A change in biophysical properties accompanies heterochromatin formation in mouse embryos.

The majority of our genome is composed of repeated DNA sequences that assemble into heterochromatin, a highly compacted structure that constrains their mutational potential. How heterochromatin forms during development and how its structure is maintained are not fully understood. Here, we show that mouse heterochromatin phase-separates after fertilization, during the earliest stages of mammalian embryogenesis. Using high-resolution quantitative imaging and molecular biology approaches, we show that pericentromeric heterochromatin displays properties consistent with a liquid-like state at the two-cell stage, which change at the four-cell stage, when chromocenters mature and heterochromatin becomes silent. Disrupting the condensates results in altered transcript levels of pericentromeric heterochromatin, suggesting a functional role for phase separation in heterochromatin function. Thus, our work shows that mouse heterochromatin forms membrane-less compartments with biophysical properties that change during development and provides new insights into the self-organization of chromatin domains during mammalian embryogenesis.

Histone post-translational modifications - cause and consequence of genome function.

Much has been learned since the early 1960s about histone post-translational modifications (PTMs) and how they affect DNA-templated processes at the molecular level. This understanding has been bolstered in the past decade by the identification of new types of histone PTM, the advent of new genome-wide mapping approaches and methods to deposit or remove PTMs in a locally and temporally controlled manner. Now, with the availability of vast amounts of data across various biological systems, the functional role of PTMs in important processes (such as transcription, recombination, replication, DNA repair and the modulation of genomic architecture) is slowly emerging. This Review explores the contribution of histone PTMs to the regulation of genome function by discussing when these modifications play a causative (or instructive) role in DNA-templated processes and when they are deposited as a consequence of such processes, to reinforce and record the event. Important advances in the field showing that histone PTMs can exert both direct and indirect effects on genome function are also presented.

Chromatin dynamics in the regulation of cell fate allocation during early embryogenesis.

Following fertilization, gametes undergo epigenetic reprogramming in order to revert to a totipotent state. How embryonic cells subsequently acquire their fate and the role of chromatin dynamics in this process are unknown. Genetic and experimental embryology approaches have identified some of the players and morphological changes that are involved in early mammalian development, but the exact events underlying cell fate allocation in single embryonic cells have remained elusive. Experimental and technological advances have recently provided novel insights into chromatin dynamics and nuclear architecture in single cells; these insights have reshaped our understanding of the mechanisms underlying cell fate allocation and plasticity in early mammalian development.

SUV4-20 activity in the preimplantation mouse embryo controls timely replication.

Extensive chromatin remodeling after fertilization is thought to take place to allow a new developmental program to start. This includes dynamic changes in histone methylation and, in particular, the remodeling of constitutive heterochromatic marks such as histone H4 Lys20 trimethylation (H4K20me3). While the essential function of H4K20me1 in preimplantation mouse embryos is well established, the role of the additional H4K20 methylation states through the action of the SUV4-20 methyltransferases has not been addressed. Here we show that Suv4-20h1/h2 are mostly absent in mouse embryos before implantation, underscoring a rapid decrease of H4K20me3 from the two-cell stage onward. We addressed the functional significance of this remodeling by introducing Suv4-20h1 and Suv4-20h2 in early embryos. Ectopic expression of Suv4-20h2 leads to sustained levels of H4K20me3, developmental arrest, and defects in S-phase progression. The developmental phenotype can be partially overcome through inhibition of the ATR pathway, suggesting that the main function for the remodeling of H4K20me3 after fertilization is to allow the timely and coordinated progression of replication. This is in contrast to the replication program in somatic cells, where H4K20me3 has been shown to promote replication origin licensing, and anticipates a different regulation of replication during this early developmental time window.

Center-level and region-level variations in liver transplantation practices following acuity circles policy change.

Although early studies suggest the Acuity Circles (AC) allocation policy has increased access to deceased donor liver transplants (DDLTs) for patients with the highest MELD scores, changes in center- and region-level practices among patients with the highest MELD scores in response to AC are not well-characterized. OPTN/UNOS data were analyzed to compare center-level changes in the number of DDLTs based on allocation-MELD (aMELD) categories used for AC sharing performed in the 18-month periods before and after AC enactment on February 4, 2020. There was large center-level variation in the number and proportion of aMELD ≥ 37 DDLTs performed from pre-AC to AC period; 13 centers accounted for 196 of the 198 total net increase in aMELD ≥ 37 DDLTs performed after AC, 5 of these being from UNOS region 5. Similar center-level variation was seen for MELD 33-36 and MELD 29-32 DDLTs, with 17 centers and 14 centers, respectively, accounting for the entire net increase in DDLTs in the aMELD categories. In conclusion, AC increased access to livers for transplantation for high MELD patients nationally, but imbalances remain in transplant practice patterns at the center and regional levels. Longer-term study is necessary to assess effectiveness of AC in improving equitability of liver transplantations.

