The potato StMKK5-StSIPK module enhances resistance to Phytophthora pathogens through activating the salicylic acid and ethylene signalling pathways

Mitogen-activated protein kinase (MAPK) cascades play pivotal roles in plant responses to both biotic and abiotic stress. A screen of a Nicotiana benthamiana cDNA virus-induced gene silencing (VIGS) library for altered plant responses to inoculation with Phytophthora infestans previously identified an NbMKK gene, encoding a clade D MAPKK that we renamed as NbMKK5, which is involved in immunity to P. infestans. To study the role of the potato orthologous gene, referred to as StMKK5, in the response to P. infestans, we transiently overexpressed StMKK5 in N. benthamiana and observed that cell death occurred at 2 days postinfiltration. Silencing of the highly conserved eukaryotic protein SGT1 delayed the StMKK5-induced cell death, whereas silencing of the MAPK-encoding gene NbSIPK completely abolished the cell death response. Further investigations showed that StMKK5 interacts with, and directly phosphorylates, StSIPK. Furthermore, both StMKK5 and StSIPK trigger salicylic acid (SA)- and ethylene (Eth)-related gene expression, and co-expression of the salicylate hydroxylase NahG with the negative regulator of Eth signalling CTR1 hampers StSIPK-triggered cell death. This observation indicates that the cell death triggered by StMKK5-StSIPK is dependent on the combination of SA- and Eth-signalling. By introducing point mutations, we showed that the kinase activity of both StMKK5 and StSIPK is required for triggering cell death. Genetic analysis showed that StMKK5 depends on StSIPK to trigger plant resistance. Thus, our results define a potato StMKK5-SIPK module that positively regulates immunity to P. infestans via activation of both the SA and Eth signalling pathways.     

Genomic insights into selection for heterozygous alleles and woody traits in Populus tomentosa

Heterozygous alleles are widespread in outcrossing and clonally propagated woody plants. The variation in heterozygosity that underlies population adaptive evolution and phenotypic variation, however, remains largely unknown. Here, we describe a de novo chromosome-level genome assembly of Populus tomentosa, an economic and ecologically important native tree in northern China. By resequencing 302 natural accessions, we determined that the South subpopulation (Pop_S) encompasses the ancestral strains of P. tomentosa, while the Northwest subpopulation (Pop_NW) and Northeast subpopulation (Pop_NE) experienced different selection pressures during population evolution, resulting in significant population differentiation and a decrease in the extent of heterozygosity. Analysis of heterozygous selective sweep regions (HSSR) suggested that selection for lower heterozygosity contributed to the local adaptation of P. tomentosa by dwindling gene expression and genetic load in the Pop_NW and Pop_NE subpopulations. Genome-wide association studies (GWAS) revealed that 88 single nucleotide polymorphisms (SNPs) within 63 genes are associated with nine wood composition traits. Among them, the selection for the homozygous AA allele in PtoARF8 is associated with reductions in cellulose and hemicellulose contents by attenuating PtoARF8 expression, and the increase in lignin content is attributable to the selection for decreases in exon heterozygosity in PtoLOX3 during adaptive evolution of natural populations. This study provides novel insights into allelic variations in heterozygosity associated with adaptive evolution of P. tomentosa in response to the local environment and identifies a series of key genes for wood component traits, thereby facilitating genomic-based breeding of important traits in perennial woody plants.     

Human appropriation of net primary production as driver of change in landscape-scale vertebrate richness

Aim

Land use is the most pervasive driver of biodiversity loss. Predicting its impact on species richness (SR) is often based on indicators of habitat loss. However, the degradation of habitats, especially through land-use intensification, also affects species. Here, we evaluate whether an integrative metric of land-use intensity, the human appropriation of net primary production, is correlated with the decline of SR in used landscapes across the globe.

Location

Global.

Time period

Present.

Major taxa studied

Birds, mammals and amphibians.

