The Face of Chromatin Variants.

A developmental program affecting human face shape is shown by Greenberg et al. (2019) to hinge on the ability to distinguish a single methyl group between two histone variant isoforms and the action of the chromatin-remodeling enzyme SRCAP. This challenges researchers to link atomic structure to a morphological defect.

Mechanisms and functions of ATP-dependent chromatin-remodeling enzymes.

Chromatin provides both a means to accommodate a large amount of genetic material in a small space and a means to package the same genetic material in different chromatin states. Transitions between chromatin states are enabled by chromatin-remodeling ATPases, which catalyze a diverse range of structural transformations. Biochemical evidence over the last two decades suggests that chromatin-remodeling activities may have emerged by adaptation of ancient DNA translocases to respond to specific features of chromatin. Here, we discuss such evidence and also relate mechanistic insights to our understanding of how chromatin-remodeling enzymes enable different in vivo processes.

RNA polymerase II promotes the organization of chromatin following DNA replication.

Understanding how chromatin organisation is duplicated on the two daughter strands is a central question in epigenetics. In mammals, following the passage of the replisome, nucleosomes lose their defined positioning and transcription contributes to their re-organisation. However, whether transcription plays a greater role in the organization of chromatin following DNA replication remains unclear. Here we analysed protein re-association with newly replicated DNA upon inhibition of transcription using iPOND coupled to quantitative mass spectrometry. We show that nucleosome assembly and the re-establishment of most histone modifications are uncoupled from transcription. However, RNAPII acts to promote the re-association of hundreds of proteins with newly replicated chromatin via pathways that are not observed in steady-state chromatin. These include ATP-dependent remodellers, transcription factors and histone methyltransferases. We also identify a set of DNA repair factors that may handle transcription-replication conflicts during normal transcription in human non-transformed cells. Our study reveals that transcription plays a greater role in the organization of chromatin post-replication than previously anticipated.

ATRX: Put me on repeat.

Mutations in the chromatin-remodeling protein ATRX cause alpha thalassaemia and mental retardation, but the severity of the disorder is independent of the specific mutation. In this issue of Cell, Law et al. (2010) demonstrate that ATRX alters gene expression by binding to G-rich tandem repeats, and the degree of transcriptional silencing caused by ATRX mutations correlates with the number of repeats.

Cerebrospinal fluid HIV RNA escape syndrome.

A 54-year-old man with treated HIV developed a subacute deterioration of speech, mobility and cognition. Cerebrospinal fluid (CSF) analysis showed a raised protein and a discordant CSF HIV RNA paired with plasma HIV RNA, confirming the diagnosis of CSF HIV RNA escape syndrome. It is important to consider this diagnosis in people with treated HIV who develop new neurological symptoms.

Classification of new morbillivirus and jeilongvirus sequences from bats sampled in Brazil and Malaysia.

As part of a broad One Health surveillance effort to detect novel viruses in wildlife and people, we report several paramyxovirus sequences sampled primarily from bats during 2013 and 2014 in Brazil and Malaysia, including seven from which we recovered full-length genomes. Of these, six represent the first full-length paramyxovirid genomes sequenced from the Americas, including two that are the first full-length bat morbillivirus genome sequences published to date. Our findings add to the vast number of viral sequences in public repositories, which have been increasing considerably in recent years due to the rising accessibility of metagenomics. Taxonomic classification of these sequences in the absence of phenotypic data has been a significant challenge, particularly in the subfamily Orthoparamyxovirinae, where the rate of discovery of novel sequences has been substantial. Using pairwise amino acid sequence classification (PAASC), we propose that five of these sequences belong to members of the genus Jeilongvirus and two belong to members of the genus Morbillivirus. We also highlight inconsistencies in the classification of Tupaia virus and Mòjiang virus using the same demarcation criteria and suggest reclassification of these viruses into new genera. Importantly, this study underscores the critical importance of sequence length in PAASC analysis as well as the importance of biological characteristics such as genome organization in the taxonomic classification of viral sequences.

Implementing public involvement throughout the research process-Experience and learning from the GPs in EDs study.

