On the dependent recognition of some long zinc finger proteins.

The human genome contains about 800 C2H2 zinc finger proteins (ZFPs), and most of them are composed of long arrays of zinc fingers. Standard ZFP recognition model asserts longer finger arrays should recognize longer DNA-binding sites. However, recent experimental efforts to identify in vivo ZFP binding sites contradict this assumption, with many exhibiting short motifs. Here we use ZFY, CTCF, ZIM3, and ZNF343 as examples to address three closely related questions: What are the reasons that impede current motif discovery methods? What are the functions of those seemingly unused fingers and how can we improve the motif discovery algorithms based on long ZFPs' biophysical properties? Using ZFY, we employed a variety of methods and find evidence for 'dependent recognition' where downstream fingers can recognize some previously undiscovered motifs only in the presence of an intact core site. For CTCF, high-throughput measurements revealed its upstream specificity profile depends on the strength of its core. Moreover, the binding strength of the upstream site modulates CTCF's sensitivity to different epigenetic modifications within the core, providing new insight into how the previously identified intellectual disability-causing and cancer-related mutant R567W disrupts upstream recognition and deregulates the epigenetic control by CTCF. Our results establish that, because of irregular motif structures, variable spacing and dependent recognition between sub-motifs, the specificities of long ZFPs are significantly underestimated, so we developed an algorithm, ModeMap, to infer the motifs and recognition models of ZIM3 and ZNF343, which facilitates high-confidence identification of specific binding sites, including repeats-derived elements. With revised concept, technique, and algorithm, we can discover the overlooked specificities and functions of those 'extra' fingers, and therefore decipher their broader roles in human biology and diseases.

Anthropogenic disturbance driving population decline of a dominant tree in East Asia evergreen broadleaved forests over the last 11,000 years.

Current biodiversity loss is generally considered to have been caused by anthropogenic disturbance, but it is unclear when anthropogenic activities began to affect biodiversity loss. One hypothesis suggests it began with the Industrial Revolution, whereas others propose that anthropogenic disturbance has been associated with biodiversity decline since the early Holocene. To test these hypotheses, we examined the unique vegetation of evergreen broadleaved forests (EBLFs) in East Asia, where humans have affected landscapes since the early Holocene. We adopted a genomic approach to infer the demographic history of a dominant tree (Litsea elongata) of EBLFs. We used Holocene temperature and anthropogenic disturbance factors to calculate the correlation between these variables and the historical effective population size of L. elongata with Spearman statistics and integrated the maximum-entropy niche model to determine the impact of climate change and anthropogenic disturbance on fluctuation in its effective population size. We identified 9 well-defined geographic clades for the populations of L. elongata. Based on the estimated historical population sizes of these clades, all the populations contracted, indicating persistent population decline over the last 11,000 years. Demographic history of L. elongata and human population change, change in cropland use, and change in irrigated rice area were significantly negatively correlated, whereas climate change in the Holocene was not correlated with demographic history. Our results support the early human impact hypothesis and provide comprehensive evidence that early anthropogenic disturbance may contribute to the current biodiversity crisis in East Asia.

Energetic and Kinetic Competition on the Stability of Pd13 Clusters: Ab Initio Molecular Dynamics Simulations.

Material stability is the focus on both experiments and calculations, which includes the energetic stability at the static state and the thermodynamic stability at the kinetic state. To show whether energetics or kinetics dominates on material stability, this study focuses on the Pd13 clusters, because of their observable magnetic moment in experiment. Energetically, the CALYPSO searching method and first-principles calculations find that Pd13(C2) is the ground state at 0 K while the static frequency calculations demonstrate that the icosahedron Pd13(Ih) becomes more favorable on free energy as temperature increases. However, their magnetic moments (8 µB) are not in agreement with the experimental value (<5.2 µB). Kinetically, ab initio molecular dynamics simulations reveal that Pd13(C3v) (6 µB) has supreme isomerization temperature and the other 11 low-lying isomers transform to Pd13(C3v) directly or indirectly, demonstrating that Pd13(C3v) has the maximum probability to be observed in experiment. The magnetic moment difference between experiment (<5.2 µB) and this calculation (6 µB) may be due to the spin multiplicities. Our result suggests that the magnetic moment disparity between theory and experiment (in Pd13 clusters) originates from the kinetic stability.

