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Proteolysis targeting chimera (PROTAC) is a state‐of‐the‐art technology for ablating undruggable targets. A PROTAC degrader achieves targeted protein degradation (TPD) through the simultaneous binding of a protein of interest (POI) and an E3 ligase to form a ternary complex. A nanofibril‐based PROTAC strategy to form a polynary (E3)m : PROTAC : (POI)n complex has not been reported in the TPD field up to this point. A recent innovation shows that a POI ligand and E3 ligase ligand don't have to be within a fused degrader molecule. Instead, they can be recruited to cellular proximity by a self‐assembly‐driving peptide and click chemistry. The resulting nanofibrils can recruit multiple POI and E3 ligase molecules to form a polynary complex as a degradation center. The so‐called Nano‐PROTAC provides a novel approach for TPD in cancer therapy. [ABSTRACT FROM AUTHOR]

Proteolysis targeting chimera (PROTAC) is a state‐of‐the‐art technology for ablating undruggable targets. A PROTAC degrader achieves targeted protein degradation (TPD) through the simultaneous binding of a protein of interest (POI) and an E3 ligase to form a ternary complex. A nanofibril‐based PROTAC strategy to form a polynary (E3)m : PROTAC : (POI)n complex has not been reported in the TPD field up to this point. A recent innovation shows that a POI ligand and E3 ligase ligand don't have to be within a fused degrader molecule. Instead, they can be recruited to cellular proximity by a self‐assembly‐driving peptide and click chemistry. The resulting nanofibrils can recruit multiple POI and E3 ligase molecules to form a polynary complex as a degradation center. The so‐called Nano‐PROTAC provides a novel approach for TPD in cancer therapy. [ABSTRACT FROM AUTHOR]

Epigenetics, the inheritance of genomic information independent of DNA sequence, controls the interpretation of extracellular and intracellular signals in cell homeostasis, proliferation and differentiation. On the chromatin level, signal transduction leads to changes in epigenetic marks, such as histone post-translational modifications (PTMs), DNA methylation and chromatin accessibility to regulate gene expression. Crosstalk between different epigenetic mechanisms, such as that between histone PTMs and DNA methylation, leads to an intricate network of chromatin-binding proteins where pre-existing epigenetic marks promote or inhibit the writing of new marks. The recent technical advances in mass spectrometry (MS) -based proteomic methods and in genome-wide DNA sequencing approaches have broadened our understanding of epigenetic networks greatly. However, further development and wider application of these methods is vital in developing treatments for disorders and pathologies that are driven by epigenetic dysregulation. [ABSTRACT FROM AUTHOR]

Complex vector modes of light, non-separable in their spatial and polarisation degrees of freedom, are revolutionising a wide variety of research fields. It is therefore not surprising that the generation techniques have evolved quite dramatically since their inception. At present it is common to use computer-controlled devices, among which digital micromirror devices have become popular. Some of the reason for this are their low-cost, their polarisation-insensitive and their high-refresh rates. As such, in this manuscript we put forward a novel technique characterised by its high stability, which is achieved through a common-path interferometer. We demonstrate the capabilities of this technique experimentally, first by generating arbitrary vector modes on a higher-order Poincaré sphere, secondly, by generating vector modes in different coordinates systems and finally, by generating various vector modes simultaneously. Our technique will find applications in fields such as optical manipulations, optical communications, optical metrology, among others. [ABSTRACT FROM AUTHOR]

In this work, we present an introduction to the field of spatially partial coherent beams, while keeping in mind the transverse structure of an optical field. We look closely at the concept of spatial coherence and show some strategies to deal with it. We work stepâ€"byâ€"step with the reader and construct as an example, a partially coherent vortex beam. Finally, using numerical simulations, the richness in structure of a partially coherent field is revealed through its crossâ€"correlation function. [ABSTRACT FROM AUTHOR]

Sequencing technologies, in particular RNASeq, have become critical tools in the design, build, test and learn cycle of synthetic biology. They provide a better understanding of synthetic designs, and they help identify ways to improve and select designs. While these data are beneficial to design, their collection and analysis is a complex, multistep process that has implications on both discovery and reproducibility of experiments. Additionally, tool parameters, experimental metadata, normalization of data and standardization of file formats present challenges that are computationally intensive. This calls for high-throughput pipelines expressly designed to handle the combinatorial and longitudinal nature of synthetic biology. In this paper, we present a pipeline to maximize the analytical reproducibility of RNASeq for synthetic biologists. We also explore the impact of reproducibility on the validation of machine learning models. We present the design of a pipeline that combines traditional RNASeq data processing tools with structured metadata tracking to allow for the exploration of the combinatorial design in a high-throughput and reproducible manner. We then demonstrate utility via two different experiments: a control comparison experiment and a machine learning model experiment. The first experiment compares datasets collected from identical biological controls across multiple days for two different organisms. It shows that a reproducible experimental protocol for one organism does not guarantee reproducibility in another. The second experiment quantifies the differences in experimental runs from multiple perspectives. It shows that the lack of reproducibility from these different perspectives can place an upper bound on the validation of machine learning models trained on RNASeq data. Graphical Abstract [ABSTRACT FROM AUTHOR]

