ScanNeo2: a comprehensive workflow for neoantigen detection and immunogenicity prediction from diverse genomic and transcriptomic alterations.

MOTIVATION: Neoantigens, tumor-specific protein fragments, are invaluable in cancer immunotherapy due to their ability to serve as targets for the immune system. Computational prediction of these neoantigens from sequencing data often requires multiple algorithms and sophisticated workflows, which are currently restricted to specific types of variants, such as single-nucleotide variants or insertions/deletions. Nevertheless, other sources of neoantigens are often overlooked. RESULTS: We introduce ScanNeo2 an improved and fully automated bioinformatics pipeline designed for high-throughput neoantigen prediction from raw sequencing data. Unlike its predecessor, ScanNeo2 integrates multiple sources of somatic variants, including canonical- and exitron-splicing, gene fusion events, and various somatic variants. Our benchmark results demonstrate that ScanNeo2 accurately identifies neoantigens, providing a comprehensive and more efficient solution for neoantigen prediction. AVAILABILITY AND IMPLEMENTATION: ScanNeo2 is freely available at https://github.com/ylab-hi/ScanNeo2/ and is accompanied by instruction and application data.

Eight-Electron Copper Nanoclusters for Photothermal Conversion.

Owing to distinct physicochemical properties in comparison to gold and silver counterparts, atomically precise copper nanoclusters are attracting embryonic interest in material science. The introduction of copper cluster nanomaterials in more interesting fields is currently urgent and desired. Reported in this work are novel copper nanoclusters of [XCu54Cl12(tBuS)20(NO3)12] (X=S or none, tBuSH=2-methyl-2-propanethiol), which exhibit high performance in photothermal conversion. The clusters have been prepared in one pot and characterized by combinatorial techniques including ultraviolet-visible spectroscopy (UV-vis), electrospray ionization mass spectrometry (ESI-MS), and X-ray photoelectron spectroscopy (XPS). The molecular structure of the clusters, as revealed by single crystal X-ray diffraction analysis (SCXRD), shows the concentric three-shell Russian doll arrangement of X@Cu14@Cl12@Cu40. Interestingly, the [SCu54Cl12(tBuS)20(NO3)12] cluster contains 8 free valence electrons in its structure, making it the first eight-electron copper nanocluster stabilized by thiolates. More impressively, the clusters possess an effective photothermal conversion (temperature increases by 71 °C within ~50 s, λex=445 nm, 0.5 W cm-2) in a wide wavelength range (either blue or near-infrared). The photothermal conversion can be even driven under irradiation of simulated sunlight (3 sun), endowing the clusters with great potency in solar energy utilization.

Decorating an Anticuboctahedral Copper Kernel with Labile Surface Coatings for Controlling Optical and Catalytic Properties.

Manipulating the interfacial/surface structure of ligand-stabilized atomically precise metal nanoclusters (NCs) is one of the central tasks in nanoscience because surface motifs are directly related to key properties of nanomaterials. Although great progress has been made in engineering the surface of gold and silver nanoclusters, parallel studies on lighter copper analogues hitherto remain unexplored. In this work, we report the design, synthesis, and structure of a new class of copper nanoclusters featuring virtually identical kernels but different surface motifs. The four Cu29 nanoclusters share the same Cu13 kernel with unprecedented anticuboctahedral architecture. Finely modulating synthetic parameters endows the Cu13 core with diverse surface structures, thus affording the Cu29 series with labile surface coatings. More interestingly, the slight surface modification results in distinct optical and catalytic properties of the cluster compounds, highlighting the importance of the surface structure in shaping the behaviors of copper nanomolecules. This work not only exemplifies the efficiency of surface engineering for controlling properties of well-defined copper nanoclusters but also provides a new family of Cu materials with a clear molecular structure and controlled surface motifs that hold great promise in studying structure-property relationships.

A myxozoan genome reveals mosaic evolution in a parasitic cnidarian.