Phylogenomic insights into the origin and evolutionary history of evergreen broadleaved forests in East Asia under Cenozoic climate change.

The evergreen versus deciduous leaf habit is an important functional trait for adaptation of forest trees and has been hypothesized to be related to the evolutionary processes of the component species under paleoclimatic change, and potentially reflected in the dynamic history of evergreen broadleaved forests (EBLFs) in East Asia. However, knowledge about the shift of evergreen versus deciduous leaf with the impact of paleoclimatic change using genomic data remains rare. Here, we focus on the Litsea complex (Lauraceae), a key lineage with dominant species of EBLFs, to gain insights into how evergreen versus deciduous trait shifted, providing insights into the origin and historical dynamics of EBLFs in East Asia under Cenozoic climate change. We reconstructed a robust phylogeny of the Litsea complex using genome-wide single-nucleotide variants (SNVs) with eight clades resolved. Fossil-calibrated analyses, diversification rate shifts, ancestral habit, ecological niche modelling and climate niche reconstruction were employed to estimate its origin and diversification pattern. Taking into account studies on other plant lineages dominating EBLFs of East Asia, it was revealed that the prototype of EBLFs in East Asia probably emerged in the Early Eocene (55-50 million years ago [Ma]), facilitated by the greenhouse warming. As a response to the cooling and drying climate in the Middle to Late Eocene (48-38 Ma), deciduous habits were evolved in the dominant lineages of the EBLFs in East Asia. Up to the Early Miocene (23 Ma), the prevailing of East Asian monsoon increased the extreme seasonal precipitation and accelerated the emergence of evergreen habits of the dominant lineages, and ultimately shaped the vegetation resembling that of today.

Effect of genetically determined BCAA levels on cardiovascular diseases and their risk factors: A Mendelian randomization study.

BACKGROUND AND AIMS: Observational studies have demonstrated that serum branched-chain amino acids (BCAAs) are associated with the risk of various cardiovascular diseases (CVDs) and their risk factors. However, the causal effect is unclear. The aim of this study was to investigate the effect of genetically determined BCAA levels on CVDs and their risk factors using Mendelian randomization (MR). METHODS AND RESULTS: We performed univariable and multivariable MR analyses using summary-level data from multiple GWASs and the FinnGen consortium to investigate the association between BCAA levels and the risk of CVDs (myocardial infarction, ischemic stroke, and intracerebral hemorrhage) and their risk factors (atrial fibrillation, hypertension, heart failure, and valvular heart disease). We used the random-effects IVW approach as the primary statistical method and incorporated MR estimates from different data sources using the fixed-effects model. We found genetically determined total and individual BCAA levels and a high risk of hypertension. However, there is no evidence of a causal relationship between BCAA levels and 3 cardiovascular diseases and other their risk factors. The odds of hypertension increased per 1-SD increase in BCAA levels (OR = 1.02 95% CI: 1.01, 1.04; P = 0.005), valine (OR = 1.02 95% CI: 1.01, 1.03; P<0.0001), leucine (OR = 1.02 95% CI: 1.01, 1.04; P＜0.01), and isoleucine (OR = 1.02 95% CI: 1.01, 1.03; P < 0.0001). This result was also significant in the multivariable MR. CONCLUSIONS: This MR study suggests that total and individual BCAA levels could be associated with a high risk of hypertension.

Plastome phylogenomic analysis reveals evolutionary divergences of Polypodiales suborder Dennstaedtiineae.