BACKGROUND: Public involvement in health services research is encouraged. Descriptions of public involvement across the whole research cycle of a major study are uncommon and its effects on research conduct are poorly understood. AIM: This study aimed to describe how we implemented public involvement, reflect on process and effects in a large-scale multi-site research study and present learning for future involvement practice. METHOD: We recorded public involvement roles and activities throughout the study and compared these to our original public involvement plan included in our project proposal. We held a group interview with study co-applicants to explore their experiences, transcribed the recorded discussion and conducted thematic analysis. We synthesized the findings to develop recommendations for future practice. RESULTS: Public contributors' activities went beyond strategic study planning and management to include active involvement in data collection, analysis and dissemination. They attended management, scrutiny, planning and task meetings. They also facilitated public involvement through annual planning and review sessions, conducted a Public Involvement audit and coordinated public and patient input to stakeholder discussions at key study stages. Group interview respondents said that involvement exceeded their expectations. They identified effects such as changes to patient recruitment, terminology clarification and extra dissemination activities. They identified factors enabling effective involvement including team and leader commitment, named support contact, building relationships and demonstrating equality and public contributors being confident to challenge and flexible to meet researchers' timescales and work patterns. There were challenges matching resources to roles and questions about the risk of over-professionalizing public contributors. CONCLUSION: We extended our planned approach to public involvement and identified benefits to the research process that were both specific and general. We identified good practice to support effective public involvement in health services research that study teams should consider in planning and undertaking research. PUBLIC CONTRIBUTION: This paper was co-conceived, co-planned and co-authored by public contributors to contribute research evidence, based on their experiences of active involvement in the design, implementation and dissemination of a major health services research study.

Young people's experiences of COVID-19 messaging at the start of the UK lockdown: lessons for positive engagement and information sharing.

BACKGROUND: To reduce COVID-19 infection rates during the initial stages of the pandemic, the UK Government mandated a strict period of restriction on freedom of movement or 'lockdown'. For young people, closure of schools and higher education institutions and social distancing rules may have been particularly challenging, coming at a critical time in their lives for social and emotional development. This study explored young people's experiences of the UK Government's initial response to the pandemic and related government messaging. METHODS: This qualitative study combines data from research groups at the University of Southampton, University of Edinburgh and University College London. Thirty-six online focus group discussions (FGDs) were conducted with 150 young people (Southampton: n = 69; FGD = 7; Edinburgh: n = 41; FGD = 5; UCL: n = 40; FGD = 24). Thematic analysis was conducted to explore how young people viewed the government's response and messaging and to develop recommendations for how to best involve young people in addressing similar crises in the future. RESULTS: The abrupt onset of lockdown left young people shocked, confused and feeling ignored by government and media messaging. Despite this, they were motivated to adhere to government advice by the hope that life might soon return to normal. They felt a responsibility to help with the pandemic response, and wanted to be productive with their time, but saw few opportunities to volunteer. CONCLUSIONS: Young people want to be listened to and feel they have a part to play in responding to a national crisis such as the COVID-19 epidemic. To reduce the likelihood of disenfranchising the next generation, Government and the media should focus on developing messaging that reflects young people's values and concerns and to provide opportunities for young people to become involved in responses to future crises.

URSID γ-HERPESVIRUS TYPE 1-RELATED VIRUS IN CAPTIVE BORNEAN SUN BEARS (HELARCTOS MALAYANUS EURYSPILUS) IN SABAH, MALAYSIA.

