Miracle-dz/UniFAR-FTUs huggingface dataset:
RNA-directed DNA methylation prevents rapid and heritable reversal of transposon silencing under heat stress in Zea mays

In large complex plant genomes, RNA-directed DNA methylation (RdDM) ensures that epigenetic silencing is maintained at the boundary between genes and flanking transposable elements. In maize, RdDM is dependent on Modifer of Paramutation 1 (Mop1), a putative RNA dependent RNA polymerase. Here we show that although RdDM is essential for the maintenance of DNA methylation of a silenced MuDR transposon in maize, a loss of that methylation does not result in a restoration of activity. Instead, heritable maintenance of silencing is maintained by histone modifications. At one terminal inverted repeat (TIR) of this element, heritable silencing is mediated via H3K9 and H3K27 dimethylation, even in the absence of DNA methylation. At the second TIR, heritable silencing is mediated by H3K27 trimethylation, a mark normally associated with somatically inherited gene silencing. We find that a brief exposure of high temperature in a mop1 mutant rapidly reverses both of these modifications in conjunction with a loss of transcriptional silencing. These reversals are heritable, even in mop1 wild type progeny in which methylation is restored at both TIRs. These observations suggest that DNA methylation is neither necessary to maintain silencing, nor is it sufficient to initiate silencing once has been reversed. However, given that heritable reactivation only occurs in a mop1 mutant background, these observations suggest that DNA methylation is required to buffer the effects of environmental stress on transposable elements. Author Summary Most plant genomes are mostly transposable elements (TEs), most of which are held in check by modifications of both DNA and histones. The bulk of silenced TEs are associated with methylated DNA and histone H3 lysine 9 demethylation (H3K9me2). In contrast, epigenetically silenced genes are often associated with histone lysine 27 trimethylation (H3K27me3). Although stress can affect each of these modifications, plants are generally competent to rapidly reset them following that stress. Here we demonstrate that although DNA methylation is not required to maintain silencing of the MuDR element, it is essential for preventing heat-induced, stable and heritable changes in both H3K9me2 and H3K27me3 at this element, and for concomitant changes in transcriptional activity. These finding suggest that RdDM acts to buffer the effects of heat on silenced transposable elements, and that a loss of DNA methylation under conditions of stress can have profound and long lasting effects on epigenetic silencing in maize.
Differential phase register of Hes1 oscillations with mitoses underlies cell-cycle heterogeneity in ER+ breast cancer cells

Significance Tumors exhibit heterogeneities that are not due to mutations, including cancer stem cells with different potencies. We show that the cancer stem-cell state predisposed to dormancy in vivo has a highly variable and long cell cycle. Using single-cell live imaging for the transcriptional repressor Hes1 (a key molecule in cancer), we show a type of circadian-like oscillatory expression of Hes1 in all cells in the population. The most potent cancer stem cells tend to divide around the trough of the Hes1 oscillatory wave, a feature predictive of a long cell cycle. A concept proposed here is that the position of cell division with respect to the Hes1 wave is predictive of its prospective cell-cycle length and cancer cellular substate. Here, we study the dynamical expression of endogenously labeled Hes1, a transcriptional repressor implicated in controlling cell proliferation, to understand how cell-cycle length heterogeneity is generated in estrogen receptor (ER)+ breast cancer cells. We find that Hes1 shows oscillatory expression with ∼25 h periodicity and during each cell cycle has a variable peak in G1, a trough around G1–S transition, and a less variable second peak in G2/M. Compared to other subpopulations, the cell cycle in CD44HighCD24Low cancer stem cells is longest and most variable. Most cells divide around the peak of the Hes1 expression wave, but preceding mitoses in slow dividing CD44HighCD24Low cells appear phase-shifted, resulting in a late-onset Hes1 peak in G1. The position, duration, and shape of this peak, rather than the Hes1 expression levels, are good predictors of cell-cycle length. Diminishing Hes1 oscillations by enforcing sustained expression slows down the cell cycle, impairs proliferation, abolishes the dynamic expression of p21, and increases the percentage of CD44HighCD24Low cells. Reciprocally, blocking the cell cycle causes an elongation of Hes1 periodicity, suggesting a bidirectional interaction of the Hes1 oscillator and the cell cycle. We propose that Hes1 oscillations are functionally important for the efficient progression of the cell cycle and that the position of mitosis in relation to the Hes1 wave underlies cell-cycle length heterogeneity in cancer cell subpopulations.
Seeing the World Like Never Before: Human stereovision through perfect optics