We present an experimental realization of the first iteration in Grover's quantum algorithm using classical light and linear optical elements. The algorithm serves to find an entry marked by an oracle in an unstructured database. In our scheme, the quantum states encoding the database are represented by helical modes generated by means of a Spatial Light Modulator, while the marking corresponds to a π phase shift of the hidden mode. The optical implementation of Grover's algorithm then selectively amplifies the intensity of the marked mode such that it can be revealed by a modal decomposition. The core of the algorithm - a geometrical reflection of the point representing all database entries - is implemented in a single step independent of the size of the database. Moreover, we demonstrate experimentally that one iteration of the algorithm is enough to identify the marked entry, as a consequence of using classical states of light. [ABSTRACT FROM AUTHOR]

Drought stress is a recurring feature of world climate and the single most important factor influencing agricultural yield worldwide. Plants display highly variable, species-specific responses to drought and these responses are multifaceted, requiring physiological and morphological changes influenced by genetic and molecular mechanisms. Moreover, the reproducibility of water deficit studies is very cumbersome, which significantly impedes research on drought tolerance, because how a plant responds is highly influenced by the timing, duration, and intensity of the water deficit. Despite progress in the identification of drought-related mechanisms in many plants, the molecular basis of drought resistance remains to be fully understood in trees, particularly in poplar species because their wide geographic distribution results in varying tolerances to drought. Herein, we aimed to better understand this complex phenomenon in eastern cottonwood (Populus deltoides) by performing a detailed contrast of the proteome changes between two different water deficit experiments to identify functional intersections and divergences in proteome responses. We investigated plants subjected to cyclic water deficit and compared these responses to plants subjected to prolonged acute water deficit. In total, we identified 108,012 peptide sequences across both experiments that provided insight into the quantitative state of 22,737 Populus gene models and 8,199 functional protein groups in response to drought. Together, these datasets provide the most comprehensive insight into proteome drought responses in poplar to date and a direct proteome comparison between short period dehydration shock and cyclic, post-drought re-watering. Overall, this investigation provides novel insights into drought avoidance mechanisms that are distinct from progressive drought stress. Additionally, we identified proteins that have been associated as drought-relevant in previous studies. Importantly, we highlight the RD26 transcription factor as a gene regulated at both the transcript and protein level, regardless of species and drought condition, and, thus, represents a key, universal drought marker for Populus species. [ABSTRACT FROM AUTHOR]

Mass spectrometry (MS) is an essential part of the cell biologist's proteomics toolkit, allowing analyses at molecular and system-wide scales. However, proteomics still lag behind genomics in popularity and ease of use. We discuss key differences between MS-based -omics and other booming -omics technologies and highlight what we view as the future of MS and its role in our increasingly deep understanding of cell biology. [ABSTRACT FROM AUTHOR]

In 1804, Thomas Young reported the observation of fringes in the intensity of light, and attributed it to the concept of interference between coherent sources. In this paper, we revisit this famous experiment and show how it can easily be demonstrated with digital holography. We look closely at the concept of interference with light and ask, "fringes in what?" We then show that depending on how light interferes, fringe patterns in observables other than intensity can be seen. We explain this conceptually and demonstrate it experimentally. We provide a holistic approach to the topic, aided by modern laboratory practices for a straightforward demonstration of the underlying physics. [ABSTRACT FROM AUTHOR]

We propose an optical implementation of the Deutsch–Jozsa Algorithm using classical light in a binary decision-tree scheme. Our approach uses a ring cavity and linear optical devices in order to efficiently query the oracle functional values. In addition, we take advantage of the intrinsic Fourier transforming properties of a lens to read out whether the function given by the oracle is balanced or constant. [ABSTRACT FROM AUTHOR]