The Bornean sun bear (Helarctos malayanus euryspilus) is the smallest subspecies of sun bear. Their numbers are declining, and more research is needed to better understand their health and biology. Forty-four bears housed at the Bornean Sun Bear Conservation Centre (BSBCC) in Sabah, Malaysia, were screened for known and novel viruses in November 2018. Ursid γ-herpesvirus type 1 (UrHV-1) is a herpesvirus that has been detected from swab samples of clinically healthy sun bears and biopsy samples of oral squamous cell carcinoma in sun bears. We detected an UrHV-1-related virus from throat and rectal swabs by molecular viral screening in samples from 15.9% of the sun bears at BSBCC. None of the bears with the UrHV-1-related virus in this study had oral lesions. There is no known report of UrHV-1 detection in the wild sun bear population, and its association with oral squamous cell carcinoma is not fully understood. Finding an UrHV-1-related virus in a rehabilitation center is a concern because conditions in captivity may contribute to spreading this virus, and there is the potential of introducing it into wild populations when a bear is released. This study demonstrates an urgent need to carry out similar surveillance for sun bears in captivity as well as those in the wild, to better understand the impact of captivity on the prevalence and spread of UrHV-1-related viruses. Positive bears also should be monitored for oral lesions to better understand whether there is a causal relationship.

Publisher Correction: BAF complex vulnerabilities in cancer demonstrated via structure-based PROTAC design.

In the version of this article originally published, several lines of text in the last paragraph of the right column on page 1 of the PDF were transposed into the bottom paragraph of the left column. The affected text of the left column should read "The ATP-dependent activities of the BAF (SWI/SNF) chromatin remodeling complexes affect the positioning of nucleosomes on DNA and thereby many cellular processes related to chromatin structure, including transcription, DNA repair and decatenation of chromosomes during mitosis12,13." The affected text of the right column should read "SMARCA2/4BD inhibitors are thus precluded from use for the treatment of SMARCA4 mutant cancers but could provide attractive ligands for PROTAC conjugation. Small molecules binding to other bromodomains have been successfully converted into PROTACs by conjugating them with structures capable of binding to the E3 ligases von Hippel-Lindau (VHL) or cereblon5,6,10,11,25,26,27." The errors have been corrected in the PDF version of the paper.

BAF complex vulnerabilities in cancer demonstrated via structure-based PROTAC design.

Targeting subunits of BAF/PBAF chromatin remodeling complexes has been proposed as an approach to exploit cancer vulnerabilities. Here, we develop proteolysis targeting chimera (PROTAC) degraders of the BAF ATPase subunits SMARCA2 and SMARCA4 using a bromodomain ligand and recruitment of the E3 ubiquitin ligase VHL. High-resolution ternary complex crystal structures and biophysical investigation guided rational and efficient optimization toward ACBI1, a potent and cooperative degrader of SMARCA2, SMARCA4 and PBRM1. ACBI1 induced anti-proliferative effects and cell death caused by SMARCA2 depletion in SMARCA4 mutant cancer cells, and in acute myeloid leukemia cells dependent on SMARCA4 ATPase activity. These findings exemplify a successful biophysics- and structure-based PROTAC design approach to degrade high profile drug targets, and pave the way toward new therapeutics for the treatment of tumors sensitive to the loss of BAF complex ATPases.

The first reported cases of elephant endotheliotropic herpesvirus infectious haemorrhagic disease in Malaysia: case report.

BACKGROUND: Elephant endotheliotropic herpesvirus haemorrhagic disease (EEHV HD) is the leading cause of death in captive Asian elephant calves in Asia, North America, and Europe with a mortality rate of ~ 65% in calves that are under human care. Although EEHV HD was first found in elephant camps, more recently it was identified in wild populations which poses a greater threat to the elephant population. Deaths due to EEHV HD have been seen in wild elephants, but the in-situ prevalence and mortality rate is unknown. We report the first EEHV HD cases in Malaysia from 3 wild born endangered Bornean elephant calves from Sabah with known typical clinical signs. CASE PRESENTATION: The first calf died within 24 h of the onset of clinical signs; the second calf died within 12 h of the onset of clinical signs. The third calf succumbed within 72 h. Necropsies revealed that all 3 calves had similar presentations of EEHV HD but in the third calf with less severity. We conducted conventional polymerase chain reaction (cPCR) assays and found EEHV DNA at all 7 loci in the 3 calves; it was identified as EEHV1A, the virus type that has been found in most other reported cases. CONCLUSION: Typical EEHV HD clinical signs and the molecular confirmation of EEHV by cPCR and sequencing point to EEHV as the cause of death. Further genetic investigation of the strain is in progress.

PICH: a DNA translocase specially adapted for processing anaphase bridge DNA.