BACKGROUND: Parasite evolution has been conceptualized as a process of genetic loss and simplification. Contrary to this model, there is evidence of expansion and conservation of gene families related to essential functions of parasitism in some parasite genomes, reminiscent of widespread mosaic evolution-where subregions of a genome have different rates of evolutionary change. We found evidence of mosaic genome evolution in the cnidarian Myxobolus honghuensis, a myxozoan parasite of fish, with extremely simple morphology. RESULTS: We compared M. honghuensis with other myxozoans and free-living cnidarians, and determined that it has a relatively larger myxozoan genome (206 Mb), which is less reduced and less compact due to gene retention, large introns, transposon insertion, but not polyploidy. Relative to other metazoans, the M. honghuensis genome is depleted of neural genes and has only the simplest animal immune components. Conversely, it has relatively more genes involved in stress resistance, tissue invasion, energy metabolism, and cellular processes compared to other myxozoans and free-living cnidarians. We postulate that the expansion of these gene families is the result of evolutionary adaptations to endoparasitism. M. honghuensis retains genes found in free-living Cnidaria, including a reduced nervous system, myogenic components, ANTP class Homeobox genes, and components of the Wnt and Hedgehog pathways. CONCLUSIONS: Our analyses suggest that the M. honghuensis genome evolved as a mosaic of conservative, divergent, depleted, and enhanced genes and pathways. These findings illustrate that myxozoans are not as genetically simple as previously regarded, and the evolution of some myxozoans is driven by both genomic streamlining and expansion.

Identification of Myxobolus distalisensis n. sp. (Cnidaria: Myxozoa) infecting yellow catfish Tachysurus fulvidraco (Richardson), with a supplement description of M. voremkhai (Akhmerov, 1960) Landsberg and Lom, 1991.

With growing scale of intensive fish cultivation, the risk of parasite infection in commercial fish is increased. Precisely identifying and characterizing the parasites that infect the farmed fish is critical to understanding the dynamics of their communities. Here, two species of Myxobolus were identified in farmed yellow catfish Tachysurus fulvidraco (Richardson) in China. Myxobolus distalisensis n. sp. developed plasmodia in the gill filaments, with oval to elliptical myxospores measuring 11.3 ± 0.6 (10.4-12.6) × 8.1 ± 0.3 (7.5-8.6) × 5.5 ± 0.2 (5.2-5.8) µm. Two pyriform polar capsules of equal size were measured 5.3 ± 0.4 (4.5-6.3) × 2.7 ± 0.1 (2.3-3) µm. Myxobolus voremkhai (Akhmerov, 1960) Landsberg and Lom, 1991 developed plasmodia in the gill arch and had a myxospore morphology similar to the conspecific isolates described in previous studies. The consensus sequences of M. distalisensis was remarkably distinct from those deposited in the GenBank, with exception of whereas M. voremkhai showing 99.84% identity. The genetic data on both isolates differed considerably from each other, revealing only 86.96% molecular identity. Histologically, M. distalisensis resided in the filament cartilage, and the aggressive proliferation of the sporogenic stages led to lytic cartilage corrosion. In contrast, plasmodia of M. voremkhai grossly observed at the base of the gill filament were embedded by the connective tissue in the gills arch. Phylogenetically, both isolates were separately placed in different subclades, indicating difference in their evolutionary history. Besides, the taxon under the family Myxobolidae was demonstrated non-monophyletic origins, and parasite radiation largely followed their host affinity.

Paclitaxel-based supramolecular hydrogel loaded with mifepristone for the inhibition of breast cancer metastasis.

Breast cancer is the leading cause of cancer death among women and almost all of the breast cancer-caused mortality is related to metastasis. It has been reported that glucocorticoid facilitates the metastasis of breast cancer in mice, and mifepristone can antagonize the effect of glucocorticoid. Paclitaxel is one of the important drugs in the treatment of breast cancer. Mifepristone combined with paclitaxel could be an effective strategy for inhibiting breast cancer metastasis. However, their inherent defects, in terms of short blood circulation half-life and lack of tumor targeting, not only limit their effectiveness but also cause adverse reactions. Therefore, our aim is to explore a novel protocol against breast cancer metastasis, further optimize its therapeutic efficacy by a nanodelivery system, and explore its mechanism. Herein, a paclitaxel-conjugated and mifepristone-loaded hydrogel (PM-nano) was prepared by self-assembly. Its characterizations were studied. The antimetastatic effect was evaluated in vitro and in vivo and its mechanism was also explored by western blot assay. The resultant PM-nano was developed with favorable water solubility and good biocompatibility. Moreover, PM-nano displayed increased cell uptake properties and stimulated drug release in the tumor micro-acidic environment. The PM-nano was more effective in inhibiting the proliferation and metastasis of breast cancer than other groups in vitro and in vivo. The PM-nano might inhibit metastasis through glucocorticoid receptor/receptor tyrosine kinase-like orphan receptor 1 and MMPs. Taken together, PM-nano showed superior antimetastatic effects against breast cancer and excellent biocompatibility in vitro and in vivo, providing a new option for limiting metastasis.