Stereovision is the ability to perceive fine depth variations from small differences in the two eyes’ images. Using adaptive optics, we show that even minute optical aberrations that are not clinically correctable, and go unnoticed in everyday vision, can affect stereo acuity. Hence, the human binocular system is capable of using unnaturally fine details that are not encountered in everyday vision. More importantly, stereoacuity was still considerably variable even with perfect optics. This variability can be attributed to neural adaptation. Our visual system tries to compensate for these aberrations through neural adaptation that optimizes stereovision when viewing stimuli through one’s habitual optics. However, the same adaptation becomes ineffective when the optics are changed, even if improved. Beyond optical imperfections, we show that stereovision is limited by neural adaptation to one’s own optics. Significance statement Humans, and animals with front-facing eyes, view the world from slightly different vantage points. This creates small differences in the left and right images that can be utilized for fine depth perception (stereovision). Retinal images are also subject to imperfections that are often different in the optics of the two eyes. Using advanced optical correction techniques, we show that even the smallest imperfections that escape clinical detection affect stereovision. We also find that neural processes become adapted to a person’s own optics. Hence, stereovision is directly impacted by the optics of the eyes, and indirectly via neural adaptation. Since the optics change over the lifespan, our results imply that the adult binocular system is adaptable with possibilities for binocular rehabilitation.
Evolutionary Processes Driving the Rise and Fall of Staphylococcus aureus ST239, a Dominant Hybrid Pathogen

The rise of antibiotic resistance in most pathogenic bacteria has been driven by the spread of a small number of epidemically successful resistant strains. However, the processes that drive the rise and fall of these “superbugs” remain poorly understood. ABSTRACT Selection plays a key role in the spread of antibiotic resistance, but the evolutionary drivers of clinically important resistant strains remain poorly understood. Here, we use genomic analyses and competition experiments to study Staphylococcus aureus ST239, a prominent MRSA strain that is thought to have been formed by large-scale recombination between ST8 and ST30. Genomic analyses allowed us to refine the hybrid model for the origin of ST239 and to date the origin of ST239 to 1920 to 1945, which predates the clinical introduction of methicillin in 1959. Although purifying selection has dominated the evolution of ST239, parallel evolution has occurred in genes involved in antibiotic resistance and virulence, suggesting that ST239 has evolved toward an increasingly pathogenic lifestyle. Crucially, ST239 isolates have low competitive fitness relative to both ST8 and ST30 isolates, supporting the idea that fitness costs have driven the demise of this once-dominant pathogen strain. IMPORTANCE The rise of antibiotic resistance in most pathogenic bacteria has been driven by the spread of a small number of epidemically successful resistant strains. However, the processes that drive the rise and fall of these “superbugs” remain poorly understood. In our study, we investigated Staphylococcus aureus ST239, an important MRSA strain that has been a leading cause of serious hospital-acquired infections. We show here that ST239 was formed by the exchange of a very large fragment of DNA carrying resistance genes between strains of S. aureus sometime before 1945. The introduction of the antibiotic methicillin in 1959 provided a key advantage for Methicillin-Resistant Staphylococcus aureus (MRSA) strains. Interestingly, we found that ST239 has low competitive ability compared to other strains of S. aureus, and this evolutionary hindrance may explain why the prevalence of ST239 has declined precipitously at a global scale.
Impact of South African 501.V2 Variant on SARS-CoV-2 Spike Infectivity and Neutralization: A Structure-based Computational Assessment

