XRN2-mediated regulation of let-7 microRNAs is of critical pathophysiological significance in Humans MicroRNAs are critical regulators of diverse developmental and physiological processes, and they themselves get regulated both at the level of biogenesis and turnover. We demonstrate that the ribonuclease XRN2 can degrade the mature forms of certain let-7 family members in multiple human cancer cell lines, without affecting their precursors. XRN2 depletion results in a reduction in the expression of a number of oncogenes, and diminishes the proliferative and metastatic potential of cancer cells. The clinical relevance of these observations is also verified in tumour transcriptomics data from public RNA-sequencing datasets, where XRN2 mRNA expression is inversely correlated with the levels of a large number of miRNAs, including let-7 members, and high XRN2 mRNA levels are associated with poor survival in hepatocellular carcinoma, lung adenocarcinoma, and glioblastoma. We demonstrate that the miRNA is released by an as-yet unidentified proteinaceous  miRNA release factor from the grasp of Argonaute before its degradation. Our analyses of the patient-derived transcriptomics data also show that XRN2, via its regulation of let-7, affects multiple pathways in a consistent manner across epithelial and glial cell lineages, and thus, is of critical pathophysiological significance.
Catchet-MS identifies IKZF1-targeting Thalidomide analogues as novel HIV-1 latency reversal agents A major pharmacological strategy toward HIV cure aims to reverse latency in infected cells as a first step leading to their elimination. While the unbiased identification of molecular targets physically associated with the latent HIV-1 provirus would be highly valuable to unravel the molecular determinants of HIV-1 transcriptional repression and latency reversal, due to technical limitations, this has not been possible. Here we use a dCas9 targeted chromatin and histone enrichment strategy coupled to mass spectrometry (Catchet-MS) to describe the protein composition of the latent and activated HIV-1 5LTR. Catchet-MS identified known and novel latent 5LTR-associated host factors. Among these, IKZF1 is a novel HIV-1 transcriptional repressor, required for Polycomb Repressive Complex 2 recruitment to the LTR. We find the clinically advanced thalidomide analogue iberdomide, and the FDA approved analogues lenalidomide and pomalidomide, to be novel LRAs that, by targeting IKZF1 for degradation, reverse HIV-1 latency in CD4+T-cells isolated from virally suppressed people living with HIV-1.

One Sentence SummarydCas9 targeted chromatin and histone enrichment for mass spectrometry (Catchet-MS) led to the identification of IKZF1-targeting thalidomide analogues as novel HIV-1 latency reversal agents
The important choice of reference environment in microevolutionary climate response predictions It is well documented that individuals of wild populations can adjust to climate change by means of phenotypic plasticity, but few reports on adaptation by means of genetically based microevolution caused by selection. Disentanglement of these separate effects requires that the reference environment (the environmental zero-point) is defined, and this should not be done arbitrarily. The problem is that an error in the reference environment may lead to large errors in predicted microevolution. Together with parameter values and initial mean trait values, the reference environment can be estimated from environmental, phenotypic and fitness data. A prediction error method for this purpose is described, with the feasibility shown by simulations. As shown in a toy example, an estimated reference environment may have large errors, especially for small populations. This may still be a better choice than use of an initial environmental value in a recorded time series, or the mean value, which is often used. Another alternative may be to use the mean value of a past and stationary stochastic environment, which the population is judged to have been fully adapted to, in the sense that the expected geometric mean fitness was at a global maximum. Exceptions are cases with constant phenotypic plasticity, where the microevolutionary changes per generation follow directly from phenotypic and environmental data, independent of the chosen reference environment. Trimeric Photosystem I facilitates energy transfer from phycobilisomes in Synechocystis sp. PCC 6803 In cyanobacteria, phycobilisomes serve as peripheral light-harvesting complexes of the two photosystems, extending their antenna size and the wavelength range of photons available for photosynthesis. The abundance of phycobilisomes, the number of phycobiliproteins they contain, and their light-harvesting function are dynamically adjusted in response to the physiological conditions. Phycobilisomes are also thought to be involved in state transitions that maintain the excitation balance between the two photosystems. Unlike its eukaryotic counterpart, PSI is trimeric in many cyanobacterial species and the physiological significance of this is not well understood. Here we compared the composition and light-harvesting function of phycobilisomes in cells of Synechocystis sp. PCC 6803, which has primarily trimeric PSI, and the{Delta} psaL mutant unable to form trimers. We also investigated a mutant additionally lacking the PsaJ and PsaF subunits of PSI. Both strains with monomeric PSI accumulated significantly more allophycocyanin per chlorophyll, indicating higher abundance of phycobilisomes. On the other hand, a higher phycocyanin:allophycocyanin ratio in WT suggests larger phycobilisomes or the presence of APC-less phycobilisomes (CpcL-type), that are not assembled in cells with monomeric PSI. Steady-state and time-resolved fluorescence spectroscopy at room temperature and 77 K revealed that PSII receives more energy from the phycobilisomes at the expense of PSI in cells with monomeric PSI, regardless of the presence of PsaF. Taken together, these results show that the oligomeric state of PSI has an impact on the excitation energy flow in Synechocystis.