Methods

Based on species range maps (spatial resolution: 20 km × 20 km) and an area-of-habitat approach, we calibrated a “species–energy model” by correlating the SR of three groups of vertebrates with net primary production and biogeographical covariables in “wilderness” areas (i.e., those where available energy is assumed to be still at pristine levels). We used this model to project the difference between pristine SR and the SR corresponding to the energy remaining in used landscapes (i.e., SR loss expected owing to human energy extraction outside wilderness areas). We validated the projected species loss by comparison with the realized and impending loss reconstructed from habitat conversion and documented by national Red Lists.

Results

Species–energy models largely explained landscape-scale variation of mapped SR in wilderness areas (adjusted R²-values: 0.79–0.93). Model-based projections of SR loss were lower, on average, than reconstructed and documented ones, but the spatial patterns were correlated significantly, with stronger correlation in mammals (Pearson's r = 0.68) than in amphibians (r = 0.60) and birds (r = 0.57).

Main conclusions

Our results suggest that the human appropriation of net primary production is a useful indicator of heterotrophic species loss in used landscapes, hence we recommend its inclusion in models based on species–area relationships to improve predictions of land-use-driven biodiversity loss.     

Functional cis-element sequence requirements for suppression of gene expression by the TNPA protein of the Zea mays transposon En/Spm

TNPA, one of the two transposition proteins encoded by the En/Spm transposable elements of Zea mays, suppresses the expression of genes that contain an appropriate cis element. Suppression can be monitored in tobacco protoplasts in a transient expression assay as follows. The plant promoter-driven expression of the Escherichia coli-glucuronidase (GUS)-encoding gene, uidA, is repressed in the presence of TNPA if the GUS gene contains a functional cis element in the untranslated RNA leader sequence. Earlier, we found that the minimal cis element is composed of two 12 by sequences in a tail-to-tail inverted orientation. Each 12 by sequence is sufficient to bind TNPA in vitro and can be thought of as a half-site in the cis element. Here, we investigated the sequence requirements of the minimal cis element. Our observations support our expectations that a functional cis element must provide a template to which two TNPA molecules can bind in the correct orientation. Sequences within the half-sites can be altered as long as the eight bases that make up the consensus binding sites are not changed. However, we found the following unexpected sequence specificities. Firstly, some changes to the consensus binding sequence can be tolerated in one half-site, as long as the other site matches the consensus. Secondly, although the region between the half-sites can vary in sequence and in length between two and four bases, a thymidine residue is not tolerated directly 5′ preceding the second half-site. Since many variants of the cis element sequence remain functional, the suppressor response element provides a flexible tool for artificially manipulating the expression of genes.     

ATP稳定肌浆网膜蛋白色氨酸相关构象的机理的探讨

在无ＡＴＰ存在时,带相同正电荷、但饱和不同的两种胺类两亲物,即Ｃ１８饱和的硬脂胺与单不饱和的油胺引起肌浆网蛋白内源荧光强度降低,当ＡＴＰ或一些阴离子化合物先与肌浆网作用,再加入硬脂胺或油胺,则肌浆网蛋白内源荧光下降幅度明显减小,即存在拮抗作用。腺苷对硬脂胺或油胺均无此拮抗作用。肌浆网与油胺先保温,再加入ＡＴＰ或阴离子化合物,ＡＴＰ仍有拮抗,阴离子化合物对油胺则无拮抗作用。然而在肌浆网钙泵蛋白上存在     

The agony of choice: Impact of the host animal species on the enzyme-linked immunosorbent assay performance for host cell protein quantification

