Characterization of Nucleotide Binding ­Site-Encoding Genes in Sweetpotato, Ipomoea batatas(L.) Lam., and Their Response to Biotic and Abiotic Stresses

Sweetpotato, <i>Ipomoea batatas</i> (L.) Lam., is an important and widely grown crop, yet its production is affected severely by biotic and abiotic stresses. The nucleotide binding site (NBS)-encoding genes have been shown to improve stress tolerance in several plant species. However, the characterization of NBS-encoding genes in sweetpotato is not well-documented to date. In this study, a comprehensive analysis of NBS-encoding genes has been conducted on this species by using bioinformatics and molecular biology methods. A total of 315 NBS-encoding genes were identified, and 260 of them contained all essential conserved domains while 55 genes were truncated. Based on domain architectures, the 260 NBS-encoding genes were grouped into 6 distinct categories. Phylogenetic analysis grouped these genes into 3 classes: TIR, CC (I), and CC (II). Chromosome location analysis revealed that the distribution of NBS-encoding genes in chromosomes was uneven, with a number ranging from 1 to 34. Multiple stress-related regulatory elements were detected in the promoters, and the NBS-encoding genes’ expression profiles under biotic and abiotic stresses were obtained. According to the bioinformatics analysis, 9 genes were selected for RT-qPCR analysis. The results revealed that <i>IbNBS75</i>, <i>IbNBS219</i>, and <i>IbNBS256</i> respond to stem nematode infection; <i>Ib­NBS240</i>, <i>IbNBS90</i>, and <i>IbNBS80</i> respond to cold stress, while <i>IbNBS208</i>, <i>IbNBS71</i>, and <i>IbNBS159</i> respond to 30% PEG treatment. We hope these results will provide new insights into the evolution of NBS-encoding genes in the sweetpotato genome and contribute to the molecular breeding of sweetpotato in the future.

Structure and comparison of the motor domain of centromere-associated protein E

Centromere-associated protein E (CENP-E) plays an essential role in mitosis and is a target candidate for anticancer drugs. However, it is difficult to design small-molecule inhibitors of CENP-E kinesin motor ATPase activity owing to a lack of structural information on the CENP-E motor domain in complex with its inhibitors. Here, the CENP-E motor domain was crystallized in the presence of an ATP-competitive inhibitor and the crystal structure was determined at 1.9 Å resolution. In the determined structure, ADP was observed instead of the inhibitor in the nucleotide-binding site, even though no ADP was added during protein preparation. Structural comparison with the structures of previously reported CENP-E and those of other kinesins indicates that the determined structure is nearly identical except for several loop regions. However, the retention of ADP in the nucleotide-binding site of the structure strengthens the biochemical view that the release of ADP is a rate-limiting step in the ATPase cycle of CENP-E. These results will contribute to the development of anticancer drugs targeting CENP-E and to understanding the function of kinesin motor domains.

Deletion of Arno Reduces Eosinophilic Inflammation and Interleukin-5 Expression in a Murine Model of Rhinitis

Background: ARF nucleotide-binding site opener (ARNO) is a guanine nucleotide-exchange factor for ADP-ribosylation factor proteins. ARF nucleotide-binding site opener also binds MyD88, and small-molecule inhibition of ARNO reduces inflammation in animal models of inflammatory arthritis and acute inflammation. However, whether genetic deletion of Arno in mice reduces pathologic inflammation has not yet been reported. Furthermore, its role in the nasal cavity has yet to be investigated. Objective: To generate Arno knockout mice and to determine whether genetic loss of ARNO reduces eosinophilic inflammation in the ovalbumin (OVA) murine model of rhinitis. Methods: Arno knockout mice were generated and wild type and knockout littermates were subjected to the OVA-induced mouse model of rhinosinutitis. Eosinophilic inflammation was assessed through immunofluorescent quantification of EMBP+ eosinophils in the septal mucosa and cytokine expression was assessed by quantitative polymerase chain reaction. Results: Arno knockout mice are viable and fertile without any noted deficits. Arno wild type and knockout mice subjected to the OVA-induced model of rhinitis demonstrated an average of 314.5 and 153.8 EMBP+ cells per mm2 septal tissue, respectively ( P < .05). Goblet cells per mm of basal lamina were assessed via Alcian blue and there was no statistically significant difference between Arno wild type and knockout mice. Ovalbumin-induced expression of interleukin-5 (IL-5) was significantly reduced in Arno knockout mice ( P < .05). There was no statistically significant reduction in IL-4, IL-13, or eotaxin-1 expression. Conclusions: These data demonstrate that deletion of Arno reduces eosinophilic inflammation and IL-5 expression in an OVA-induced model of rhinitis.

The N-terminal domain in TRPM2 channel is a conserved nucleotide binding site

This study by Tóth et al. has defined that the N-terminal MHR1/2 domain is a conserved ADPR binding site in TRPM2 from ancient cnidarians to vertebrate, and that it is the key ligand binding site for invertebrate TRPM2 channel activation by ADPR, the same as observed in human and zebrafish TRPM2.

Transcriptome-wide analysis of expressed resistance gene analogs (RGAs) in mango

Mango is an economically important fruit crop largely cultivated in the (sub)tropics and thus, is constantly challenged by a myriad of insect pests and diseases. Here, we identified and characterized the resistance gene analogs (RGAs) of mango from de novo assembly of transcriptomic sequences. A core RGA database of mango with 747 protein models was established and classified based on conserved domains and motifs: 53 nucleotide binding site proteins (NBS); 27 nucleotide binding site-leucine rich repeat proteins (NBS-LRR); 17 coiled-coil NBS-LRR (CNL); 2 toll/interleukin-1 receptor NBS-LRR (TNL); 29 coiled-coil NBS (CN); 4 toll/interleukin-1 receptor NBS (TN); 17 toll/interleukin-1 receptor with unknown domain (TX); 158 receptor-like proteins (RLP); 362 receptor-like kinases (RLK); 72 transmembrane coiled-coil domain protein (TM-CC), and 6 NBS-encoding proteins with other domains. The various molecular functions, biological processes, and cellular localizations of these RGAs were functionally well-annotated through gene ontology (GO) analysis, and their expression profiles across different mango varieties were also determined. Phylogenetic analysis broadly clustered the core RGAs into 6 major clades based on their domain classification, while TM-CC proteins formed subclades all across the tree. The phylogenetic results suggest highly divergent functions of the RGAs which also provide insights into the mango-pest co-evolutionary arms race. From the mango RGA transcripts, 134 unique EST-SSR loci were identified, and primers were designed targeting these potential markers. To date, this is the most comprehensive analysis of mango RGAs which offer a trove of markers for utilization in resistance breeding of mango.