One-sentence summaryCyanobacterial mutants with monomeric PSI show changes in the composition and abundance of phycobilisomes and in the excitation energy transfer to PSII and PSI. Structural basis for targeting the human T-cell leukemia virus Tax oncoprotein and syntenin-1 interaction using a small molecule Human T-cell leukemia virus type-1 (HTLV-1) is the causative agent of adult T-cell leukemia (ATL). Although ATL is a well-characterized T-cell neoplasm, linked to intermittent expression of the viral Tax-1 protein, there is currently no strategy to target Tax-1 functions using small molecules. Here, we report a comprehensive interaction map between Tax-1 and human PDZ domain-containing proteins (hPDZome). We show that Tax-1 interacts with more than one-third of hPDZome components, including proteins involved in cell cycle, cell-cell junctions, cytoskeleton organization, and membrane complex assembly. Using nuclear magnetic resonance (NMR) spectroscopy, we have determined the structural basis of the interaction between the C-terminal PDZ binding motif (PBM) of Tax-1, and the PDZ domains of syntenin-1, an evolutionary conserved hub that controls exosome trafficking. Finally, we have used confocal imaging, molecular modelling, NMR and mammalian cell-based assays to demonstrate that the Tax-1/syntenin-1 interaction is amenable to small-molecule inhibition. Altogether, our study highlights the biological significance of Tax-PDZ interactome and its interplay with exosome formation. It shows a direct link between extracellular vesicles and HTLV-1 transmission, providing a novel framework for the design of targeted therapies for HTLV-1-induced diseases. Robust retrieval of data stored in DNA by de Bruijn graph-based de novo strand assembly DNA data storage is a rapidly developing technology with great potential due to its high density, long-term durability, and low maintenance cost. The major technical challenges include various errors, such as the strand breaks, rearrangements, and indels that frequently arise during DNA synthesis, amplification, sequencing, and preservation. Through a de novo assembly strategy, we developed an algorithm based on the de Bruijn graph and greedy path search (DBGPS) to address these issues. DBGPS shows distinct advantages in handling DNA breaks, rearrangements, and indels. The robustness of DBGPS is demonstrated by accelerated aging, multiple independent data retrievals, deep error-prone PCR, and large data scale simulations. Remarkably, 6.8 MB of data can be retrieved accurately from a seriously corrupted sample that has been treated at 70 {degrees}C for 70 days. With DBGPS, we were able to achieve a logical density of 1.30 bits/cycle and a physical density of 295 PB/g.

One-Sentence SummaryA de Bruijn graph-based de novo assembly algorithm for DNA data storage enables fast and robust data readouts even with DNA samples that have been severely corrupted. Soluble ECM promotes organotypic formation in lung alveolar model Micropatterned suspension culture creates consistently sized and shaped cell aggregates but has not produced organotypic structures from stable cells, thus restricting its use in accurate disease modeling. Here, we show that organotypic structure is achieved in hybrid suspension culture via supplementation of soluble extracellular matrix (ECM). We created a viable lung organoid from epithelial, endothelial, and fibroblast human stable cell lines in suspension culture. We demonstrate the importance of soluble ECM in organotypic patterning with the emergence of lumen-like structures with airspace showing feasible gas exchange units, formation of branching, perfusable vasculature, and long-term 70-day maintenance of lumen structure. Our results show a dependent relationship between enhanced fibronectin fibril assembly and the incorporation of ECM in the organoid. We successfully applied this technology in modeling lung fibrosis via bleomycin induction and test a potential antifibrotic drug in vitro while maintaining fundamental cell-cell interactions in lung tissue. Our human fluorescent lung organoid (hFLO) model represents features of pulmonary fibrosis which were ameliorated by fasudil treatment. We also demonstrate a 3D culture method with potential of creating organoids from mature cells, thus opening avenues for disease modeling and regenerative medicine, enhancing understanding of lung cell biology in health and lung disease. Evolution of thermal physiology alters the projected range of threespine stickleback under climate change Species distribution models (SDMs) are widely used to predict range shifts but could be unreliable under climate change scenarios because they do not account for evolution. The thermal physiology of a species is a key determinant of range and thus incorporating thermal trait evolution into SDMs might be expected to alter projected ranges. We identified a genetic basis for physiological and behavioural traits that evolve in response to temperature change in natural populations of threespine stickleback (Gasterosteus aculeatus). Using these data, we created geographic range projections using a mechanistic niche area approach under two climate change scenarios. Under both scenarios, trait data was either static ( no evolution models), allowed to evolve at observed evolutionary rates ( evolution models), or allowed to evolve at a rate of evolution scaled by the trait variance that is explained by quantitative trait loci (QTL;  scaled evolution models). We show that incorporating these traits and their evolution substantially altered the projected ranges for a widespread panmictic marine population, with over 7-fold increases in area under climate change projections when traits are allowed to evolve. Evolution-informed SDMs should improve the precision of forecasting range dynamics