Structure of Self-assembled Peptide Determines the Activity of Aggregation-Induced Emission Luminogen-Peptide Conjugate for Detecting Alkaline Phosphatase.

The unique property of turning on their fluorescence after aggregation or assembly makes aggregation-induced emission luminogens (AIEgens) ideal luminescent molecules for the construction of self-assembled peptide-based nanoprobes. However, the characteristic highly twisted or propeller-shaped molecular conformation of AIEgens tends to prevent the assembly of AIEgen-peptides. Here, we show that (i) the distance between tetraphenylethene (TPE) and assembled peptides should not be too far (less than five glycines), otherwise the self-assembly of peptides cannot limit the intramolecular rotation of conjugated TPE and the luminous efficiency of TPE-peptide to alkaline phosphatase (ALP) will decrease; (ii) properly increasing the number of amino acids with self-assembly ability (three phenylalanines) can improve their ALP-responsive self-assembly and luminescence ability; (iii) the strategy of co-assembly with a non-AIEgen-capped self-assembled peptide is a simple and effective way to realize the efficient assembly and luminescence of AIEgen-peptides; and (iv) the hydrophilic and hydrophobic balance of the probe should always be considered in the construction of an efficient AIEgen-peptide probe. In addition, AIEgen-peptide probes show good selectivity and sensitivity for ALP detection both in vitro and in live bacteria. These insights illustrated here are crucial for guiding the design of AIEgen-conjugated supramolecular materials, especially for the construction of AIEgen-peptides, for enzymes detection, biomarker imaging, diseases therapy, and other biomedical fields.

Transcriptomic Insights into the Diversity and Evolution of Myxozoa (Cnidaria, Endocnidozoa) Toxin-like Proteins.

Myxozoa is a speciose group of endoparasitic cnidarians that can cause severe ecological and economic effects. Their cnidarian affinity is affirmed by genetic relatedness and the presence of nematocysts, historically called "polar capsules". Previous studies have revealed the presence of toxin-like proteins in myxozoans; however, the diversity and evolution of venom in Myxozoa are not fully understood. Here, we performed a comparative analysis using the newly sequenced transcriptomes of five Myxobolidae species as well as some public datasets. Toxin mining revealed that myxozoans have lost most of their toxin families, while most species retained Kunitz, M12B, and CRISP, which may play a role in endoparasitism. The venom composition of Endocnidozoa (Myxozoa + Polypodium) differs from that of free-living cnidarians and may be influenced by ecological and environmental factors. Phylogenetic analyses showed that toxin families of myxozoans and free-living cnidarians were clustered into different clades. Selection analyses showed that purifying selection was the dominant evolutionary pressure in toxins, while they were still influenced by episodic adaptive selection. This suggests that the potency or specificity of a particular toxin or species might increase. Overall, our findings provide a more comprehensive framework for understanding the diversity and evolution of Myxozoa venoms.

Fins infestation induced by Myxobolus xiantaoensis in yellow catfish Tachysurus fulvidraco Richardson, 1846: Some pathophysiological and molecular insights.