BACKGROUND: Polypodiales suborder Dennstaedtiineae contain a single family Dennstaedtiaceae, eleven genera, and about 270 species, and include some groups that were previously placed in Dennstaedtiaceae, Hypolepidaceae, Monachosoraceae, and Pteridaceae. The classification and phylogenetic relationships among these eleven genera have been poorly understood. To explore the deep relationships within suborder Dennstaedtiineae and estimate the early diversification of this morphologically heterogeneous group, we analyzed complete plastomes of 57 samples representing all eleven genera of suborder Dennstaedtiineae using maximum likelihood and Bayesian inference. RESULTS: The phylogenetic relationships of all the lineages in the bracken fern family Dennstaedtiaceae were well resolved with strong support values. All six genera of Hypolepidoideae were recovered as forming a monophyletic group with full support, and Pteridium was fully supported as sister to all the other genera in Hypolepidoideae. Dennstaedtioideae (Dennstaedtia s.l.) fell into four clades with full support: the Microlepia clade, the northern Dennstaedtia clade, the Dennstaedtia globulifera clade, and the Dennstaedtia s.s. clade. Monachosorum was strongly resolved as sister to all the remaining genera of suborder Dennstaedtiineae. Based on the well resolved relationships among genera, the divergence between Monachosorum and other groups of suborder Dennstaedtiineae was estimated to have occurred in the Early Cretaceous, and all extant genera (and clades) in Dennstaedtiineae, were inferred to have diversified since the Late Oligocene. CONCLUSION: This study supports reinstating a previously published family Monachosoraceae as a segregate from Dennstaedtiaceae, based on unique morphological evidence, the shady habitat, and the deep evolutionary divergence from its closest relatives.

Target-Based Virtual Screening and LC/MS-Guided Isolation Procedure for Identifying Phloroglucinol-Terpenoid Inhibitors of SARS-CoV-2.

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has led to more than 5 million deaths worldwide to date. Due to the limited therapeutic options so far available, target-based virtual screening with LC/MS support was applied to identify the novel and high-content compounds 1-4 with inhibitory effects on SARS-CoV-2 in Vero E6 cells from the plant Dryopteris wallichiana. These compounds were also evaluated against SARS-CoV-2 in Calu-3 cells and showed unambiguous inhibitory activity. The inhibition assay of targets showed that compounds 3 and 4 mainly inhibited SARS-CoV-2 3CLpro, with effective Kd values. Through docking and molecular dynamics modeling, the binding site is described, providing a comprehensive understanding of 3CLpro and interactions for 3, including hydrogen bonds, hydrophobic bonds, and the spatial occupation of the B ring. Compounds 3 and 4 represent new, potential lead compounds for the development of anti-SARS-CoV-2 drugs. This study has led to the development of a target-based virtual screening method for exploring the potency of natural products and for identifying natural bioactive compounds for possible COVID-19 treatment.

Assessment of Morphological Features and Imaging Characteristics of Patients with Intracranial Artery Dissection: A High-Resolution MRI Study.

BACKGROUND: Intracranial artery dissection (IAD) is a pathological dissection of the arterial wall. .However, the morphological features and imaging characteristics of patients with intracranial artery dissection (IAD) remain poorly understood. METHODS: The study reports on 70 IAD patients (30 culprit and 40 non-culprit). All participants underwent high-resolution magnetic resonance imaging (HR-MRI) scans. The morphological features and imaging characteristics of artery dissection were carefully investigated. Demographics and clinical characteristics of culprit and non-culprit patients were also collected. Apparent differences between the two groups, which could be used as biomarkers for ischemic event caused by the culprit dissection, were identified by receiver operating characteristic (ROC) curve analysis. RESULTS: The IAD patients studied could be classified into five different types on the basis of morphological features: classical dissection (n = 31), fusiform aneurysm (n = 2), long dissected aneurysm (n = 9), dolichoectatic dissecting aneurysm (n = 6), and saccular aneurysm (n = 22). The direct sites of artery dissection (double lumen and intimal flap) can be seen in most IAD patients on HR-MRI. Additionally, the presence of hypertension, double lumen and intimal flap were associated with culprit lesions and might be considered biomarkers for the ischemic event caused by the culprit dissection. CONCLUSIONS: Analysis showed that HR-MRI allowed easy visualization of abnormal morphology of artery dissection lesions. This was of great significance for the diagnosis of IAD and gave a better understanding of its pathophysiological mechanism.