Tuberculosis and nontuberculous mycobacterial infections constitute a high burden of pulmonary disease in humans, resulting in over 1.5 million deaths per year. Building on the premise that genetic factors influence the instance, progression, and defense of infectious disease, we undertook a systems biology approach to investigate relationships among genetic factors that may play a role in increased susceptibility or control of mycobacterial infections. We combined literature and database mining with network analysis and pathway enrichment analysis to examine genes, pathways, and networks, involved in the human response to Mycobacterium tuberculosis and nontuberculous mycobacterial infections. This approach allowed us to examine functional relationships among reported genes, and to identify novel genes and enriched pathways that may play a role in mycobacterial susceptibility or control. Our findings suggest that the primary pathways and genes influencing mycobacterial infection control involve an interplay between innate and adaptive immune proteins and pathways. Signaling pathways involved in autoimmune disease were significantly enriched as revealed in our networks. Mycobacterial disease susceptibility networks were also examined within the context of gene-chemical relationships, in order to identify putative drugs and nutrients with potential beneficial immunomodulatory or anti-mycobacterial effects. [ABSTRACT FROM AUTHOR]

Background: Central to most omic scale experiments is the interpretation and examination of resulting gene lists corresponding to differentially expressed, regulated, or observed gene or protein sets. Complicating interpretation is a lack of functional annotation assigned to a large percentage of many microbial genomes. This is particularly noticeable in mycobacterial genomes, which are significantly divergent from many of the microbial model species used for gene and protein functional characterization, but which are extremely important clinically. Mycobacterial species, ranging from M. tuberculosis to M. abscessus, are responsible for deadly infectious diseases that kill over 1.5 million people each year across the world. A better understanding of the coding capacity of mycobacterial genomes is therefore necessary to shed increasing light on putative mechanisms of virulence, pathogenesis, and functional adaptations. Description: Here we describe the improved functional annotation coverage of 11 important mycobacterial genomes, many involved in human diseases including tuberculosis, leprosy, and nontuberculous mycobacterial (NTM) infections. Of the 11 mycobacterial genomes, we provide 9899 new functional annotations, compared to NCBI and TBDB annotations, for genes previously characterized as genes of unknown function, hypothetical, and hypothetical conserved proteins. Functional annotations are available at our newly developed web resource MycoBASE (Mycobacterial Annotation Server) at strong.ucdenver.edu/mycobase. Conclusion: Improved annotations allow for better understanding and interpretation of genomic and transcriptomic experiments, including analyzing the functional implications of insertions, deletions, and mutations, inferring the function of understudied genes, and determining functional changes resulting from differential expression studies. MycoBASE provides a valuable resource for mycobacterial researchers, through improved and searchable functional annotations and functional enrichment strategies. MycoBASE will be continually supported and updated to include new genomes, enabling a powerful resource to aid the quest to better understand these important pathogenic and environmental species. [ABSTRACT FROM AUTHOR]

We present an implementation of the Deutsch Algorithm using linear optical elements and laser light. We encoded two quantum bits in form of superpositions of electromagnetic fields in two degrees of freedom of the beam: its polarisation and orbital angular momentum. Our approach, based on a Sagnac interferometer, offers outstanding stability and demonstrates that optical quantum computation is possible using classical states of light. [ABSTRACT FROM AUTHOR]

We propose an off-axis superposition of vector modes with orthogonal polarizations, constructed from a general scalar helical vortex mode with unknown topological charge m, as a method to measure its orbital angular momentum. We derived analytic expressions for sets of solutions to find lines of linear polarization (L lines) within the composite polarization field. We found that the solutions corresponding to the angular component of the composite field depend only on the displacement of the beams and the topological charge m, and they are invariant under propagation and changes in the relative amplitude and phase between the beams. [ABSTRACT FROM AUTHOR]

Background: Although respiratory diseases exhibit in a wide array of clinical manifestations, certain respiratory diseases may share related genetic mechanisms or may be influenced by similar chemical stimuli. Here we explore and infer relationships among genes, diseases, and chemicals using network and matrix based clustering methods. Results: In order to better understand and elucidate these shared genetic mechanisms and chemical relationships we analyzed a comprehensive collection of gene, disease, and chemical relationships pertinent to respiratory disease, using network and matrix based analysis approaches. Our methods enabled us to analyze relationships and make biological inferences among over 200 different respiratory and related diseases, involving thousands of gene-chemical-disease relationships. Conclusions: The resulting networks provided insight into shared mechanisms of respiratory disease and in some cases suggest novel targets or repurposed drug strategies. [ABSTRACT FROM AUTHOR]

Histone post-translational modifications (PTMs) have been linked to a variety of biological processes and disease states, thus making their characterization a critical field of study. In the last 5 years, a number of novel sites and types of modifications have been discovered, greatly expanding the histone code. Mass spectrometric methods are essential for finding and validating histone PTMs. Additionally, novel proteomic, genomic and chemical biology tools have been developed to probe PTM function. In this snapshot review, proteomic tools for PTM identification and characterization will be discussed and an overview of PTMs found in the last 5 years will be provided. [ABSTRACT FROM AUTHOR]