The myxozoan parasite Myxobolus xiantaoensis is a fin pathogen of commercially important yellow catfish Tachysurus fulvidraco Richardson, 1846, in the freshwater ponds of China. In the present work, four geographical isolates of M. xiantaoensis were sampled from the fins of yellow catfish. It was found that the spores of four isolates exhibited few markable differences in morphometrics. The small subunit ribosomal DNA (SSU rDNA) sequences of four isolates were conspecific to the SSU rDNA sequence of M. xiantaoensis. No genetic level variation was observed, even in the characteristically more variable internal transcribed spacer (ITS) region. This absence of variability suggests high gene flow as a result of panmixia in the parasitic populations. ITS phylogeny placed four isolates of M. xiantaoensis in a clade together with myxozoans species infecting Siluriformes. The M. xiantaoensis infection inflicted severe hemorrhages on epidermis of ray-fins, which grew into inflammatory epithelial hyperplasia and lytic cartilage signs. The histochemical analysis of infected fins biopsies is characterized by damage of collagen components of cartilage, resulting in weakness, breaks, and missing fin rays. These tissue sections also had a remarkable inflammatory response around the fin cartilage, with the absence of mature spores and chondrocytes. These results indicate that the fin cartilage damage appeared before the development of tissue inflammation and the parasitic infestation of the fins. The present four geographical isolates of M. xiantaoensis were identified by a holistic approach of species characterization based on biological, morphological, and molecular evidence. These four isolates showed some morphological and genetic variations but within the intraspecific range.

A fast and effective method for dissecting parasitic spores: myxozoans as an example.

Disassembling the parasitic spores and acquiring the main subunits is a prerequisite for deep understanding of the basic biology of parasites. Herein we present a fast and efficient method to dissect the myxospores in a few steps, which mainly involved sonication, sucrose density gradient and Percoll density gradient. We tested our method on three myxozoans species and demonstrated this method allows the dismembering of myxospores, isolation of intact and clean nematocysts and shell valves within 2h by low-cost. This new tool will facilitate subsequent analyses and enable a better understanding of the ecological and evolutionary significance of parasitic spores.

Molecular and light microscopy evidence for the transfer of Myxobolus honghuensis from Carassius auratus gibelio broodfish to progeny.

Myxozoans usually have a complex life cycle involving indirect transmission between vertebrate and invertebrate hosts. The vertical transmission of these parasites in vertebrate hosts has not been documented so far. Here, we assessed whether the myxozoan parasite Myxobolus honghuensis is vertically transmitted in naturally infected allogynogenetic gibel carp Carassius auratus gibelio (Bloch). M. honghuensis infection of broodfish, fertilized eggs and laboratory-cultured progeny was monitored in 2018 and 2019. The presporogonic stage was microscopically observed in the pharynx of broodfish and their progeny. In situ hybridization confirmed the presence of M. honghuensis presporogonic stage in the pharynx of broodfish and progeny. Nested PCR results showed that M. honghuensis was present in tissues and eggs of broodfish, fertilized eggs and their corresponding progeny. The sequences obtained from broodfish and progeny showed 98.0-99.8% similarity with ITS-5.8S rDNA of M. honghuensis. This study provides molecular and light microscopy evidence for the transfer of M. honghuensis from broodfish to progeny via the eggs, but it is insufficient to assert that M. honghuensis can transmit vertically in naturally infected allogynogenetic gibel carp. This is the first record about vertical transfer of myxozoan in the vertebrate host.

Description of Myxobolus xiantaoensis n. sp. from the fins of yellow catfish in China: a species previously attributed to Myxobolus physophilus Reuss, 1906 in Chinese records.

Myxozoans are economically important cnidarian endoparasites. Members of this group have been traditionally characterized by a morphology-based taxonomic system. Because myxozoans possess few morphological characters, these data are routinely accompanied by biological traits (host/organ/tissue specificity) and molecular data when describing or identifying myxozoan species. In the present study, a species of Myxobolus was collected from the fins of yellow catfish Tachysurus fulvidraco Richardson, 1846, which was consistent in spore morphology and host/organ specificity with Chinese records of Myxobolus physophilus Reuss, 1906. However, these earlier records and our own findings are inconsistent with the original description of M. physophilus from Russia. Specifically, there are differences in spore morphology (shape, intercapsular appendix, and polar capsule size), the infection site (air bladder vs. fins), and the host affinity (common rudd vs. yellow catfish). The inconsistencies allow us to conclude that both the present Myxobolus species and Chinese records of M. physophilus are distinct from the original description of M. physophilus and represent a new Myxobolus species, which we named Myxobolus xiantaoensis n. sp.

Description of a New Freshwater Ciliate Epistylis wuhanensis n. sp. (Ciliophora, Peritrichia) from China, with a Focus on Phylogenetic Relationships within Family Epistylididae.