An Authentication and Secure Communication Scheme for In-Vehicle Networks Based on SOME/IP.

The rapid development of intelligent networked vehicles (ICVs) has brought many positive effects. Unfortunately, connecting to the outside exposes ICVs to security threats. Using secure protocols is an important approach to protect ICVs from hacker attacks and has become a hot research area for vehicle security. However, most of the previous studies were carried out on V2X networks, while those on in-vehicle networks (IVNs) did not involve Ethernet. To this end, oriented to the new IVNs based on Ethernet, we designed an efficient secure scheme, including an authentication scheme using the Scalable Service-Oriented Middleware over IP (SOME/IP) protocol and a secure communication scheme modifying the payload field of the original SOME/IP data frame. The security analysis shows that the designed authentication scheme can provide mutual identity authentication for communicating parties and ensure the confidentiality of the issued temporary session key; the designed authentication and secure communication scheme can resist the common malicious attacks conjointly. The performance experiments based on embedded devices show that the additional overhead introduced by the secure scheme is very limited. The secure scheme proposed in this article can promote the popularization of the SOME/IP protocol in IVNs and contribute to the secure communication of IVNs.

Design of a CANFD to SOME/IP Gateway Considering Security for In-Vehicle Networks.

In recent years, Ethernet has been introduced into vehicular networks to cope with the increasing demand for bandwidth and complexity in communication networks. To exchange data between controller area network (CAN) and Ethernet, a gateway system is required to provide a communication interface. Additionally, the existence of networked devices exposes automobiles to cyber security threats. Against this background, a gateway for CAN/CAN with flexible data-rate (CANFD) to scalable service-oriented middleware over IP (SOME/IP) protocol conversion is designed, and security schemes are implemented in the routing process to provide integrity and confidentiality protections. Based on NXP-S32G, the designed gateway is implemented and evaluated. Under most operating conditions, the CPU and the RAM usage are less than 5% and 20 MB, respectively. Devices running a Linux operating system can easily bear such a system resource overhead. The latency caused by the security scheme accounts for about 25% of the entire protocol conversion latency. Considering the security protection provided by the security scheme, this overhead is worthwhile. The results show that the designed gateway can ensure a CAN/CANFD to SOME/IP protocol conversion with a low system resource overhead and a low latency while effectively resisting hacker attacks such as frame forgery, tampering, and sniffing.

Design, synthesis, antitumor activity and theoretical calculation of novel PI3Ka inhibitors.

PI3Kα has been identified as an ideal target to treat with PIK3CA gene mutation disease, including drugs such as Alpelisib and Copanlisib. Five purine analogues and four thiazole analogues were designed and synthesized. Their enzymaticactivity against PI3Ka/ß/γ/δ were tested, respectively. All compounds showed excellent selectivity in modulating PI3Ka activity, and parts of the compounds showed good inhibition. Meanwhile, we used Autodock 4.2 to explore the binding mode of the most potential compound Tg with the target protein. In addition, DFT was used to calculate the HOMO-LUMO maps of the compounds Tf, Tg and positive control. This paper will provide some useful information for further drug design of PI3Kα inhibitors.

The Antithrombotic Effects of Low Molecular Weight Fragment from Enzymatically Modified of Laminaria Japonica Polysaccharide.