The role of ZFP57 and additional KRAB-zinc finger proteins in the maintenance of human imprinted methylation and multi-locus imprinting disturbances.

Genomic imprinting is an epigenetic process regulated by germline-derived DNA methylation that is resistant to embryonic reprogramming, resulting in parental origin-specific monoallelic gene expression. A subset of individuals affected by imprinting disorders (IDs) displays multi-locus imprinting disturbances (MLID), which may result from aberrant establishment of imprinted differentially methylated regions (DMRs) in gametes or their maintenance in early embryogenesis. Here we investigated the extent of MLID in a family harbouring a ZFP57 truncating variant and characterize the interactions between human ZFP57 and the KAP1 co-repressor complex. By ectopically targeting ZFP57 to reprogrammed loci in mouse embryos using a dCas9 approach, we confirm that ZFP57 recruitment is sufficient to protect oocyte-derived methylation from reprogramming. Expression profiling in human pre-implantation embryos and oocytes reveals that unlike in mice, ZFP57 is only expressed following embryonic-genome activation, implying that other KRAB-zinc finger proteins (KZNFs) recruit KAP1 prior to blastocyst formation. Furthermore, we uncover ZNF202 and ZNF445 as additional KZNFs likely to recruit KAP1 to imprinted loci during reprogramming in the absence of ZFP57. Together, these data confirm the perplexing link between KZFPs and imprint maintenance and highlight the differences between mouse and humans in this respect.

SCUBA Diver's Knee: A Case Report.

ABSTRACT: With more than 9 million recreational certified self-contained underwater breathing apparatus divers in the United States, clinicians should be aware of the unique diving-related injuries. One of the most common diving-related injuries is type 1 decompression sickness, or "the bends." The bends commonly manifest as localized joint pain, most often occurring within 24 h of surfacing and resolving over the following 1 to 2 d. We report a unique case of a patient who experienced an exacerbation of musculoskeletal joint pain following initial recovery. This 35-year-old man had nearly complete resolution of his joint pain following the bends, then developed severe right knee pain with swelling after a high-volume lower body workout. Following unremarkable imaging and unsuccessful conservative treatment, ultrasound-guided aspiration of his right knee was performed, which resolved the patient's symptoms. This case highlights a unique presentation of the bends and demonstrates a potentially beneficial treatment if recurrence of the bends is suspected.

Expression and phase separation potential of heterochromatin proteins during early mouse development.

In most eukaryotes, constitutive heterochromatin is associated with H3K9me3 and HP1α. The latter has been shown to play a role in heterochromatin formation through liquid-liquid phase separation. However, many other proteins are known to regulate and/or interact with constitutive heterochromatic regions in several species. We postulate that some of these heterochromatic proteins may play a role in the regulation of heterochromatin formation by liquid-liquid phase separation. Indeed, an analysis of the constitutive heterochromatin proteome shows that proteins associated with constitutive heterochromatin are significantly more disordered than a random set or a full nucleome set of proteins. Interestingly, their expression begins low and increases during preimplantation development. These observations suggest that the preimplantation embryo is a useful model to address the potential role for phase separation in heterochromatin formation, anticipating exciting research in the years to come.

The role of PR-Set7 in replication licensing depends on Suv4-20h.

PR-Set7 is the sole monomethyltransferase responsible for H4K20 monomethylation (H4K20me1) that is the substrate for further methylation by Suv4-20h1/h2. PR-Set7 is required for proper cell cycle progression and is subject to degradation by the CRL4(Cdt2) ubiquitin ligase complex as a function of the cell cycle and DNA damage. This report demonstrates that PR-Set7 is an important downstream effector of CRL4(Cdt2) function during origin of DNA replication licensing, dependent on Suv4-20h1/2 activity. Aberrant rereplication correlates with decreased levels of H4K20me1 and increased levels of H4K20 trimethylation (H4K20me3). Expression of a degradation-resistant PR-Set7 mutant in the mouse embryo that is normally devoid of Suv4-20 does not compromise development or cell cycle progression unless Suv4-20h is coexpressed. PR-Set7 targeting to an artificial locus results in recruitment of the origin recognition complex (ORC) in a manner dependent on Suv4-20h and H4K20me3. Consistent with this, H4K20 methylation status plays a direct role in recruiting ORC through the binding properties of ORC1 and ORCA/LRWD1. Thus, coordinating the status of H4K20 methylation is pivotal for the proper selection of DNA replication origins in higher eukaryotes.