We study the dynamics of polarization singularities in the superposition of two off-axis optical vortices with orthogonal polarization states. The motion of C points, states of circular polarization, is described by varying parameters such as separation distance, constant phase difference and topological charge. We analyze the zeros of the Stokes parameters and find analytical solutions for particular cases. The results could be used to realize an alternative method to measure the vortex topological charge. [ABSTRACT FROM AUTHOR]

Protein methylation is a post-translational modification (PTM) which modulates cellular and biological processes including transcription, RNA processing, protein interactions and protein dynamics. Methylation, catalyzed by highly specific methyltransferase enzymes, occurs on several amino acids including arginine, lysine, histidine and dicarboxylic amino acids like glutamate. Mass spectrometry (MS)-based techniques continue to be the methods of choice for the study of protein PTMs. These approaches are powerful and sensitive tools that have been used to identify, quantify and characterize protein methylation. In addition, metabolic labeling strategies can be coupled to MS detection in order to measure dynamic and differential in vivo protein methylation rates. In this review, different applications of mass spectrometry technologies and methods to study protein methylation are discussed. [ABSTRACT FROM AUTHOR]

Mass spectrometry (MS) research has revolutionized modern biological and biomedical fields. At the heart of the majority of mass spectrometry experiments is the use of Bottom Up mass spectrometry methods where proteins are first proteolyzed into smaller fragments before MS interrogation. The advent of electron capture dissociation and, more recently, electron-transfer dissociation, however, has allowed Top Down (analysis of intact proteins) or middle down (analysis of large polypeptides) mass spectrometry to both experience large increases in development, growth, and overall usage. Nevertheless, for high-throughput large-scale proteomic studies, Bottom Up mass spectrometry has easily dominated the field. As Top Down mass spectrometry methodology and technology continue to develop, will it genuinely be able to compete with Bottom Up mass spectrometry for whole proteome analysis? Discussed here are the current approaches, applications, issues, and future view of high-throughput Top Down mass spectrometry. [Copyright &y& Elsevier]

Post-translational modifications (PTMs) of histones play an important role in many cellular processes, notably gene regulation. Using a combination of mass spectrometric and immunobiochemical approaches, we show that the PTM profile of histone H3 differs significantly among the various model organisms examined. Unicellular eukaryotes, such as Saccharomyces cerevisiae (yeast) and Tetrahymena thermophila (Tet), for example, contain more activation than silencing marks as compared with mammalian cells (mouse and human), which are generally enriched in PTMs more often associated with gene silencing. Close examination reveals that many of the better-known modified lysines (Lys) can be either methylated or acetylated and that the overall modification patterns become more complex from unicellular eukaryotes to mammals. Additionally, novel species-specific H3 PTMs from wild-type asynchronously grown cells are also detected by mass spectrometry. Our results suggest that some PTMs are more conserved than previously thought, including H3K9me1 and H4K20me2 in yeast and H3K27me1, -me2, and -me3 in Tet. On histone H4, methylation at Lys-20 showed a similar pattern as H3 methylation at Lys-9, with mammals containing more methylation than the unicellular organisms. Additionally, modification profiles of H4 acetylation were very similar among the organisms examined. [ABSTRACT FROM AUTHOR]

Histone proteins and their accompanying post-translational modifications have received much attention for their ability to affect chromatin structure and, hence, regulate gene expression. Recently, mass spectrometry has become an important complementary tool for the analysis of histone variants and modification sites, for determining the degree of occupancy of these modifications and for quantifying differential expression of these modifications from various samples. Additionally, as advancements in mass spectrometry technologies continue, the ability to read entire ‘histone codes’ across large regions of histone polypeptides or intact protein is possible. As chromatin biology demands, mass spectrometry has adapted and continues as a key technology for the analysis of gene regulation networks involving histone modifications. [Copyright &y& Elsevier]

Abstract: Post-translational modifications (PTMs) of histones are intimately involved in chromatin structure and thus have roles in cellular processes through their impact on gene activation or repression. At the forefront in histone PTM analysis are mass spectrometry-based techniques, which have capabilities to produce improved views of processes affected by chromatin remodeling via histone modifications. In this report, we take the first mass spectrometric look at histone variant expression and post-translational modifications from histones isolated from rat brain tissue. Analyses of whole rat brain identified specific histone H2A and H2B gene family members and several H4 and H3 post-translational modification sites by electron capture dissociation (ECD) mass spectrometry. We subsequently compared these results to selected rat brain regions. Major differences in the expression profiles of H2A and H2B gene family members or in the post-translational modifications on histone H4 were not observed from the different brain regions using a Top Down approach. However, “Middle Down” mass spectrometry facilitating improved characterization of the histone H3 tail (1–50 residues), revealed an enrichment of trimethylation on Lys9 from cerebellum tissue compared to H3 extracted from whole brain, cerebral cortex or hypothalamus tissue. We forward this study in honor of Professor Donald F. Hunt, whose pioneering efforts in protein and PTM analyses have spawned new eras and numerous careers, many exemplified in this special issue. [Copyright &y& Elsevier]