Motivation The SARS-CoV-2 variants emerging from South Africa (501.V2) and the UK (B.1.1.7) necessitate rapid assessment of the effects of the corresponding amino acid substitutions in the spike (S) receptor-binding domain (RBD) of the variants on the interactions with the human ACE2 receptor and monoclonal antibodies (mAbs) reported earlier to neutralize the spike. Results Molecular modeling and simulations reveal that N501Y, shared by both variants, increases ACE2 binding affinity, and may impact the collective dynamics of the ACE2-RBD complex, occupying a central hinge site that modulates the overall dynamics of the complex. In contrast, the substitutions K417N and E484K in the South African variant 501.V2 would reduce the ACE2-binding affinity by abolishing two interfacial salt bridges that facilitate RBD binding to ACE2, K417(S)-D30(ACE2) and E484 (S)-K31(ACE2). These two mutations may thus be more than compensating the attractive effect induced by N501Y, overall resulting in an ACE2-binding affinity comparable to that of the wildtype RBD. Further analysis of the impact of these mutations on the interactions with mAbs targeting the spike indicate that the substitutions K417N and E484K may also abolish the salt bridges between the spike and selected mAbs, such as REGN10933, BD23, H11_H4, and C105, thus reducing the binding affinity and effectiveness of these mAbs. Contact bahar@pitt.edu Supplementary information Supplementary data are available at Bioinformatics online.
Transposable element landscape changes are buffered by RNA silencing in aging Drosophila

Genetic mechanisms that repress transposable elements (TEs) in young animals decline during aging, as reflected by increased TE expression in aged animals. Does increased TE expression during aging lead to more genomic TE copies in older animals? To answer this question, we quantified TE Landscapes (TLs) via whole genome sequencing of young and aged Drosophila strains of wild-type and mutant backgrounds. We quantified TLs in whole flies and dissected brains and validated the feasibility of our approach in detecting new TE insertions in aging Drosophila genomes when natural defenses like RNA interference (RNAi) pathways are compromised. By also incorporating droplet digital PCR to validate genomic TE loads, we confirm TL changes can occur in a single lifespan of Drosophila when TEs are not suppressed. We also describe improved sequencing methods to quantify extra-chromosomal DNA circles (eccDNAs) in Drosophila as an additional source of TE copies that accumulate during aging. Lastly, to combat the natural progression of aging-associated TE expression, we show that knocking down PAF1, a conserved transcription elongation factor that antagonizes RNAi pathways, may bolster suppression of TEs during aging and extend lifespan. Our study suggests that RNAi mechanisms generally mitigate genomic TL expansion despite the increase in TE transcripts during aging.
T cell stiffness is enhanced upon formation of immunological synapse

T cells are activated by cognate target cells via an intimate contact, termed immunological synapse (IS). Cellular mechanical properties, especially stiffness, are essential to regulate cell functions, T cell stiffness at a subcellular level at the IS still remains largely elusive. In this work, we established an atomic force microscopy (AFM)-based elasticity mapping method on whole T cells to obtain an overview of the stiffness with a resolution of ~ 60 nm. Using Jurkat T-cells and primary human CD4+ T cells, we show that in the T cells in contact with functionalized surfaces, the lamellipodia are stiffer than the cell body. Upon IS formation, T cell stiffness is substantially enhanced both at the lamellipodia and in cell body. Chelation of intracellular Ca2+ abolishes IS-induced stiffening at the lamellipodia but has no influence on cell body-stiffening, suggesting different regulatory mechanism of IS-induced stiffening between the lamellipodia and the cell body.
Single Cell RNA-seq and Mass Cytometry Reveals a Novel and a Targetable Population of Macrophages in Idiopathic Pulmonary Fibrosis