Two populations of Epistylis wuhanensis n. sp., a new freshwater peritrich ciliate, were isolated from different freshwater ponds located in Hubei, China. Their morphological characteristics were investigated using live observation, protargol impregnation, and scanning electron microscopy (SEM). Specimens from the two populations showed identical arrangement of the infraciliature and identical small subunit ribosomal RNA (SSU rRNA) gene and ITS1-5.8S-ITS2 sequences. The zooids present bell-shaped and 90-175 × 27-54 µm in vivo. Macronucleus is variable in shape and located in the middle of cell. Pellicle is usually smooth with 139-154 and 97-105 striations above and below the trochal band, respectively. SSU rRNA gene and ITS1-5.8S-ITS2 sequences of E. wuhanensis n. sp. did not match any available sequences in GenBank. Phylogenetically, E. wuhanensis n. sp. clusters with the other Epistylis within the family Epistylididae, but is distinct from the major clades of Epistylis. Above all, the morphological characteristics and molecular analyses support that the present Epistylis is a new species. Expanded phylogenetic analyses of sessilids based on both SSU rRNA gene sequences and ITS1-5.8S-ITS2 sequences reveal that the genus Epistylis consists of Epistylis morphospecies and taxonomic revision of the genus is needed.

Discovery of a highly potent, selective and novel CDK9 inhibitor as an anticancer drug candidate.

A series of novel hybrid structure derivatives, containing both LEE011 and Cabozantinib pharmacophore, were designed, synthesized and evaluated. Surprisingly, a compound 4d was discovered that highly exhibited effective and selective activity of CDK9 inhibition with IC50=12nM. It effectively induced apoptosis in breast and lung cancer cell lines at nanomolar level. Molecular docking of 4d to ATP binding site of CDK9 kinase demonstrated a new hydrogen bonding between F atom of 4-(3-fluorobenzyloxy) group and ASN116 residue, compared with the positive control, LEE011. The compound 4d could block the cell cycle both in G0/G1 and G2/M phase to prevent the proliferation and differentiation of cancer cells. Mice bared-breast cancer treated with compound 4d showed significant suppression of cancer with low toxicity. Taken together, this novel compound 4d could be a promising drug candidate for clinical application.

Histopathological and ultrastructural studies of Myxobolus turpisrotundus from allogynogenetic gibel carp Carassius auratus gibelio in China.

During an ongoing systematic survey on species diversity of myxozoans parasitising allogynogenetic gibel carp Carassius auratus gibelio (Bloch) in China, plasmodia were detected in the fins, lip, jaw, gill chamber, gill arches, operculum and oral cavity of infected fish. Combining the morphological and molecular data, the present species was identified as Myxobolus turpisrotundus Zhang, Wang, Li et Gong, 2010. Histopathological examination revealed that despite infecting different organs, M. turpisrotundus always occurred in dermis, demonstrating its affinity to this tissue. Histopathological effect of M. turpisrotundus on the host is relatively mild except parasites in the gill arches producing compression of the adipose tissue and heavy adductor muscles deformation with lymphohistiocytic infiltrates. In addition, the plasmodia in different sites were with the same complex structure arrangement: cup-like cells with unknown derivation, a thin collagenous fibril layer, areolar connective tissue, basement membrane and host epithelial cell. Ultrastructural analysis showed that the parasite has monosporic pansporoblast and sporogenesis followed the usual pattern of most of the myxosporeans.

Intraspecific morphometric variation in myxosporeans.

Morphometric data from spores of ten myxosporean species were statistically analysed to explore myxosporean intraspecific variation in measurements when obtained from a sample from: (1) the same plasmodium, (2) different plasmodia from the same host and (3) different host individuals and localities. In some cases, significant differences in spore dimensions were found between samples from the same plasmodium, highlighting the difficulty of obtaining representative measurements of myxosporean spore. In addition, significant differences in spore dimensions were found when plasmodia from the same site of infection were compared, suggesting that measurements of spores should come from several different plasmodia of the sampling to increase the reliability of the morphology data. Moreover, significant differences in spore dimensions were observed for most spore dimensions when data were compared between localities. In all cases, there was clear overlap in ranges of dimensions even when means differed significantly. The present statistical analysis shows that intraspecific morphometric variation of myxosporean species commonly occurs, highlighting the importance of reporting ranges of measurements for a species, not just the mean dimensions, and taking into account all evidence when assigning or describing myxosporean species.