Careful regulation of the Wnt–Β-catenin signaling pathway is critical to many aspects of development and cancer. Casein kinase Iε is a Wnt-activated positive regulator of this pathway. Members of the Dishevelled family have been identified as key substrates of casein kinase I (CKI). However, the specific sites phosphorylated in vivo by CKI and their relative importance in the physiologic regulation of these proteins in the canonical Wnt–β-catenin signaling pathway remain unclear. To address this question, recombinant mouse Dishevelled (mDvl-1) was phosphorylated by CKI in vitro and phosphorylation sites were identified by MS. CKI phosphorylation of mDvl-1 at two highly conserved residues, serines 139 and 142, was observed by MS and confirmed by phosphopeptide mapping of in vivo phosphorylated protein. Phosphorylation of these sites is dependent on casein kinase I epsilon activity in vivo. Phenotypic analysis of mutant mDvl-1 indicates that phosphorylation of these sites stimulates the Dvl-activated β-catenin-dependent Wnt signaling pathway in both cell culture and in Xenopus development. Casein kinase I epsilon is a Wnt-regulated kinase, and regulated phosphorylation of Dvl allows fine tuning of the Wnt–β-catenin signaling pathway. [ABSTRACT FROM AUTHOR]

Covalent histone modifications and the incorporation of histone variants bring about changes in chromatin structure that in turn alter gene expression. Interest in non-allelic histone variants has been renewed, in part because of recent work on H3 (and other) histone variants. However, only in mammals do three non-centromeric H3 variants (H3.1, H3.2, and H3.3) exist. Here, we show that mammalian cell lines can be separated into two different groups based on their expression of H3.1, H3.2, and H3.3 at both mRNA and protein levels. Additionally, the ratio of these variants changes slightly during neuronal differentiation of murine ES cells. This difference in H3 variant expression between cell lines could not be explained by changes in growth rate, cell cycle stages, or chromosomal ploidy, but rather suggests other possibilities, such as changes in H3 variant incorporation during differentiation and tissue- or species-specific H3 variant expression. Moreover, quantitative mass spectrometry analysis of human H3.1, H3.2, and H3.3 showed modification differences between these three H3 variants, suggesting that they may have different biological functions. Specifically, H3.3 contains marks associated with transcriptionally active chromatin, whereas H3.2, in contrast, contains mostly silencing modifications that have been associated with facultative heterochromatin. Interestingly, H3.1 is enriched in both active and repressive marks, although the latter marks are different from those observed in H3.2. Although the biological significance as to why mammalian cells differentially employ three highly similar H3 variants remains unclear, our results underscore potential functional differences between them and reinforce the general view that H3.1 and H3.2 in mammalian cells should not be treated as equivalent proteins. [ABSTRACT FROM AUTHOR]

Phosphorylation of histone H3 is a hallmark event in mitosis and is associated with chromosome condensation. Here, we use a combination of immobilized metal affinity chromatography and tandem mass spectrometry to characterize post-translational modifications associated with phosphorylation on the N-terminal tails of histone H3 variants purified from mitotically arrested HeLa cells. Modifications observed in vivo on lysine residues adjacent to phosphorylated Ser and Thr provide support for the existence of the "methyl/phos", binary-switch hypothesis [Fischle, W., Wang, Y., and Allis, C. D. (2003) Nature 425, 475-479]. ELISA with antibodies selective for H3 at Ser10, Ser28, and Thr3 show reduced activity when adjacent Lys residues are modified. When used together, mass spectrometry and immunoassay methods provide a powerful approach for elucidation of the histone code and identification of histone post-translational modifications that occur during mitosis and other specific cellular events. [ABSTRACT FROM AUTHOR]

The article demonstrates that the serine 31 phosphorylation of histone variant H3.3 is specific to regions bordering centromeres in metaphase chromosomes. Histones are the fundamental components of the nucleosome. The histone H3 family contains an evolutionary conserved variant H3.3, which differs in sequence in only five amino acids from the canonical H3, H3.1, and was shown to play a role in the transcriptional activation of genes. Histone H3.3 contains a serine to alanine replacement at amino acid position 31.