Inositol pyrophosphates regulate JMJD2C-dependent histone demethylation.

Epigenetic modifications of chromatin represent a fundamental mechanism by which eukaryotic cells adapt their transcriptional response to developmental and environmental cues. Although an increasing number of molecules have been linked to epigenetic changes, the intracellular pathways that lead to their activation/repression have just begun to be characterized. Here, we demonstrate that inositol hexakisphosphate kinase 1 (IP6K1), the enzyme responsible for the synthesis of the high-energy inositol pyrophosphates (IP7), is associated with chromatin and interacts with Jumonji domain containing 2C (JMJD2C), a recently identified histone lysine demethylase. Reducing IP6K1 levels by RNAi or using mouse embryonic fibroblasts derived from ip6k1(-/-) knockout mice results in a decreased IP7 concentration that epigenetically translates to reduced levels of trimethyl-histone H3 lysine 9 (H3K9me3) and increased levels of acetyl-H3K9. Conversely, expression of IP6K1 induces JMJD2C dissociation from chromatin and increases H3K9me3 levels, which depend on IP6K1 catalytic activity. Importantly, these effects lead to changes in JMJD2C-target gene transcription. Our findings demonstrate that inositol pyrophosphate signaling influences nuclear functions by regulating histone modifications.

Brief exposure to aversive stimuli impairs visual selective attention.

Although it is well established that stress can disrupt complex cognitive functions, relatively little is known about how it influences visual processing, especially in terms of visual selective attention. In the current study, we used highly aversive images, taken from the International Affective Picture System, to induce acute, low-intensity stress while participants performed a visual discrimination task. Consistent with prior research, we found that anticipation of aversive stimuli increased overall amplitude of the N170, suggesting an increase in early sensory gain. More importantly, we found that stress disrupted visual selective attention. While in no-stress blocks, the amplitude of the face-sensitive N170 was higher when participants attended to faces rather than scenes in face-scene overlay images; this effect was absent under stress. This was because of an increase in N170 amplitude in the scene-attend condition under stress. We interpret these findings as suggesting that even low-intensity acute stress can impair participants' ability to filter out task-irrelevant information. We discuss our findings in relation to how even brief exposure to low-intensity stress may adversely impact both healthy and clinical populations.

Heterochromatin protein ERH represses alternative cell fates during early mammalian differentiation.

During development, H3K9me3 heterochromatin is dynamically rearranged, silencing repeat elements and protein coding genes to restrict cell identity. Enhancer of Rudimentary Homolog (ERH) is an evolutionarily conserved protein originally characterized in fission yeast and recently shown to be required for H3K9me3 maintenance in human fibroblasts, but its function during development remains unknown. Here, we show that ERH is required for proper segregation of the inner cell mass and trophectoderm cell lineages during mouse development by repressing totipotent and alternative lineage programs. During human embryonic stem cell (hESC) differentiation into germ layer lineages, ERH is crucial for silencing naïve and pluripotency genes, transposable elements, and alternative lineage genes. Strikingly, ERH depletion in somatic cells reverts the H3K9me3 landscape to an hESC state and enables naïve and pluripotency gene and transposable element activation during iPSC reprogramming. Our findings reveal a role for ERH in initiation and maintenance of developmentally established gene repression.

Inositol pyrophosphate mediated pyrophosphorylation of AP3B1 regulates HIV-1 Gag release.

High-energy inositol pyrophosphates, such as IP(7) (diphosphoinositol pentakisphosphate), can directly donate a beta-phosphate to a prephosphorylated serine residue generating pyrophosphorylated proteins. Here, we show that the beta subunit of AP-3, a clathrin-associated protein complex required for HIV-1 release, is a target of IP(7)-mediated pyrophosphorylation. We have identified Kif3A, a motor protein of the kinesin superfamily, as an AP3B1-binding partner and demonstrate that Kif3A, like the AP-3 complex, is involved in an intracellular process required for HIV-1 Gag release. Importantly, IP(7)-mediated pyrophosphorylation of AP3B1 modulates the interaction with Kif3A and, as a consequence, affects the release of HIV-1 virus-like particles. This study identifies a cellular process that is regulated by IP(7)-mediated pyrophosphorylation.