Two-photon fluorescent probes for biological Mg(2+) detection based on 7-substituted coumarin.

Two novel water-soluble coumarin-based compounds (OC7, NC7) were designed and synthesized as two-photon fluorescent probes for biological Mg(2+) detection. The compounds feature a ß-keto acid as a high selective binding site for Mg(2+) and the coumarin framework as the two-photon fluorophore. OC7 and NC7 show significant "off-on" detecting signals (9.05-fold and 23.8-fold fluorescence enhancement) and lower detection limits compared with previous reported two-photon fluorescent probes for Mg(2+). Moreover, OC7-Mg(2+) and NC7-Mg(2+) exhibit large two-photon absorption cross sections (340 and 615 GM) at the near-infrared wavelengths (740 and 860 nm), which indicates that the probes are very suitable for detection of Mg(2+) in vivo. Both OC7 and NC7 are pH-insensitive and of low cytotoxicity and can be applied to image intracellular Mg(2+) under two-photon microscopy (TPM). Our results provide a strategy to modify the coumarin fluorophore to get better two-photon fluorescent properties. And the results also suggest that electronic density of ß-keto acid plays a very important role in the recognition of Mg(2+).

Accelerated Fmoc solid-phase synthesis of peptides with aggregation-disrupting backbones.

In this work, we describe an accelerated solid-phase synthetic protocol for ordinary or difficult peptides involving air-bath heating and amide protection. For the Hmsb-based backbone amide protection, an optimized acyl shift condition using 1,4-dioxane was discovered. The efficiency and robustness of the protocol was validated in the course of preparation of classical difficult peptides and ubiquitin protein segments.

Diaminodiacid-based solid-phase synthesis of all-hydrocarbon stapled α-helical peptides.

An alternative stapling strategy is described herein using Fmoc-solid phase peptide synthesis (SPPS) that employed pre-prepared diaminodiacid building blocks to introduce all-hydrocarbon staples into peptides by on-resin cyclization. Compared to unstapled native peptides, diaminodiacid-based stapled peptides exhibited an increased α-helicity ratio and stability toward protease. Moreover, the linkage length was found to affect the bioactivity of the peptides and their ability to inhibit the Wnt pathway. Therefore, the new stapling method provides an alternative way to obtain stapled peptides with tunable linkers of diaminodiacids.

First record of Chilodonella piscicola (Ciliophora: Chilodonellidae) from two endangered fishes, Schizothorax o'connori and Oxygymnocypris stewartii in Tibet.

Schizothorax o'connori and Oxygymnocypris stewartii are two endangered endemic Tibetan fishes that thrive in the Lhasa River at an average altitude over 4000 m. During artificial reproduction of endemic Tibetan fishes, the juvenile fish of S. o'connori and O. stewartii experienced mass mortality event. The causative agent is diagnosed to be a ciliate parasite, Chilodonella piscicola (syn. C. cyprini), which is common in various fishes. Here, we supplemented its description based on the morphological and molecular data. The body of C. piscicola is oval, 30-60 × 25-40 µm in vivo. Cyrtos is hook-like, composed of 9-10 toothed nematodesmal rods. Somatic kineties usually contain seven right kineties and nine left kineties. Two parallel circumoral kineties revolve round the cyrtos, and one preoral kinety extends to the anterior end of the fourth left-most kinety. Terminal fragment kinety is linear and on the top left of dorsal side. Sequence alignments revealed that the present SSU rDNA and ITS1-5.8S-ITS2 sequences are both most similar to the sequences of C. uncinata with the similarities of 98.2 and 99.5%. The phylogenetic analyses showed that C. piscicola is sister to other Chilodonella species, whereas C. cyprini (FJ873805) cluster with Tetrahymena species. Molecular analysis shows that the ITS1-5.8S-ITS2 sequence of C. cyprini in GenBank is unreliable. Our study extended the host range of C. piscicola and supplemented and revised the molecular data. Besides, as far as we know, this is the first record of C. piscicola in Tibetan plateau.