under climate change, and aid in their application to management and the protection of biodiversity. ISL1 is necessary for auditory neuron development and contributes towards tonotopic organization A cardinal feature of the auditory pathway is frequency selectivity, represented in a tonotopic map from the cochlea to the cortex. The molecular determinants of the auditory frequency map are unknown. Here, we discovered that the transcription factor ISL1 regulates the molecular and cellular features of auditory neurons, including the formation of the spiral ganglion and peripheral and central processes that shape the tonotopic representation of the auditory map. We selectively knocked out Isl1 in auditory neurons using Neurod1Cre strategies. In the absence of Isl1, spiral ganglion neurons migrate into the central cochlea and beyond, and the cochlear wiring is profoundly reduced and disrupted. The central axons of Isl1 mutants lose their topographic projections and segregation at the cochlear nucleus. Transcriptome analysis of spiral ganglion neurons shows that Isl1 regulates neurogenesis, axonogenesis, migration, neurotransmission-related machinery, and synaptic communication patterns. We show that peripheral disorganization in the cochlea affects the physiological properties of hearing in the midbrain and auditory behavior. Surprisingly, auditory processing features are preserved despite the significant hearing impairment, revealing central auditory pathway resilience and plasticity in Isl1 mutant mice. Mutant mice have a reduced acoustic startle reflex, altered prepulse inhibition, and characteristics of compensatory neural hyperactivity centrally. Our findings show that ISL1 is one of the obligatory factors required to sculpt auditory structural and functional tonotopic maps. Still, upon Isl1 deletion, the ensuing central compensatory plasticity of the auditory pathway does not suffice to overcome developmentally induced peripheral dysfunction of the cochlea. Comparison of hybrid learning and remote education in the implementation of the 'Adopt a Microorganism' methodology The Internet has changed the way teachers and students access information and build knowledge. The recent COVID-19 pandemic has created challenges for both teachers and students and a demand for new methodologies of remote learning. In the life sciences, mixing online content with practical activities represents an even greater challenge. In microbiology, the implementation of an active teaching methodology, the #Adopt project, based on the social network Facebook(R), represents an excellent option for connecting remote education with classroom activities. In 2020, the version applied in high school, "Adopt a Microorganism", was adapted to meet the demands of emergency remote education owing to the suppression of face-to-face activities caused by the pandemic. In the present study, we assessed how the change in methodology impacted the discourse richness of students from high school integrated with technical education in the Business Administration program of the Federal Institute of Sao Paulo, Sorocaba Campus. Three questionnaires related to the groups of microorganisms (Archaea, Bacteria, Virus, Fungi, and Protozoan) were applied. The students responses in the 2019 and 2020 classes were compared concerning content richness and multiplicity of concepts through the application of the Shannon diversity index, an approach that is generally used to assess biodiversity in different environments. The observed results suggest that remote learning provided students with a conceptual basis and richness of content equivalent to that achieved by students subjected to the hybrid teaching model. In conclusion, this study suggests that the #Adopt project methodology increases students discourse richness in microbiology even without face-to-face traditional classes. First Report of Lactoria cornuta from Karnataka, West coast of India, Role of tropical cyclones in Range Extension India is known for its rich biodiversity and is fortunate to have several endemic species from different classes of vertebrates. India is home to 7.5% of the global fish diversity, with 91 endemic species of ray finned fishes, the actinopterygians. Some fish species have never been reported until recently, one such example is the long-horned Cowfish (Ostracidae), best known for inhabiting only certain areas of the Indo-Pacific oceans. However, this fish has been reported from Bay of Bengal, Costs of Goa and Lakshadweep. This is the first time this fish has been found on the Coasts of Karnataka, west coast of India. Given its morphology, the migration seems highly improbable since it can only endure lethargic swimming. In this article, we have explored the role of other external forces that could have contributed to its range extension in the east and west coast of India. In this paper we are exploring the role of Super Cyclones as a vital force in determining the expansion of its range. The fact of collection longhorn cowfish for the first time from the coast of Karwar coincides with the passage of Cyclone Ockhi on the west coast of India this suggests the possible role of super cyclone in its range extension. A year later, another specimen was recovered from a location very close to the first, which indicates the successful establishment of Lacturia cornuta in its new environment. Related morphometric and meristic studies of our specimens are consistent with previous studies reported in the Bay of Bengal. Electrocommunication signals indicate motivation to compete during dyadic interactions of an electric fish Animals across species compete for limited resources. While in some species competition behavior is solely based on own abilities, others assess their opponents to facilitate these interactions. Using cues and