The Diagnosis and Management of Cardiometabolic Risk and Cardiometabolic Syndrome after Spinal Cord Injury.

Individuals with spinal cord injuries (SCI) commonly present with component risk factors for cardiometabolic risk and combined risk factors for cardiometabolic syndrome (CMS). These primary risk factors include obesity, dyslipidemia, dysglycemia/insulin resistance, and hypertension. Commonly referred to as "silent killers", cardiometabolic risk and CMS increase the threat of cardiovascular disease, a leading cause of death after SCI. This narrative review will examine current data and the etiopathogenesis of cardiometabolic risk, CMS, and cardiovascular disease associated with SCI, focusing on pivotal research on cardiometabolic sequelae from the last five years. The review will also provide current diagnosis and surveillance criteria for cardiometabolic disorders after SCI, a novel obesity classification system based on percent total body fat, and lifestyle management strategies to improve cardiometabolic health.

Demographics of Surgical Specialty Residency Program Directors in the United States: A Cross-sectional Analysis.

OBJECTIVE: To evaluate program director (PD) demographics, training background, and academic productivity in 11 surgical specialties. SUMMARY BACKGROUND DATA: There is currently no comprehensive study comparing educational background, research output, and gender differences between PDs of surgical residencies in the United States. METHODS: The Accreditation Council for Graduate Medical Education (ACGME) and the Association of American Medical Colleges (AAMC) websites were used to identify residency PDs. Age, information related to service as PD, educational background, and research output were collected utilizing online searches including Doximity, PubMed, and Scopus.. The ACGME Data Resource Book was used to obtain data on the gender makeup of residents in each surgical specialty. Data collection occurred between December 14, 2019, and May 9, 2020. RESULTS: 1571 residency PDs across 11 surgical specialties were included. Significant differences between specialties were found with respect to PD gender, current age, age at appointment, years between residency and assignment, term duration, number of PubMed publications, and Scopus h-index. The current age (mean ± SD) ranged from 46.8 ± 8.5 years among Interventional Radiology (IR) PDs to 53.4 ± 9.1 years among Neurological Surgery (NEUROSURG) PDs. The proportion of female PDs ranged from 5.9% in NEUROSURG to 63.5% in Obstetrics and Gynecology (OB-GYN). Completion of a post-residency fellowship was least common for OB-GYN PDs at 9.1%, and most common for IR PDs at 98.8%. The number (mean ± SD) of PubMed publications and Scopus h-index ranged from 13.1 ± 22.3 publications and h index 4.5 ± 5.7 among OB-GYN PDs to 112.5 ± 103.0 publications and h index 27.4 ± 16.7 among Thoracic Surgery PDs. Age and academic productivity as measured by PubMed publications and Scopus h-index were significantly lower among female PDs in multiple surgical specialties. CONCLUSIONS: There were significant variations in the PDs of surgical specialties, particularly with respect to gender and academic productivity. Efforts should be made to support and encourage greater female representation in the role of surgical residency PD.

Age-, sex-, and region-specific differences in skeletal muscle size and quality.

Ultrasonography-derived cross-sectional area (CSA) and echo intensity (EI) are increasingly utilized by investigators to study muscle size and quality, respectively. We sought to examine age, sex, and region (proximal, middle, distal) differences in vastus lateralis and rectus femoris CSA and EI, and determine whether correction for subcutaneous fat thickness influences the magnitude of EI differences. Fifteen younger men (mean age = 23 years), 15 younger women (aged 21 years), 11 older men (aged 74 years), and 15 older women (aged 70 years) participated. Clear differences were observed among age, sex, and region for vastus lateralis CSA (p ≤ 0.013, d = 0.38-0.73), whereas rectus femoris CSA was only different between younger and older participants at the proximal region (p = 0.017, d = 0.65). Uncorrected EI was greatest at the distal region of both muscles (p < 0.001, d = 0.59-1.38), with only the younger men having significantly lower EI values than the other groups (p ≤ 0.043, d = 0.37-0.63). Subcutaneous fat correction resulted in a marked increase in the magnitude of sex-specific EI differences (p ≤ 0.032, d ≥ 0.42). Additionally, subcutaneous fat correction increased the uniformity of EI throughout the thigh. These findings highlight considerable region-specific differences in muscle size and quality among younger and older men and women and highlight the need to correct for subcutaneous fat thickness when examining EI. Novelty Rectus femoris CSA is similar between younger and older adults except at the most proximal site evaluated. Age- and sex-specific differences in uncorrected EI are nonuniform across the thigh. Correction for subcutaneous fat thickness substantially increased EI in women, resulting in greater sex differences.

