB-LRR proteins (RNLs), named MdRNL1, MdRNL2, and MdRNL3, that contribute to Alternaria leaf spot (ALT1) resistance in apple. However, the role of NB-LRR proteins in resistance to fungal diseases in apple remains poorly understood. We therefore used MdRNL1/2/3 as baits to screen ALT1-inoculated leaves for interacting proteins and identified only MdRNL6 (another RNL) as an interactor of MdRNL2. Protein interaction assays demonstrated that MdRNL2 and MdRNL6 interact through their NB-ARC domains. Transient expression assays in apple indicated that complexes containing both MdRNL2 and MdRNL6 are necessary for resistance to Alternaria leaf spot. Intriguingly, the same complexes were also required to confer resistance to Glomerella leaf spot and Marssonina leaf spot in transient expression assays. Furthermore, stable transgenic apple plants with suppressed expression of MdRNL6 showed hypersensitivity to Alternaria leaf spot, Glomerella leaf spot, and Marssonina leaf spot; these effects were similar to the effects of suppressing MdRNL2 expression in transgenic apple plantlets. The identification of these novel broad-spectrum fungal resistance genes will facilitate breeding for fungal disease resistance in apple.