In this study, we leveraged a combination of single cell RNAseq, cytometry by time of flight (CyTOF), and flow cytometry to study the biology of a unique macrophage population in pulmonary fibrosis. Using the profiling data from 312,928 cells derived from 32 idiopathic pulmonary fibrosis (IPF), 29 healthy control and 18 chronic obstructive pulmonary disease (COPD) lungs, we identified an expanded population of macrophages in IPF that have a unique transcriptional profile associated with pro-fibrotic signature. These macrophages attain a hybrid transitional state between alveolar and interstitial macrophages, are enriched with biological processes of pro-fibrotic immune cells, and express novel surface markers and genes that have not been previously reported. We then applied single cell CyTOF to simultaneously measure 37 markers to precisely phenotype the uniquely expanded macrophage subset in IPF lungs. The SPADE algorithm independently identified an expanded macrophage cluster, and validated CD84 and CD36 as novel surface markers that highly label this cluster. Using a separate validation cohort, we confirmed an increase in CD84++CD36++ macrophage population in IPF compared to control and COPD lungs by flow cytometry. Further, using the signature from the IPF-specific macrophages and the LINCS drug database, we predicted small molecules that could reverse the signature of IPF-specific macrophages, and validated two molecules, CRT and Cucur, using THP-1 derived human macrophages and precision-cut lung slices (PCLS) from IPF patients. Utilizing a multi-dimensional translational approach, our work identified a novel and targetable population of macrophages found in end-stage pulmonary fibrosis. One Sentence Summary Single cell RNAseq, CyTOF, and flow cytometry reveal the presence of an aberrant macrophage population in pulmonary fibrosis
Report on the 20th International Radiocarbon Conference Graphitization Workshop

A sample preparation workshop was convened at the 20th International Radiocarbon conference, with about 30–40 attendees. The term “sample preparation” was not further specified to allow participants to address the laboratory issues most important to them. Given the short time available, the focus quickly narrowed to details of graphite preparation for accelerator mass spectrometry (AMS), with particular interest in procedures for small samples, and hence we have changed the title to reflect the focus. Here, we summarize the workshop discussion, adding additional background information and references to publications. Several aspects of graphite preparation are included: catalyst types and ratio of catalyst to carbon; optimal reaction temperatures for various sample sizes; methods for water removal; types of pressure transducers; preconditioning of catalyst; and problems with blank values in very small samples. The pros and cons of sample dilution were also discussed.
CcLBD25 functions as a key regulator of haustorium development in Cuscuta campestris

Parasitic plants reduce yield of crops worldwide. Cuscuta campestris is a stem parasite that attaches to its host, using haustoria to extract nutrients and water. We analyzed the transcriptome of six C. campestris tissues and identified a key gene, CcLBD25, as highly expressed in prehaustoria and haustoria. Our gene co-expression networks (GCN) from different tissue types and laser-capture microdissection (LCM) RNA-Seq data indicate that CcLBD25 could be essential for regulating cell wall loosening and organogenesis. We employed host-induced gene silencing (HIGS) by generating transgenic tomato hosts that express hairpin RNAs to target and down-regulate CcLBD25 in the parasite. Our results showed that C. campestris growing on CcLBD25 RNAi transgenic tomatoes transited to the flowering stage earlier and had less biomass compared with C. campestris growing on wild type host. This suggests that the parasites growing on the transgenic plants were stressed due to insufficient nutrient acquisition. Anatomy of C. campestris haustoria growing on CcLBD25 RNAi tomatoes showed reduced pectin digestion and lack of searching hyphae, which interfered with haustorium penetration and the formation of vascular connections. We developed an in vitro haustorium (IVH) system to assay the number of prehaustoria produced on strands from C. campestris. When C. campestris was grown on CcLBD25 RNAi tomatoes or wild type tomatoes, the former produce fewer prehaustoria than the latter, indicating that down-regulating CcLBD25 may affect haustorium initiation. The results of this study shed light on the role of CcLBD25 in haustorium development and might help to develop a parasite-resistant system in crops. One-sentence summary CcLBD25 plays a pivotal role in haustorium initiation, regulating pectin digestion, and searching hyphae development during the haustorium penetration process.
The pathway of starch synthesis in Arabidopsis thaliana leaves