BACKGROUND Laminaria japonica polysaccharide (LJP), a fucose enriched sulfated polysaccharide has been demonstrated to have excellent anticoagulant and antithrombotic activities. However, the antithrombotic effect of low molecular weight polysaccharide from enzymatically modified of LJP (LMWEP) remains unknown. MATERIAL AND METHODS LMWEP was prepared by fucoidanase enzymatic hydrolysis, and the antithrombotic and anticoagulant activities, and the underlying mechanism were investigated thoroughly. Rats were randomly divided into 6 groups (8 rats in each group): the blank control group, the blank control group treated with LMWEP (20 mg/kg), the model group, the model group treated with heparin (2 mg/kg), the model group treated with LJP (20 mg/kg), and the model group treated with LMWEP (20 mg/kg). After 7 days of intravenous administration, blood was collected for biochemical parameters examinations. RESULTS LMWEP increased the activated partial thromboplastin time (APTT), thrombin time (TT), prothrombin time (PT), 6-keto prostaglandin F1alpha (6-Keto-PGF1alpha), and endothelial nitric oxide synthase (eNOS). In addition, LMWEP decreased fibrinogen (FIB), endothelin-1 (ET-1), thromboxane B2 (TXB2), erythrocyte sedimentation rate (ESR), and hematocrit (HCT). CONCLUSIONS LMWEP, an enzymatically modified fragment with a molecular weight of 25.8 kDa, is a potential antithrombotic candidate for treatment of thrombosis related diseases.

Quantitative specificity of STAT1 and several variants.

The quantitative specificity of the STAT1 transcription factor was determined by measuring the relative affinity to hundreds of variants of the consensus binding site including variations in the length of the site. The known consensus sequence is observed to have the highest affinity, with all variants decreasing binding affinity considerably. There is very little loss of binding affinity when the CpG within the consensus binding site is methylated. Additionally, the specificity of mutant proteins, with variants of amino acids that interact with the DNA, was determined and nearly all of them are observed to lose specificity across the entire binding site. The change of Asn at position 460 to His, which corresponds to the natural amino acid at the homologous position in STAT6, does not change the specificity nor does it change the length preference to match that of STAT6. These results provide the first quantitative analysis of changes in binding affinity for the STAT1 protein, and several variants of it, to hundreds of different binding sites including different spacer lengths, and the effect of CpG methylation.

Quantitative profiling of selective Sox/POU pairing on hundreds of sequences in parallel by Coop-seq.

Cooperative binding of transcription factors is known to be important in the regulation of gene expression programs conferring cellular identities. However, current methods to measure cooperativity parameters have been laborious and therefore limited to studying only a few sequence variants at a time. We developed Coop-seq (cooperativity by sequencing) that is capable of efficiently and accurately determining the cooperativity parameters for hundreds of different DNA sequences in a single experiment. We apply Coop-seq to 12 dimer pairs from the Sox and POU families of transcription factors using 324 unique sequences with changed half-site orientation, altered spacing and discrete randomization within the binding elements. The study reveals specific dimerization profiles of different Sox factors with Oct4. By contrast, Oct4 and the three neural class III POU factors Brn2, Brn4 and Oct6 assemble with Sox2 in a surprisingly indistinguishable manner. Two novel half-site configurations can support functional Sox/Oct dimerization in addition to known composite motifs. Moreover, Coop-seq uncovers a nucleotide switch within the POU half-site when spacing is altered, which is mirrored in genomic loci bound by Sox2/Oct4 complexes.

Quantitative profiling of BATF family proteins/JUNB/IRF hetero-trimers using Spec-seq.