communication signals, contestants gather information about their opponent, adjust their behavior accordingly, and can thereby avoid high costs of escalating fights. We tracked electrocommunication signals, in particular "rises", and agonistic behaviors of the gymnotiform electric fish Apteronotus leptorhynchus in staged competition experiments. A larger body size relative to the opponent was the sole significant predictor for winners. Sex and the frequency of the continuously emitted electric field were only mildly influencing competition outcome. In males, correlations of body-size and winning were stronger than in females and, especially when losing against females, communication and agonistic interactions were enhanced, hinting towards males being more motivated to compete. Fish that lost competitions emitted the majority of rises in dependence on the competitors relative size and sex. The emission of rises was costly since it provoked ritualized biting or chasing behaviors by the other fish. Despite winners being accurately predictable based on rise numbers already after the initial 25 minutes, losers continued to emit rises. The number of rises emitted by losers and the duration of chasing behaviors depended in similar ways on physical attributes of contestants. The detailed evaluation of these correlations hint towards A. leptorhynchus adjusting their competition behavior according to mutual assessment, where rises could signal a losers motivation to continue assessment through ritualized fighting. Modulations in motor unit discharge are related to changes in fascicle length during isometric contractions The integration of electromyography (EMG) and ultrasound imaging has provided important information about the mechanisms of muscle activation and contraction. Unfortunately, EMG does not allow an accurate assessment of the interplay between the neural drive received by muscles, changes in fascicle length (FL) and the force/torque produced. We aimed to assess the relationship between modulations in tibialis anterior (TA) motor unit (MU) firing rate, FL and dorsiflexion torque (DT) using ultrasound-transparent high-density EMG electrodes. EMG and ultrasound images were recorded simultaneously from TA, using a 32-electrode silicon matrix, while performing isometric dorsiflexion, at diverse ankle joint positions (0{degrees} and 30{degrees} plantar flexion) and torques (20% and 40% of maximum). EMG signals were decomposed into individual MUs and changes in FL were assessed with a fascicle-tracking algorithm. MU firings were converted into a cumulative spike train (CST) that was cross-correlated with DT (CST-DT) and FL (CST-FL). High cross-correlations were found for CST-FL, 0.60 (range: 0.31-0.85) and CST-DT 0.71 (range: 0.31-0.88). Cross-correlation lags revealed that the delay between CST-FL (~75ms) was significantly smaller than CST-DT (~150ms, p<0.001). These delays affected the interpretation of MU recruitment/de-recruitment thresholds, with FL showing similar lengths for both recruitment and de-recruitment. This study is the first to demonstrate that changes in TA FL are closely related to both modulations in MU firing frequency and DT. These relationships allow assessment of the interplay between neural drive, muscle contraction and resultant torque, thereby providing a better understanding of the mechanisms responsible for the generation of muscle force.

NEW AND NOTEWORTHYBy employing ultrasound-transparent high-density surface EMG electrodes, we show that modulations in tibialis anterior motor unit discharge rate were closely related to both changes in its fascicle length and resultant torque. These relationships allowed quantifying delays between neural drive and muscle shortening as well as muscle shortening and torque during submaximal isometric contractions, providing an accurate estimation of the time required to generate muscle force and subsequent production of torque via the tendon. Male age and Wolbachia dynamics: Investigating how fast and why bacterial densities and cytoplasmic incompatibility strengths vary Endosymbionts can influence host reproduction and fitness to favor their maternal transmission. For example, endosymbiotic Wolbachia bacteria often cause cytoplasmic incompatibility (CI) that kills uninfected embryos fertilized by Wolbachia-modified sperm. Infected females can rescue CI, providing them a relative fitness advantage. Wolbachia-induced CI strength varies widely and tends to decrease as host males age. Since strong CI drives Wolbachia to high equilibrium frequencies, understanding how fast and why CI strength declines with male age is crucial to explaining age-dependent CIs influence on Wolbachia prevalence. Here, we investigate if Wolbachia densities and/or CI gene (cif) expression covary with CI-strength variation and explore covariates of age-dependent Wolbachia-density variation in two classic CI systems. wRi CI strength decreases slowly with Drosophila simulans male age (6%/ day), but wMel CI strength decreases very rapidly (19%/ day), yielding statistically insignificant CI after only three days of D. melanogaster emergence. Wolbachia densities and cif expression in testes decrease as wRi-infected males age, but both surprisingly increase as wMel-infected males age, and CI strength declines. We then tested if phage lysis, Octomom copy number (which impacts wMel density), or host immune expression covary with age-dependent wMel densities--only host immune expression correlated with density. Together, our results identify how fast CI strength declines with male age in two model systems and reveal unique relationships between male age, Wolbachia densities, cif expression, and host immunity. We discuss new hypotheses about the basis of age-dependent CI strength and its contributions to Wolbachia prevalence.