Abstract: The reversible phosphorylation of proteins is recognized as an essential post-translational modification regulating cell signaling and ultimately function of biological systems. Detection of phosphopeptides and localization of phosphorylation sites remains quite a challenge, even if the protein is purified to near homogeneity. Mass spectrometry has become a vital technique that is routinely utilized for the identification of proteins from whole cell lysates. Nonetheless, due to the minimal amount of phosphorylation found on proteins, enrichment steps for isolating phosphopeptides from complex mixtures have been the focus of many research groups world-wide. In this review, we describe some current methods for the enrichment of phosphopeptides that are compatible with mass spectrometry for assignment of phosphorylation sites. Phosphorylation modifications on proteins and peptides are either directly isolated by solid-phase approaches or chemically modified for selective isolation and/or improved characterization by mass spectrometry. These strategies hold the potential for rapid and sensitive profiling of phosphoproteins from a variety of sources and cellular conditions. [Copyright &y& Elsevier]

Examines the use of volatile solvents for the preparation of thin-layer matrix deposits of oligonucleotides analysis. Formation of thin homogeneous film; Application of matrix-assisted laser desorption/ionization mass spectrometry to nucleic acids; Local distribution of matrix crystals.

A hallmark of addiction is the ability of drugs of abuse to trigger relapse after periods of prolonged abstinence. Here, we describe an epigenetic mechanism whereby chronic cocaine exposure causes lasting chromatin and downstream transcriptional modifications in the nucleus accumbens (NAc), a critical brain region controlling motivation. We link prolonged withdrawal from cocaine to the depletion of the histone variant H2A.Z, coupled with increased genome accessibility and latent priming of gene transcription, in D1 dopamine receptor-expressing medium spiny neurons (D1 MSNs) that relate to aberrant gene expression upon drug relapse. The histone chaperone ANP32E removes H2A.Z from chromatin, and we demonstrate that D1 MSN-selective Anp32e knockdown prevents cocaine-induced H2A.Z depletion and blocks cocaine's rewarding actions. By contrast, very different effects of cocaine exposure, withdrawal, and relapse were found for D2 MSNs. These findings establish histone variant exchange as an important mechanism and clinical target engaged by drugs of abuse to corrupt brain function and behavior. [ABSTRACT FROM AUTHOR]

ABSTRACT Nucleic acids are fundamental biological molecules that encode and convey genetic information within living organisms. Over 150 modifications have been found in nucleic acids, which are involved in critical biological functions, including regulating gene expression, stabilizing nucleic acid structure, modulating protein translation, and so on. The dysregulation of nucleic acid modifications is correlated with many diseases such as cancers and neurological disorders. However, it is still challenging to simultaneously characterize and quantify diverse modifications using traditional genomic methods. Mass spectrometry (MS) has served as a crucial tool to solve this issue, and can directly identify the modified species through their distinct mass differences compared to the canonical ones and provide accurate quantitative information. This review surveys the history of nucleic acid modification discovery, advancements in MS‐based methods, nucleic acid sample preparation, and applications in biological and medical research. We expect the high‐throughput and valuable quantitative information from MS analysis will be more broadly applied to studying nucleic acid modification status in different pathological conditions, which is key to filling gaps in traditional genomics and transcriptomics research and enabling researchers to gain insights into epigenetics and epitranscriptomics. [ABSTRACT FROM AUTHOR]

Abstract: Advances in multiplex qRT-PCR have enabled increasingly accurate and robust quantification of RNA, even at lower concentrations, facilitating RNA expression profiling in clinical and environmental samples. Here we describe a data-driven qRT-PCR normalization method, the minimum variance method, and evaluate it on clinically derived Mycobacterium tuberculosis samples with variable transcript detection percentages. For moderate to significant amounts of nondetection (∼50%), our minimum variance method consistently produces the lowest false discovery rates compared to commonly used data-driven normalization methods. [Copyright &y& Elsevier]

Serial capture affinity purification (SCAP) is a powerful method to isolate a specific protein complex. When combined with cross-linking mass spectrometry and computational approaches, one can build an integrated structural model of the isolated complex. Here, we applied SCAP to dissect a subpopulation of WDR76 in complex with SPIN1, a histone reader that recognizes trimethylated histone H3 lysine4 (H3K4me3). In contrast to a previous SCAP analysis of the SPIN1:SPINDOC complex, histones and the H3K4me3 mark were enriched with the WDR76:SPIN1 complex. Next, interaction network analysis of copurifying proteins and microscopy analysis revealed a potential role of the WDR76:SPIN1 complex in the DNA damage response. Since we detected 149 pairs of cross-links between WDR76, SPIN1, and histones, we then built an integrated structural model of the complex where SPIN1 recognized the H3K4me3 epigenetic mark while interacting with WDR76. Finally, we used the powerful Bayesian Integrative Modeling approach as implemented in the Integrative Modeling Platform to build a model of WDR76 and SPIN1 bound to the nucleosome. [ABSTRACT FROM AUTHOR]