Host cell proteins (HCPs) are inevitable process-related impurities in biotherapeutics commonly monitored by enzyme-linked immunosorbent assays (ELISAs). Of particular importance for their reliable detection are the anti-HCP polyclonal antibodies (pAbs), supposed to detect a broad range of HCPs. The present study focuses on the identification of suitable host animal species for the development of high-performance CHO-HCP ELISAs, assuming the generation of pAbs with adequate coverage and specificity. Hence, antibodies derived from immunization of sheep, goats, donkeys, rabbits, and chickens were compared concerning their amount of HCP-specific antibodies, coverage, and performance in a sandwich ELISA. Immunization of sheep, goats, donkeys, and rabbits met all test criteria, whereas the antibodies from chickens cannot be recommended based on the results of this study. Additionally, a mixture of antibodies from the five host species was prepared to assess if coverage and ELISA performance can be improved by a multispecies approach. Comparable results were obtained for the single- and multispecies ELISAs in different in-process samples, indicating no substantial improvement for the latter in ELISA performance while raising ethical and financial concerns.     

Piping plover chick ecology following landscape-level disturbance

Population declines of disturbance-dependent species due to suppression of natural disturbances are realized across ecosystems. The piping plover (Charadrius melodus; plover), a disturbance-dependent and conservation-reliant shorebird that nests on sandy beaches and barrier islands on the Atlantic Coast, was listed under the United States Endangered Species Act in 1986. In 2012, Hurricane Sandy landed on Fire and Westhampton islands, barrier island nesting sites for plovers in New York, USA. Hurricane Sandy was a natural disturbance in this system, creating abundant nesting habitat. The number of chicks produced by a pair, or a population, is a direct measure of reproductive output, and gaining a better understanding of productivity and chick behavior following large-scale habitat creation may improve plover habitat management and potentially species persistence. We evaluated the effects of landscape features on habitat selection, behavior, and survival of plover broods using logistic regression, generalized linear mixed effects models, and survival models. Plover broods selected flatter sites with less dense vegetation than available at random. Chick foraging rates were highest in moist substrates and were lower in areas of higher nesting plover density. Chick survival was greater for broods that hatched earlier in the breeding season and increased as chicks aged. Generally, providing access to sites with flatter, moist substrates will likely result in higher quality brood rearing habitat on the landscape. Ultimately, vegetation removal and habitat management may be needed to reduce plover nesting density and ensure sufficient habitat, which may in turn improve plover chick survival. Moreover, sustaining natural landscape disturbances such as those resulting from storms, and not taking actions to prevent hurricane-created overwash, will allow these landscape features to persist.     

Linkage map of seven polymorphic markers on rat Chromosome 18

A genetic linkage map of seven polymorphic markers was created with F₂ intercross progeny of F344/N and LEW/N rats and assigned to rat Chromosome (Chr) 18. Five of the markers described were defined by simple sequence length polymorphisms (SSLPs) associated with five genes: transthyretin (TTR), trypsin inhibitor-like protein (TILP), 2 adrenergic receptor (ADRB2), olfactory neuron-specific G protein (OLF), and gap junction protein (GJA1). One marker was defined by a restriction fragment length polymorphism (RFLP) detected with a probe for the human colony stimulating factor 1 receptor (CSF1R) gene. The D18N1R locus was defined by an anonymous DNA fragment amplified by the randomly amplified polymorphic DNA (RAPD) technique with a single short primer. These seven DNA loci formed a single genetic linkage group 30.4 cM in length with the following order: TTR-6.8 cM-D18N1R-9.1 cM-TILP-4.3 cM-CSF1R-0 cM-ADRB2-10.2 cM-OLF-0 cM-GJA1. The five SSLP markers were highly polymorphic. In a total of 13 inbred rat strains analyzed (F344/ N, LEW/N, LOU/MN, WBB1/N, WBB2/N, MR/N, MNR/N, ACI/N, SHR/N, WKY/N, BN/SsN, BUF/N, and LER/N), three to six alleles were detected for each marker. Remarkable linkage conservation was detected between the region of rat Chr 18 mapped and a region of mouse Chr 18. However, genes associated with these markers have been mapped to three different human chromosomes (Chrs 5, 6, and 18). The markers described here should be useful for genetic mapping studies and genetic monitoring of inbred rat strains.