ImportanceWolbachia are the most common animal-associated endosymbionts due in large part to their manipulation of host reproduction. Many Wolbachia cause cytoplasmic incompatibility (CI) that kills uninfected host eggs. Infected eggs are protected from CI, favoring Wolbachia spread in natural systems and in transinfected mosquito populations where vector-control groups use strong CI to maintain pathogen-blocking Wolbachia at high frequencies for biocontrol of arboviruses. CI strength varies considerably in nature and declines as males age for unknown reasons. Here, we determine that CI strength weakens at different rates with age in two model symbioses. Wolbachia density and CI gene expression covary with wRi-induced CI strength in Drosophila simulans, but neither explain rapidly declining wMel-induced CI in aging D. melanogaster males. Patterns of host immune gene expression suggest a candidate mechanism behind age-dependent wMel densities. These findings inform how age-dependent CI may contribute to Wolbachia prevalence in natural systems and potentially in transinfected systems. Increased susceptibility to Chrysanthemum Yellows phytoplasma infection in Atcals7 ko plants is accompanied by enhanced expression of carbohydrate transporters. Callose deposition around sieve pores, under control of callose synthase 7 (AtCALS7), has been interpreted as a mechanical response to limit pathogen spread in phytoplasma-infected plants. Wild-type and Atcals7ko mutants were therefore employed to unveil the mode of involvement of CALS7 in the plants response to phytoplasma infection. The fresh weights of healthy and CY-(Chrysanthemum Yellows) phytoplasma-infected Arabidopsis wild-type and mutant plants indicated two superimposed effects of the absence of CALS7: a partial impairment of photo-assimilate transport and a stimulated phytoplasma proliferation as illustrated by a significantly increased phytoplasma titre in Atcal7ko mutants. Further studies solely dealt with the effects of CALS7 absence on phytoplasma growth. Phytoplasma infection affected sieve-element substructure to a larger extent in mutants than in wild-type plants, which was also true for the levels of some free carbohydrates. Moreover, infection induced a similar upregulation of gene expression of enzymes involved in sucrose cleavage (AtSUS5, AtSUS6) and transmembrane transport (AtSWEET11) in mutants and wild-type plants, but an increased gene expression of carbohydrate transmembrane transporters (AtSWEET12, AtSTP13, AtSUC3) in infected mutants only. It remains still unclear how the absence of AtCALS7 leads to gene upregulation and how an increased intercellular mobility of carbohydrates and possibly effectors contributes to a higher susceptibility. It is also unclear if modified sieve-pore structures in mutants allow a better spread of phytoplasmas giving rise to higher titre.

Author SummaryPhytoplasma infections are one of the most limiting factors for production of important crops all over the world. Phytoplasma disease epidemics can be handled mainly by insect-vector control using insecticides. Basic information about plant-phytoplasma interactions are still limited, nevertheless it is necessary to design new management and breeding strategies aimed to obtain more tolerant or resistant cultivars. Phytoplasmas are obligate intracellular parasites restricted to the phloem sieve tubes. Callose deposition at the sieve plates has been described since the  70s as a mechanical defence process to limit pathogen spread by occluding sieve pores. Studies reported 40 years later demonstrated that callose at the sieve pores is also involved in sieve-pore development and function and, hence, in mass-flow regulation, carbohydrate metabolism and distribution, and plant growth. Here, we reported on the role(s) of sieve-element callose in phytoplasma-infected Arabidopsis, using a mutant lacking AtCALS7, the enzyme responsible for callose synthesis in the sieve elements. The results indicate that loss of AtCAL7 appears to confer increased susceptibility to phytoplasma infection, due to alterations in expression of genes involved in sugar metabolism and membrane transport. In the long run, the identification of plant resistance or susceptibility traits against phytoplasmas will allow a complete re-organization of chemical control strategies, with obvious opportunities of reducing insecticide burden. ALLOPURINOL BLOCKS THE AORTIC ANEURYSM IN A MOUSE MODEL OF MARFAN SYNDROME VIA REDUCING AORTIC OXIDATIVE STRESS BackgroundThe pathogenesis and progression of aortic aneurysm in Marfan syndrome (MFS) involves dysregulated TGF-{beta} and nitric oxide signaling, altered hemodynamics, and biomechanical forces. Increasing evidence indicates that redox stress participates in MFS aortopathy, though its contribution is not well established. We reported elevated reactive oxygen species (ROS) formation and NADPH oxidase NOX4 upregulation in MFS patients and mice aortic samples. Here we address the contribution of xanthine oxidoreductase (XOR), which is transcribed as dehydrogenase (XDH) and then post-translationally modified to oxidase (XO), catabolizing mainly purines into uric acid and ROS.