We developed a platform using hydrophilic interaction chromatography and high-resolution tandem mass spectrometry (MS) for analyses of histone H3 that allows comprehensive characterization of 'histone codes' at the molecular level. We identified over 150 differentially modified forms of histone H3.2 in asynchronously grown and butyrate-treated HeLa cells, revealing pervasive combinatorial modification previously unaccounted for by other techniques and providing a clarified estimate of the molecular diversity of histone H3 in mammals. [ABSTRACT FROM AUTHOR]

The etiology of neurodevelopmental disorders (NDDs) remains a challenge for researchers. Human brain development is tightly regulated and sensitive to cellular alterations caused by endogenous or exogenous factors. Intriguingly, the surge of clinical sequencing studies has revealed that many of these disorders are monogenic and monoallelic. Notably, chromatin regulation has emerged as highly dysregulated in NDDs, with many syndromes demonstrating phenotypic overlap, such as intellectual disabilities, with one another. Here we discuss epigenetic writers, erasers, readers, remodelers, and even histones mutated in NDD patients, predicted to affect gene regulation. Moreover, this review focuses on disorders associated with mutations in enzymes involved in histone acetylation and methylation, and it highlights syndromes involving chromatin remodeling complexes. Finally, we explore recently discovered histone germline mutations and their pathogenic outcome on neurological function. Epigenetic regulators are mutated at every level of chromatin organization. Throughout this review, we discuss mechanistic investigations, as well as various animal and iPSC models of these disorders and their usefulness in determining pathomechanism and potential therapeutics. Understanding the mechanism of these mutations will illuminate common pathways between disorders. Ultimately, classifying these disorders based on their effects on the epigenome will not only aid in prognosis in patients but will aid in understanding the role of epigenetic machinery throughout neurodevelopment. [ABSTRACT FROM AUTHOR]

Premature birth disrupts normal lung development and places infants at risk for bronchopulmonary dysplasia (BPD), a disease disrupting lung health throughout the life of an individual and that is increasing in incidence. The TGF-β superfamily has been implicated in BPD pathogenesis, however, what cell lineage it impacts remains unclear. We show that TGFbr2 is critical for alveolar epithelial (AT1) cell fate maintenance and function. Loss of TGFbr2 in AT1 cells during late lung development leads to AT1-AT2 cell reprogramming and altered pulmonary architecture, which persists into adulthood. Restriction of fetal lung stretch and associated AT1 cell spreading through a model of oligohydramnios enhances AT1-AT2 reprogramming. Transcriptomic and proteomic analyses reveal the necessity of TGFbr2 expression in AT1 cells for extracellular matrix production. Moreover, TGF-β signaling regulates integrin transcription to alter AT1 cell morphology, which further impacts ECM expression through changes in mechanotransduction. These data reveal the cell intrinsic necessity of TGF-β signaling in maintaining AT1 cell fate and reveal this cell lineage as a major orchestrator of the alveolar matrisome. [ABSTRACT FROM AUTHOR]

Glycosylation is one of the most complex posttranslational protein modifications. Its importance has been established not only for eukaryotes but also for a variety of prokaryotic cellular processes, such as biofilm formation, motility, and mating. However, comprehensive glycoproteomic analyses are largely missing in prokaryotes. Here, we extend the phenotypic characterization of N-glycosylation pathway mutants in Haloferax volcanii and provide a detailed glycoproteome for this model archaeon through the mass spectrometric analysis of intact glycopeptides. Using in-depth glycoproteomic datasets generated for the wild-type (WT) and mutant strains as well as a reanalysis of datasets within the Archaeal Proteome Project (ArcPP), we identify the largest archaeal glycoproteome described so far. We further show that different N-glycosylation pathways can modify the same glycosites under the same culture conditions. The extent and complexity of the Hfx. volcanii N-glycoproteome revealed here provide new insights into the roles of N-glycosylation in archaeal cell biology. A comprehensive glycoproteomic analysis of Haloferax volcanii reveals the extent and complexity of glycosylation in archaea and provides new insights into the roles of this post-translational modification in various cellular processes, including cell shape determination. [ABSTRACT FROM AUTHOR]

Complex vector light modes, classically entangled in their spatial and polarization degrees of freedom (DoF), have become ubiquitous in a vast diversity of research fields. Crucially, while polarization is limited to a bi-dimensional space, the spatial mode is unbounded, and it can be specified by any of the sets of solutions the wave equation can support in different coordinate systems. Here, we report on a class of vector beams with elliptical symmetry where the spatial DoF is encoded in the Ince–Gaussian modes of the cylindrical elliptical coordinates. We outline their geometric representation on the higher-order Poincaré sphere, demonstrate their experimental generation, and analyze the quality of the generated modes via Stokes polarimetry. We anticipate that such vector modes will be of great relevance in applications, such as optical manipulations, laser material processing, and optical communications among others. [ABSTRACT FROM AUTHOR]