Many plants accumulate transitory starch reserves in their leaves during the day to buffer their carbohydrate supply against fluctuating light conditions, and to provide carbon and energy for survival at night. It is universally accepted that transitory starch is synthesized from ADP-glucose (ADPG) in the chloroplasts. However, the consensus that ADPG is made in the chloroplasts by ADPG pyrophosphorylase has been challenged by a controversial proposal that ADPG is made primarily in the cytosol, probably by sucrose synthase (SUS), and then imported into the chloroplasts. To resolve this long-standing controversy, we critically re-examined the experimental evidence that appears to conflict with the consensus pathway. We show that when precautions are taken to avoid artefactual changes during leaf sampling, Arabidopsis thaliana mutants that lack SUS activity in mesophyll cells (quadruple sus1234) or have no SUS activity (sextuple sus123456) have wild-type levels of ADPG and starch, while ADPG is 20 times lower in the pgm and adg1 mutants that are blocked in the classical pathway of starch synthesis. We conclude that the ADPG needed for starch synthesis in leaves is synthesized primarily by ADPG pyrophosphorylase in the chloroplasts. Significance statement Mutant analysis shows that sucrose synthase makes no significant contribution to transitory starch synthesis in Arabidopsis leaves, resolving a 20-year old controversy about one of the most important pathways of photosynthetic metabolism.
Development and Nutrient Evaluation of Oat Supplemented Products for Old Aged People with Constipation

Abstract Constipation refers to bowel movements that are infrequent. Severe constipation includes obstipation and faecal impaction found very commonly in old aged. The first step to treat constipation is by increasing the intake of dietary fibre. Oats can be supplemented in a normal carbohydrate meal without the loss of taste, palatability. Supplementary product samples of sweet dalia, salty daliaand khichdiwere prepared using broken wheat and rice as control while for test samples broken wheat for both sweet and salty daliaand rice were supplemented with oats at 20 percent , 25 percent and 30 percent levels. The control and the most acceptable level of oat supplementation in the products were analysed for proximate composition (moisture, crude protein, crude fat, crude fibre, total ash) carbohydrates and energy were calculated. Fibre fractions namely NDF, ADF and lignin were also estimated. Taste of salty daliasupplemented with 25 percent scored the best score of 8.0 while all the sensory parameters of sweet daliasupplemented with oats at same percentage had the highest scores. Addition of oats at 25 percent level in sweet/salty daliaand khichdiincreased the moisture, crude protein and crude fibre value. While the fibre fractions of NDF, ADF and lignin also increased in the supplemented products.
Using micro-Raman spectroscopy for nasopharyngeal cancerous tissue detection

Micro-Raman spectroscopy is widely used for non-invasive tissue diagnosis and detection, as it provides detailed information about biomolecular composition, structure, and interaction of tissue. In this work, micro-Raman spectroscopy was used to investigate non-cancerous and cancerous nasopharyngeal tissues. The obtained nasopharyngeal tissue samples in vitro are divided into two groups: cancerous (n=12, undifferentiated non-keratinizing carcinomas) and non-cancerous (n=10, 7 chronic inflammations, 2 lymphomas and 1 lymphocytosis). Firstly, we analyzed the Raman spectra in the fingerprint (FP, 400-1800cm-1) region acquired. Preliminary results showed that there are some spectral differences in different pathological conditions. Furthermore, Raman spectra from cancerous and non-cancerous nasopharyngeal tissue in the high wavenumber region (HW, 2800-3100cm-1) were also reported for the first time. After detailed analysis, we achieved significant differences in Raman bands at 2854, 2874, 2934, and 3067cm-1 between cancerous and non-cancerous nasopharyngeal tissues. This study demonstrates that both fingerprint and high wavenumber regions of micro-Raman spectroscopy have the potential for the early detection of nasopharyngeal carcinomas.