Methods and ResultsWe report that in MFS patients, XOR protein levels increased in aortic samples. In MFS mice (Fbn1C1041G/+), XOR mRNA transcripts augmented in the aorta of young but not older MFS mice. Moreover, the enzymatic activity of XO form increased with respect to XDH. The administration of the XOR inhibitor allopurinol (ALO) blocked the progression of aortic root aneurysm in MFS mice. ALO was also protective when administrated before the appearance of aneurysm. MFS-induced elastic fiber fragmentation, fibrotic remodeling, nuclear translocation of pNRF2, and increased 3-nitrotyrosine levels in the aortic tunica media as well as endothelial dysfunction were all prevented by allopurinol. The large overproduction of H2O2 in MFS aorta did not occur in allopurinol-treated mice whose uric acid plasma levels remained unaltered.

ConclusionsThis study strengthens the concept that redox stress is an important determinant of aortic aneurysm formation and progression in MFS and supports a clinical trial for ALO in the pharmacological treatment of MFS aortopathy.

GRAPHICAL ABSTRACT

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/464182v3_ufig1.gif" ALT="Figure 1">
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org.highwire.dtl.DTLVardef@eab02aorg.highwire.dtl.DTLVardef@1d3b39eorg.highwire.dtl.DTLVardef@1b54d0forg.highwire.dtl.DTLVardef@1d61ad0_HPS_FORMAT_FIGEXP  M_FIG C_FIG Venomics and antivenomics of Indian spectacled cobra (Naja naja) from the Western Ghats Venom proteome profiling of Naja naja from the Western Ghats region in Kerala was achieved through SDS-PAGE and RP-HPLC followed by Q-TOF LC-MS/MS analysis, incorporating PEAKS and Novor assisted de novo sequencing methodologies. A total of 115 proteins distributed across 17 different enzymatic and non-enzymatic venom protein families were identified through conventional and 39 peptides through homology-driven proteomics approaches. Fourteen peptides derived through de novo complements the Mascot data indicating the importance of homology-driven approaches in improving protein sequence information. Among the protein families identified, glutathione peroxidase and endonuclease were reported for the first time in the Indian cobra venom. Immunological cross-reactivity assessed using Indian polyvalent antivenoms suggested that VINS showed better EC50 (2.48 g/mL) value than that of PSAV (6.04 g/mL) and Virchow (6.03 g/mL) antivenoms. Western blotting experiments indicated that all the antivenoms elicited poor binding specificities, especially towards low molecular mass proteins. Second-generation antivenomics studies revealed that VINS antivenom was less efficient to detect many low molecular mass proteins such as three-finger toxins and Kunitz-type serine protease Inhibitors. Taken together, the present study enabled a large-scale characterization of the venom proteome of Naja naja from the Western Ghats and emphasized the need for developing more efficient antivenoms.

HighlightsO_LIProteomics of cobra venom resulted in the identification of 115 proteins representing 17 snake venom protein families.
C_LIO_LIDe novo approaches exclusively yielded 39 peptides harbouring multiple amino acid mutations.
C_LIO_LIGlutathione peroxidase and endonuclease were identified for the first time in Indian cobra venom.
C_LIO_LIIndian polyvalent antivenoms showed varying cross-reactivity towards cobra venom.
C_LIO_LIVINS antivenom was less efficient to detect many low molecular mass proteins (< 20 kDa).
C_LI Alien chromatin other than the GST-encoding Fhb7 candidate confers Fusarium head blight resistance in wheat breeding The lack of resistance resources is a major bottleneck for wheat Fusarium head blight (FHB) resistance breeding. Three wheat-Th. elongatum FHB resistant translocation lines have been developed and used for wheat breeding without yield penalty. Transcriptomic analysis identified a derivative glutathione S-transferase transcript T26102, which was homologous to Fhb7 and induced dramatically by Fusarium graminearum. Unlike other studies, Fhb7 homologs were detected not only in Thinopyrum but also in Elymus, Leymus, Pseudoroegeria and Roegeria. We also found that several wheat-Th. ponticum derivatives carrying Fhb7 and its homologs were highly susceptible to FHB. Moreover, the transgenic plants expressing Fhb7 and its homolog on different backgrounds did not improve the FHB resistance.