BACKGROUND: BATF family transcription factors (BATF, BATF2 and BATF3) form hetero-trimers with JUNB and either IRF4 or IRF8 to regulate cell fate in T cells and dendritic cells in vivo. While each combination of the hetero-trimer has a distinct role, some degree of cross-compensation was observed. The basis for the differential actions of IRF4 and IRF8 with BATF factors and JUNB is still unknown. We propose that the differences in function between these hetero-trimers may be caused by differences in their DNA binding preferences. While all three BATF family transcription factors have similar binding preferences when binding as a hetero-dimer with JUNB, the cooperative binding of IRF4 or IRF8 to the hetero-dimer/DNA complex could change the preferences. We used Spec-seq, which allows for the efficient and accurate determination of relative affinity to a large collection of sequences in parallel, to find differences between cooperative DNA binding of IRF4, IRF8 and BATF family members. RESULTS: We found that without IRF binding, all three hetero-dimer pairs exhibit nearly the same binding preferences to both expected wildtype binding sites TRE (TGA(C/G)TCA) and CRE (TGACGTCA). IRF4 and IRF8 show the very similar DNA binding preferences when binding with any of the three hetero-dimers. No major change of binding preferences was found in the half-sites between different hetero-trimers. IRF proteins bind with substantially lower affinity with either a single nucleotide spacer between IRF and BATF binding site or with an alternative mode of binding in the opposite orientation. In addition, the preference to CRE binding site was reduced with either IRF binding in all BATF-JUNB combinations. CONCLUSIONS: The specificities of BATF, BATF2 and BATF3 are all very similar as are their interactions with IRF4 and IRF8. IRF proteins binding adjacent to BATF sites increases affinity substantially compared to sequences with spacings between the sites, indicating cooperative binding through protein-protein interactions. The preference for the type of BATF binding site, TRE or CRE, is also altered when IRF proteins bind. These in vitro preferences aid in the understanding of in vivo binding activities.

Indoor Positioning Based on Bluetooth Low-Energy Beacons Adopting Graph Optimization.

Bluetooth Low-Energy (BLE) beacons-based indoor positioning is a promising method for indoor positioning, especially in applications of position-based services (PbS). It has low deployment cost and it is suitable for a wide range of mobile devices. Existing BLE beacon-based positioning methods can be categorized as range-based methods and fingerprinting-based methods. For range-based methods, the positions of the beacons should be known before positioning. For fingerprinting-based methods, a pre-requisite is the reference fingerprinting map (RFM). Many existing methods focus on how to perform the positioning assuming the beacon positions or RFM are known. However, in practical applications, determining the beacon positions or RFM in the indoor environment is normally a difficult task. This paper proposed an efficient and graph optimization-based way for estimating the beacon positions and the RFM, which combines the range-based method and the fingerprinting-based method. The method exists without need for any dedicated surveying instruments. A user equipped with a BLE-enabled mobile device walks in the region collecting inertial readings and BLE received signal strength indication (RSSI) readings. The inertial measurements are processed through the pedestrian dead reckoning (PDR) method to generate the constraints at adjacent poses. In addition, the BLE fingerprints are adopted to generate constraints between poses (with similar fingerprints) and the RSSIs are adopted to generate distance constraints between the poses and the beacon positions (according to a pre-defined path-loss model). The constraints are then adopted to form a cost function with a least square structure. By minimizing the cost function, the optimal user poses at different times and the beacon positions are estimated. In addition, the RFM can be generated through the pose estimations. Experiments are carried out, which validates that the proposed method for estimating the pre-requisites (including beacon positions and the RFM). These estimated pre-requisites are of sufficient quality for both range-based and fingerprinting-based positioning.

Response Element Composition Governs Correlations between Binding Site Affinity and Transcription in Glucocorticoid Receptor Feed-forward Loops.

Combinatorial gene regulation through feed-forward loops (FFLs) can bestow specificity and temporal control to client gene expression; however, characteristics of binding sites that mediate these effects are not established. We previously showed that the glucocorticoid receptor (GR) and KLF15 form coherent FFLs that cooperatively induce targets such as the amino acid-metabolizing enzymes AASS and PRODH and incoherent FFLs exemplified by repression of MT2A by KLF15. Here, we demonstrate that GR and KLF15 physically interact and identify low affinity GR binding sites within glucocorticoid response elements (GREs) for PRODH and AASS that contribute to combinatorial regulation with KLF15. We used deep sequencing and electrophoretic mobility shift assays to derive in vitro GR binding affinities across sequence space. We applied these data to show that AASS GRE activity correlated (r(2) = 0.73) with predicted GR binding affinities across a 50-fold affinity range in transfection assays; however, the slope of the linear relationship more than doubled when KLF15 was expressed. Whereas activity of the MT2A GRE was even more strongly (r(2) = 0.89) correlated with GR binding site affinity, the slope of the linear relationship was sharply reduced by KLF15, consistent with incoherent FFL logic. Thus, GRE architecture and co-regulator expression together determine the functional parameters that relate GR binding site affinity to hormone-induced transcriptional responses. Utilization of specific affinity response functions and GR binding sites by FFLs may contribute to the diversity of gene expression patterns within GR-regulated transcriptomes.