Muscle strength, not age, explains unique variance in echo intensity.

Echo intensity (EI) is being increasingly utilized by investigators as an index of skeletal muscle quality. Previous studies have reported independent associations between EI versus both age and muscle strength. PURPOSE: We sought to determine whether EI is more strongly associated with age or strength. METHODS: Twenty-eight younger adults (13 men, 15 women; mean age = 22 years) and 25 older adults (10 men, 15 women; age = 71 years) participated. B-mode ultrasonography was utilized to acquire images of the vastus lateralis and rectus femoris. ImageJ software was used to quantify corrected EI and muscle cross-sectional area (CSA). Each participant performed 40 maximal concentric isokinetic muscle actions of the knee extensors (velocity = 180°·s-1). The mean peak torque of the best three attempts was used to quantify muscle strength. Specific torque was calculated as strength relative to CSA. Fatigability was also quantified. Statistical analyses included independent samples t-tests and stepwise regression. RESULTS: There were large differences between age groups for strength (p < .001, d = 1.831) and CSA (p = .003, d = 0.872). When corrected for subcutaneous tissue thickness, the difference in EI between age groups was small (p = .184, d = 0.371). Stepwise regression revealed that muscle strength was the single best predictor of EI (R2 = 0.206), with age, CSA, specific torque, and fatigability explaining no unique variance. CONCLUSION: Concentric isokinetic muscle strength is a better predictor of EI than age.

Heterochromatin establishment during early mammalian development is regulated by pericentromeric RNA and characterized by non-repressive H3K9me3.

Following fertilization in mammals, the gametes are reprogrammed to create a totipotent zygote, a process that involves de novo establishment of chromatin domains. A major feature occurring during preimplantation development is the dramatic remodelling of constitutive heterochromatin, although the functional relevance of this is unknown. Here, we show that heterochromatin establishment relies on the stepwise expression and regulated activity of SUV39H enzymes. Enforcing precocious acquisition of constitutive heterochromatin results in compromised development and epigenetic reprogramming, which demonstrates that heterochromatin remodelling is essential for natural reprogramming at fertilization. We find that de novo H3K9 trimethylation (H3K9me3) in the paternal pronucleus after fertilization is catalysed by SUV39H2 and that pericentromeric RNAs inhibit SUV39H2 activity and reduce H3K9me3. De novo H3K9me3 is initially non-repressive for gene expression, but instead bookmarks promoters for compaction. Overall, we uncover the functional importance for the restricted transmission of constitutive heterochromatin during reprogramming and a non-repressive role for H3K9me3.

Are inositol pyrophosphates signalling molecules?

The inositol polyphosphate family of small, cytosolic molecules has a prominent place in the field of cell signalling, and inositol pyrophosphates are the most recent addition to this large family. First identified in 1993, they have since been found in all eukaryotic organisms studied. The defining feature of inositol pyrophosphates is the presence of the characteristic 'high energy' pyrophosphate group, which immediately attracted interest in them as possible signalling molecules. In addition to their unique 'high energy' pyrophosphate bond, their concentration in the cell is tightly regulated with an extremely rapid turnover. This, together with the history of other inositol polyphosphates, makes it likely that they have an important role in intracellular signalling involving some basic cellular processes. This hypothesis is supported by the surprisingly wide range of cellular functions where inositol pyrophosphates seem to be involved. A seminal finding was that inositol pyrophosphates are able to directly phosphorylate pre-phosphorylated proteins, thereby identifying an entirely new post-translational protein modification, namely serine-pyrophosphorylation. Rapid progress has been made in characterising the metabolism of these molecules in the 15 years since their first identification. However, their detailed signalling role in specific cellular processes and in the context of relevant physiological cues has developed more slowly, particularly in mammalian system. We will discuss inositol pyrophosphates from the cell signalling perspective, analysing how their intracellular concentration is modulated, what their possible molecular mechanisms of action are, together with the physiological consequences of this novel form of signalling.