One Sentence SummaryThe GST-encoding Fhb7 candidate cannot improve Fusarium head blight resistance in wheat breeding. Identification of two cancer stem cell-like populations in triple-negative breast cancer xenografts Gene expression analysis at the single-cell level by next generation sequencing has revealed the existence of clonal dissemination and microheterogeneity in cancer metastasis. The current spatial analysis technologies can elucidate the heterogeneity of cell-cell interactions in situ. To reveal the regional and expressional heterogeneity in primary tumors and metastases, we performed transcriptomic analysis of microtissues dissected from a triple-negative breast cancer (TNBC) cell line MDA-MB-231 xenograft model with our automated tissue microdissection punching technology. This multiple-microtissue transcriptome analysis revealed three cancer cell-type clusters in the primary tumor and axillary lymph node metastasis, two of which were cancer stem cell (CSC)-like clusters (CD44/MYC-high, HMGA1-high). Reanalysis of public single-cell RNA-seq (scRNA-seq) datasets confirmed that the two CSC-like populations existed both in TNBC xenograft models and TNBC patients. In addition, the gene signature of the HMGA1-high CSC-like cluster has the potential to serve as a novel biomarker for diagnosis. The diversity of these multiple CSC-like populations may cause differential anticancer drug resistance, increasing the difficulty of curing this cancer. Global Constraints within the Developmental Program of the Drosophila Wing Organismal development is a complex process, involving a vast number of molecular constituents interacting on multiple spatio-temporal scales in the formation of intricate body structures. Despite this complexity, development is remarkably reproducible and displays tolerance to both genetic and environmental perturbations. This robust-ness implies the existence of hidden simplicities in developmental programs. Here, using the Drosophila wing as a model system, we develop a new quantitative strategy that enables a robust description of biologically salient phenotypic variation. Analyzing natural phenotypic variation across a highly outbred population, and variation generated by weak perturbations in genetic and environmental conditions, we observe a highly constrained set of wing phenotypes. Remarkably, the phenotypic variants can be described by a single integrated mode that corresponds to a non-intuitive combination of structural variations across the wing. This work demonstrates the presence of constraints that funnel environmental inputs and genetic variation into phenotypes stretched along a single axis in morphological space. Our results provide quantitative insights into the nature of robustness in complex forms while yet accommodating the potential for evolutionary variations. Methodologically, we introduce a general strategy for finding such invariances in other developmental contexts. Diverse stem-chondrichthyan oral structures and evidence for an independently acquired acanthodid dentition The teeth of sharks famously form a series of transversely-organised files with conveyor-belt replacement that are borne directly on the jaw cartilages, in contrast to the dermal plate-borne dentition of bony fishes that undergoes site-specific replacement. A major obstacle in understanding how this system evolved is the poorly understood relationships of the earliest chondrichthyans and the profusion of morphologically and terminologically diverse bones, cartilages, splints and whorls that they possess. Here we use tomographic methods to investigate mandibular structures in several early branching  acanthodian-grade stem-chondrichthyans. We show that the dentigerous jaw bones of disparate genera of ischnacanthids are united by a common construction, being growing bones with non-shedding dentition. Mandibular splints, which support the ventro-lateral edge of the Meckels cartilage in some taxa, are formed from dermal bone and may be an acanthodid synapomorphy. We demonstrate that the teeth of Acanthodopsis are borne directly on the mandibular cartilage and that this taxon is deeply nested within an edentulous radiation, representing an unexpected independent origin of teeth. Many or even all of the range of unusual oral structures may be apomorphic, but they should nonetheless be considered when building hypotheses of tooth and jaw evolution, both in chondrichthyans and more broadly. Hnf1b-CreER causes efficient recombination of a Rosa26-RFP reporter in duct and islet δ cells The Hnf1b-CreERT2 BAC transgenic (Tg(Hnf1b-cre/ERT2)1Jfer) has been used extensively to trace the progeny of pancreatic ducts in developmental, regeneration, or cancer models. Hnf1b-CreERT2 transgenics have been used to show that cells from that form a duct-like plexus in the embryonic pancreas are bipotent duct-endocrine progenitors, whereas adult mouse duct cells are not a common source of {beta} cells in various regenerative settings. The interpretation of such genetic lineage tracing studies is critically dependent on a correct understanding of the cell type specificity of recombinase activity with each reporter system. We have re-examined the performance of Hnf1b-CreERT2 with a Rosa26-RFP reporter transgene. This showed inducible recombination of up to 96% adult duct cells, a much higher efficiency than previously used reporter transgenes. Despite this high duct-cell excision, recombination in  and {beta} cells remained very low, similar to previously used reporters. However, nearly half of somatostatin-expressing {delta} cells showed reporter activation, which was due to Cre expression in {delta} cells rather than to duct to {delta} cell conversions. The high recombination efficiency in duct cells indicates that the Hnf1b-CreERT2 model can be useful for both ductal fate mapping and genetic inactivation studies. The recombination in {delta} cells does not