Biomolecular condensates (BMCs) play important roles in diverse biological processes. Many viruses form BMCs which have been implicated in various functions critical for the productive infection of host cells. The adenovirus L1-52/55 kilodalton protein (52K) was recently shown to form viral BMCs that coordinate viral genome packaging and capsid assembly. Although critical for packaging, we do not know how viral condensates are regulated during adenovirus infection. Here we show that phosphorylation of serine residues 28 and 75 within the N-terminal intrinsically disordered region of 52K modulates viral condensates in vitro and in cells, promoting liquidlike properties. Furthermore, we demonstrate that phosphorylation of 52K promotes viral genome packaging and the production of infectious progeny particles. Collectively, our findings provide insights into how viral condensate properties are regulated and maintained in a state conducive to their function in viral progeny production. In addition, our findings have implications for antiviral strategies aimed at targeting the regulation of viral BMCs to limit viral multiplication. [ABSTRACT FROM AUTHOR]

Vector beams display varying polarisation over planes transversal to their direction of propagation. The variation of polarisation implies that the electric field cannot be expressed as a product of a spatial mode and its polarisation. This non-separability has been analysed for particular vector beams in terms of non–quantum entanglement between the spatial and the polarisation-degrees of freedom, and equivalently, with respect to the degree of polarisation of light. Here we demonstrate theoretically and experimentally that Mathieu–Gauss vector modes violate a Bell-like inequality known as the Clauser–Horn–Shimony–Holt–Bell inequality. This demonstration provides new insights on the violation of Bell inequalities by a more general class of vector modes with elliptical symmetry. [ABSTRACT FROM AUTHOR]

Arboleda-Tham Syndrome (ARTHS) is a rare genetic disorder caused by heterozygous, de novo mutations in Lysine(K) acetyltransferase 6A (KAT6A). ARTHS is clinically heterogeneous and characterized by several common features, including intellectual disability, developmental and speech delay, and hypotonia, and affects multiple organ systems. KAT6A is the enzymatic core of a histone–acetylation protein complex; however, the direct histone targets and gene regulatory effects remain unknown. In this study, we use ARTHS patient (n = 8) and control (n = 14) dermal fibroblasts and perform comprehensive profiling of the epigenome and transcriptome caused by KAT6A mutations. We identified differential chromatin accessibility within the promoter or gene body of 23% (14/60) of genes that were differentially expressed between ARTHS and controls. Within fibroblasts, we show a distinct set of genes from the posterior HOXC gene cluster (HOXC10, HOXC11, HOXC-AS3, HOXC-AS2, and HOTAIR) that are overexpressed in ARTHS and are transcription factors critical for early development body segment patterning. The genomic loci harboring HOXC genes are epigenetically regulated with increased chromatin accessibility, high levels of H3K23ac, and increased gene–body DNA methylation compared to controls, all of which are consistent with transcriptomic overexpression. Finally, we used unbiased proteomic mass spectrometry and identified two new histone post-translational modifications (PTMs) that are disrupted in ARTHS: H2A and H3K56 acetylation. Our multi-omics assays have identified novel histone and gene regulatory roles of KAT6A in a large group of ARTHS patients harboring diverse pathogenic mutations. This work provides insight into the role of KAT6A on the epigenomic regulation in somatic cell types. [ABSTRACT FROM AUTHOR]

The nucleolus is sensitive to stress and can orchestrate a chain of cellular events in response to stress signals. Despite being a growth factor, FGF2 has antiproliferative and tumor-suppressive functions in some cellular contexts. In this work, we investigated how the antiproliferative effect of FGF2 modulates chromatin-, nucleolus- and rDNA-associated proteins. The chromatin and nucleolar proteome indicated that FGF2 stimulation modulates proteins related to transcription, rRNA expression and chromatinremodeling proteins. The global transcriptional rate and nucleolus area increased along with nucleolar disorganization upon 24 h of FGF2 stimulation. FGF2 stimulation induced immature rRNA accumulation by increasing rRNA transcription. The rDNAassociated protein analysis reinforced that FGF2 stimulus interferes with transcription and rRNA processing. RNA Pol I inhibition partially reversed the growth arrest induced by FGF2, indicating that changes in rRNA expression might be crucial for triggering the antiproliferative effect. Taken together, we demonstrate that the antiproliferative FGF2 stimulus triggers significant transcriptional changes and modulates the main cell transcription site, the nucleolus. [ABSTRACT FROM AUTHOR]