The Plk1-interacting checkpoint helicase (PICH) protein localizes to ultrafine anaphase bridges (UFBs) in mitosis alongside a complex of DNA repair proteins, including the Bloom's syndrome protein (BLM). However, very little is known about the function of PICH or how it is recruited to UFBs. Using a combination of microfluidics, fluorescence microscopy, and optical tweezers, we have defined the properties of PICH in an in vitro model of an anaphase bridge. We show that PICH binds with a remarkably high affinity to duplex DNA, resulting in ATP-dependent protein translocation and extension of the DNA. Most strikingly, the affinity of PICH for binding DNA increases with tension-induced DNA stretching, which mimics the effect of the mitotic spindle on a UFB. PICH binding also appears to diminish force-induced DNA melting. We propose a model in which PICH recognizes and stabilizes DNA under tension during anaphase, thereby facilitating the resolution of entangled sister chromatids.

Identification of Elg1 interaction partners and effects on post-replication chromatin re-formation.

Elg1, the major subunit of a Replication Factor C-like complex, is critical to ensure genomic stability during DNA replication, and is implicated in controlling chromatin structure. We investigated the consequences of Elg1 loss for the dynamics of chromatin re-formation following DNA replication. Measurement of Okazaki fragment length and the micrococcal nuclease sensitivity of newly replicated DNA revealed a defect in nucleosome organization in the absence of Elg1. Using a proteomic approach to identify Elg1 binding partners, we discovered that Elg1 interacts with Rtt106, a histone chaperone implicated in replication-coupled nucleosome assembly that also regulates transcription. A central role for Elg1 is the unloading of PCNA from chromatin following DNA replication, so we examined the relative importance of Rtt106 and PCNA unloading for chromatin reassembly following DNA replication. We find that the major cause of the chromatin organization defects of an ELG1 mutant is PCNA retention on DNA following replication, with Rtt106-Elg1 interaction potentially playing a contributory role.

Unilateral akathisia and choreoathetosis as an unusual presentation of a frontal lobe dermoid cyst.

OBJECTIVE: To consider an unusual presentation of a frontal lobe dermoid cyst. Intracranial masses very rarely present with movement disorders. We describe a highly unusual presentation of an intracranial dermoid with unilateral choreoathetosis, akathisia and facial tics. MATERIAL: A 63-year-old man presented with left-sided akathisia and tardive dyskinesia of his upper limb, attributed to a dermoid cyst in the right frontal lobe. Resection of the cyst led to a complete resolution of his symptoms without neurological deficit. CONCLUSION: Pathogenic mechanisms of such symptoms may include compression and ischaemia of the basal ganglia and other nigrostriatal structures, as well as dysfunction of the supplementary motor area and mesolimbic dopaminergic pathways.

Diagnosis and management of acute ischaemic stroke.

Acute ischaemic stroke is a major public health priority and will become increasingly relevant to neurologists of the future. The cornerstone of effective stroke care continues to be timely reperfusion treatment. This requires early recognition of symptoms by the public and first responders, triage to an appropriate stroke centre and efficient assessment and investigation by the attending stroke team. The aim of treatment is to achieve recanalisation and reperfusion of the ischaemic penumbra with intravenous thrombolysis and/or endovascular thrombectomy in appropriately selected patients. All patients should be admitted directly to an acute stroke unit for close monitoring for early neurological deterioration and prevention of secondary complications. Prompt investigation of the mechanism of stroke allows patients to start appropriate secondary preventative treatment. Future objectives include improving accessibility to endovascular thrombectomy, using advanced imaging to extend therapeutic windows and developing neuroprotective agents to prevent secondary neuronal damage.

Diagnosis of spotted fever group Rickettsia infections: the Asian perspective.