modify the interpretation of studies that failed to show duct conversions to other cell types, but needs to be considered if this model is used in studies that aim to modify the plasticity of pancreatic duct cells. Early loss of endogenous NAD+ following rotenone treatment leads to Sarm1 induction that is ameliorated by PARP inhibition Sarm1 is an evolutionary conserved innate immune adaptor protein that has emerged as a primary regulator of programmed axonal degeneration over the past decade. In vitro structural insights have revealed that although Sarm1 induces energy depletion by breaking down NAD+, it is also allosterically inhibited by NAD+. However, how NAD+ levels modulate the activation of intracellular Sarm1 has not been elucidated so far. This study focuses on understanding the events leading to Sarm1 activation in both neuronal and non-neuronal cells using the mitochondrial complex I inhibitor rotenone. Here we report the regulation of rotenone-induced cell death by loss of NAD+ that may act as a "biological trigger" of Sarm1 activation. Our study revealed that early loss of endogenous NAD+ levels arising due to PARP1 hyperactivation preceded Sarm1 induction following rotenone treatment. Interestingly, replenishing NAD+ levels by the PARP1 inhibitor, PJ34 restored mitochondrial homeostasis and prevented subsequent Sarm1 activation in rotenone treated cells. These cellular data were further validated in Drosophila melanogaster where a significant reduction in rotenone mediated loss of locomotor abilities and reduced dSarm expression was observed in the flies following PARP inhibition. Taken together, these observations not only uncovers a novel regulation of Sarm1 induction by endogenous NAD+ levels but also point towards an important understanding on how PARP inhibitors could be repurposed in the treatment of mitochondrial complex I deficiency disorders mediated by Sarm1. PKR-mediated Stress Response Enhances Dengue and Zika Virus Replication The mechanisms by which Flaviviruses use non-canonical translation to support their replication in host cells are largely unknown. Here we investigated how the integrated stress response (ISR), which promotes translational arrest by eIF2 phosphorylation (p-eIF2), regulates Flavivirus replication. During Dengue virus (DENV) and Zika virus (ZIKV) infection, eIF2 activation peaked at 24 hours post infection and was dependent on PKR but not type I interferon. The ISR is activated downstream of p-eIF2 during infection with either virus, but translation arrest only occurred following DENV infection. Despite this difference, both DENV and ZIKV replication was impaired in cells lacking PKR, independent of IFN-I/NF-kB signaling or cell viability. By using a ZIKV 5UTR reporter system as a model, we found that this region of the genome is sufficient to promote an enhancement of viral mRNA translation in the presence of an active ISR. Together we provide evidence that Flaviviruses escape ISR translational arrest and co-opt this response to increase viral replication. High-throughput analysis of DNA replication in single human cells reveals the complex nature of replication timing control DNA replication initiates from replication origins firing throughout S phase. Debate remains about whether origins are a fixed set of loci, or a loose agglomeration of potential sites used stochastically in individual cells, and about how consistent their firing time is. We developed an approach to profile DNA replication from whole-genome sequencing of thousands of single cells, which includes "in silico flow cytometry", a method for discriminating replicating and non-replicating cells. Using two microfluidic platforms, we analyzed up to 2,437 replicating cells from a single sample. The resolution and scale of the data allow focused analysis of replication initiation sites, demonstrating that most occur in confined genomic regions. While initiation order is remarkably similar across cells, we unexpectedly identify several subtypes of initiation regions in late-replicating regions. Taken together, high throughput, high resolution sequencing of individual cells reveals previously underappreciated variability in replication initiation and progression. Pyroptosis-related gene signatures can robustly diagnose skin cutaneous melanoma and predict the prognosis Skin cutaneous melanoma (SKCM) is a chronically malignant tumor with a high mortality rate. Pyroptosis, a kind of pro-inflammatory programmed cell death, has been linked to cancer in recent studies. However, the value of pyroptosis in the diagnosis and prognosis of SKCM is not clear. In this study, it was discovered that 20 pyroptosis-related genes (PRGs) differed in expression between SKCM and normal tissues, which were related to diagnosis and prognosis. Firstly, based on these genes, nine machine-learning algorithms were shown to perform well in constructing diagnostic classifiers, including K-Nearest Neighbor (KNN), logistic regression, Support Vector Machine (SVM), Artificial Neural Network (ANN), decision tree, random forest, XGBoost, LightGBM, and CatBoost. Secondly, the least absolute shrinkage and selection operator (LASSO) Cox regression analysis was applied and the prognostic model was constructed based on 9 PRGs. Subgroups in low and high risks determined by the prognostic model were shown to have different survival. Thirdly, functional enrichment analyses were performed by applying the gene set enrichment analysis (GSEA), and results suggested that the risk was related to immune response. Finally, immune infiltration analysis was carried out, which showed that fractions of activated CD4+ memory T cells, {gamma}{delta} T cells, Ml macrophages, and M2 macrophages were significantly different between subgroups. In conclusion, the expression signatures of pyroptosis-related genes are effective and robust in the diagnosis and prognosis of SKCM, which is related to immunity.