Quality assessment of traditional Chinese medicine herb couple by high-performance liquid chromatography and mass spectrometry combined with chemometrics.

This study was designed to develop a simple, specific and reliable method to overall analyze the chemical constituents in clematidis radix et rhizome/notopterygii rhizome et radix herb couple using high-performance liquid chromatography coupled with tandem mass spectrometry and multiple chemometric analysis. First, the separation and qualitative analysis of herb couple was achieved on an Agilent Zorbax Eclipse Plus C18 column (250 mm × 4.6 mm, 5 µm), and 69 compounds were unambiguously or tentatively identified. Moreover, in quantitative analysis, eight ingredients including six coumarins and two triterpenoid sapogenins were quantified by high-performance liquid chromatography coupled with tandem mass spectrometry. In terms of good linearity (r(2) ≥ 0.9995) with a relatively wide concentration range, recovery (85.40-102.50%) and repeatability (0.99-4.45%), the validation results suggested the proposed method was reliable, and successfully used to analyze ten batches of herb couple samples. Then, hierarchical cluster analysis and principal component analysis were used to classify samples and search significant ingredients. The results showed that ten batches of herb couple samples were classified into three groups, and six compounds were found for its better quality control.

Dryopterisjinpingensis, a critically endangered diploid new species of Dryopteridaceae from Yunnan, China.

Dryopterisjinpingensis, a new species of diploid, sexually reproductive ferns of Dryopteridaceae from Yunnan, southwestern China, is described and illustrated. Morphologically, D.jinpingensis is similar to D.gaoligongensis but unique in elongated lanceolate laminae, sessile or subsessile pinna stalks, and overlapping membranous scales adnate to stipe base. Phylogenetic analyses based on both plastome and the nuclear AK1 gene sequences showed that D.jinpingensis is sister to D.gaoligongensis. A detailed taxonomic description with line drawings is provided, and its conservation status is evaluated to be critically endangered.

Novel structural determinants and bacterial death-related regulatory effects of the scorpion defensin BmKDfsin4 against gram-positive bacteria.

Numerous defensins constitute a family of cationic antimicrobial peptides with high degrees of sequence variability, and in-depth characterization of their structural basis and antibacterial mechanisms remains limited. Here, a representative scorpion defensin, BmKDfsin4, with two distinct hydrophobic and basic residue clusters, was extensively investigated. The hydrophobic residue cluster, formed by Phe2, Pro5, Phe6, Phe28 and Leu29 residues, strongly influences the antibacterial activity of BmKDfsin4 against Gram-positive bacteria. Compared with the three scattered Lys13, Lys30 and Arg32 residues, the basic residue cluster, consisting of the Arg19, Arg20, Arg21 and Arg37 residues, played a less important role. The synergistic interaction between the basic residue cluster and Arg32 significantly affected BmKDfsin4 function. The bacterial growth inhibition by BmKDfsin4 was associated with regulating expression levels of cell division-related genes, cell wall synthesis-related genes and bacterial autolysis-related genes rather than destroying the bacterial cell membrane. The coincubation of BmKDfsin4 with the bacterial strains induced gradual changes in the bacterial surface from a rough and thin surface to a noticeably wrinkled surface together with abundant white spots and even complete cavities within the bacteria. These findings revealed novel structural determinants and bacterial death-related regulatory effects of the defensin BmKDfsin4 and highlighted diverse antibacterial mechanisms of defensins.