Spotted fever group rickettsiae (SFG) are a neglected group of bacteria, belonging to the genus Rickettsia, that represent a large number of new and emerging infectious diseases with a worldwide distribution. The diseases are zoonotic and are transmitted by arthropod vectors, mainly ticks, fleas and mites, to hosts such as wild animals. Domesticated animals and humans are accidental hosts. In Asia, local people in endemic areas as well as travellers to these regions are at high risk of infection. In this review we compare SFG molecular and serological diagnostic methods and discuss their limitations. While there is a large range of molecular diagnostics and serological assays, both approaches have limitations and a positive result is dependent on the timing of sample collection. There is an increasing need for less expensive and easy-to-use diagnostic tests. However, despite many tests being available, their lack of suitability for use in resource-limited regions is of concern, as many require technical expertise, expensive equipment and reagents. In addition, many existing diagnostic tests still require rigorous validation in the regions and populations where these tests may be used, in particular to establish coherent and worthwhile cut-offs. It is likely that the best strategy is to use a real-time quantitative polymerase chain reaction (qPCR) and immunofluorescence assay in tandem. If the specimen is collected early enough in the infection there will be no antibodies but there will be a greater chance of a PCR positive result. Conversely, when there are detectable antibodies it is less likely that there will be a positive PCR result. It is therefore extremely important that a complete medical history is provided especially the number of days of fever prior to sample collection. More effort is required to develop and validate SFG diagnostics and those of other rickettsial infections.

The histone chaperones Nap1 and Vps75 bind histones H3 and H4 in a tetrameric conformation.

Histone chaperones physically interact with histones to direct proper assembly and disassembly of nucleosomes regulating diverse nuclear processes such as DNA replication, promoter remodeling, transcription elongation, DNA damage, and histone variant exchange. Currently, the best-characterized chaperone-histone interaction is that between the ubiquitous chaperone Asf1 and a dimer of H3 and H4. Nucleosome assembly proteins (Nap proteins) represent a distinct class of histone chaperone. Using pulsed electron double resonance (PELDOR) measurements and protein crosslinking, we show that two members of this class, Nap1 and Vps75, bind histones in the tetrameric conformation also observed when they are sequestered within the nucleosome. Furthermore, H3 and H4 trapped in their tetrameric state can be used as substrates in nucleosome assembly and chaperone-mediated lysine acetylation. This alternate mode of histone interaction provides a potential means of maintaining the integrity of the histone tetramer during cycles of nucleosome reassembly.

DNA repair factor APLF is a histone chaperone.

Poly(ADP-ribosyl)ation plays a major role in DNA repair, where it regulates chromatin relaxation as one of the critical events in the repair process. However, the molecular mechanism by which poly(ADP-ribose) modulates chromatin remains poorly understood. Here we identify the poly(ADP-ribose)-regulated protein APLF as a DNA-damage-specific histone chaperone. APLF preferentially binds to the histone H3/H4 tetramer via its C-terminal acidic motif, which is homologous to the motif conserved in the histone chaperones of the NAP1L family (NAP1L motif). We further demonstrate that APLF exhibits histone chaperone activities in a manner that is dependent on its acidic domain and that the NAP1L motif is critical for the repair capacity of APLF in vivo. Finally, we identify structural analogs of APLF in lower eukaryotes with the ability to bind histones and localize to the sites of DNA-damage-induced poly(ADP-ribosyl)ation. Collectively, these findings define the involvement of histone chaperones in poly(ADP-ribose)-regulated DNA repair reactions.

The Chromatin Remodelling Enzymes SNF2H and SNF2L Position Nucleosomes adjacent to CTCF and Other Transcription Factors.

Within the genomes of metazoans, nucleosomes are highly organised adjacent to the binding sites for a subset of transcription factors. Here we have sought to investigate which chromatin remodelling enzymes are responsible for this. We find that the ATP-dependent chromatin remodelling enzyme SNF2H plays a major role organising arrays of nucleosomes adjacent to the binding sites for the architectural transcription factor CTCF sites and acts to promote CTCF binding. At many other factor binding sites SNF2H and the related enzyme SNF2L contribute to nucleosome organisation. The action of SNF2H at CTCF sites is functionally important as depletion of CTCF or SNF2H affects transcription of a common group of genes. This suggests that chromatin remodelling ATPase's most closely related to the Drosophila ISWI protein contribute to the function of many human gene regulatory elements.