Transcriptional elongation control of hypoxic response.

The release of paused RNA polymerase II (RNAPII) from promoter-proximal regions is tightly controlled to ensure proper regulation of gene expression. The elongation factor PTEF-b is known to release paused RNAPII via phosphorylation of the RNAPII C-terminal domain by its cyclin-dependent kinase component, CDK9. However, the signal and stress-specific roles of the various RNAPII-associated macromolecular complexes containing PTEF-b/CDK9 are not yet clear. Here, we identify and characterize the CDK9 complex required for transcriptional response to hypoxia. Contrary to previous reports, our data indicate that a CDK9 complex containing BRD4 but not AFF1/4 is essential for this hypoxic stress response. We demonstrate that BRD4 bromodomains (BET) are dispensable for the release of paused RNAPII at hypoxia-activated genes and that BET inhibition by JQ1 is insufficient to impair hypoxic gene response. Mechanistically, we demonstrate that the C-terminal region of BRD4 is required for Polymerase-Associated Factor-1 Complex (PAF1C) recruitment to establish an elongation-competent RNAPII complex at hypoxia-responsive genes. PAF1C disruption using a small-molecule inhibitor (iPAF1C) impairs hypoxia-induced, BRD4-mediated RNAPII release. Together, our results provide insight into potentially targetable mechanisms that control the hypoxia-responsive transcriptional elongation.

Ether lipids influence cancer cell fate by modulating iron uptake.

Cancer cell fate has been widely ascribed to mutational changes within protein-coding genes associated with tumor suppressors and oncogenes. In contrast, the mechanisms through which the biophysical properties of membrane lipids influence cancer cell survival, dedifferentiation and metastasis have received little scrutiny. Here, we report that cancer cells endowed with a high metastatic ability and cancer stem cell-like traits employ ether lipids to maintain low membrane tension and high membrane fluidity. Using genetic approaches and lipid reconstitution assays, we show that these ether lipid-regulated biophysical properties permit non-clathrin-mediated iron endocytosis via CD44, leading directly to significant increases in intracellular redox-active iron and enhanced ferroptosis susceptibility. Using a combination of in vitro three-dimensional microvascular network systems and in vivo animal models, we show that loss of ether lipids also strongly attenuates extravasation, metastatic burden and cancer stemness. These findings illuminate a mechanism whereby ether lipids in carcinoma cells serve as key regulators of malignant progression while conferring a unique vulnerability that can be exploited for therapeutic intervention.

Therapeutic targeting of BET bromodomain and other epigenetic acetylrecognition domain-containing factors.

Development of cancer therapies targeting chromatin modifiers and transcriptional regulatory factors is rapidly expanding to include new targets and novel targeting strategies. At the same time, basic molecular research continues to refine our understanding of the epigenetic mechanisms regulating transcription, gene expression, and oncogenesis. This mini-review focuses on cancer therapies targeting the chromatin-associated factors that recognize histone lysine acetylation. Recently reported safety and efficacy are discussed for inhibitors targeting the bromodomains of bromodomain and extraterminal domain (BET) family proteins. In light of recent results indicating that the transcriptional regulator BRD4-PTEFb can function independently of BRD4's bromodomains, the clinical trial performance of these BET inhibitors is placed in a broader context of existing and potential strategies for targeting BRD4-PTEFb. Recently developed therapies targeting bromodomain-containing factors within the SWI/SNF (BAF) family of chromatin remodeling complexes are discussed, as is the potential for targeting the bromodomain-containing transcription factor TAF1 and the YEATS acetylrecognition domain-containing factor GAS41. Recent findings regarding the selectivity and combinatorial specificity of acetylrecognition are highlighted. In conclusion, the potential for further development is discussed with a focus on proximity-based therapies targeting this class of epigenetic factors.

Emergency Contraception for Patients Taking Isotretinoin.

This Viewpoint explains the need for emergency contraception as the last chance to avoid pregnancy for patients taking isotretinoin.

Lived Experience of Acne and Acne Treatment in Transgender Patients.

Importance: While acne is common in transgender and gender-diverse people and is associated with gender-affirming hormone therapy, little research has examined these factors and their impact in gender minority groups. Objective: To examine the lived experiences of acne and acne treatment in transgender and gender-diverse participants. Design, Setting, and Participants: This mixed-methods analysis was conducted at a multidisciplinary gender center at a public safety-net hospital and endocrinology and dermatology clinics at a tertiary academic center from January 4, 2021, to April 7, 2022, using semistructured interviews and surveys. Participants were transgender or gender-diverse adults who had received gender-affirming hormone therapy. Data analysis was performed from November 11, 2021, to March 31, 2023. Exposure: Current diagnosis of acne. Main Outcomes and Measures: Interviews exploring the experience of acne and acne treatment were audio recorded, transcribed verbatim, coded using minority stress theory and the socioecological model, and analyzed using thematic analysis. Themes were triangulated with survey data on gender, self-reported acne severity, skin-specific quality-of-life impact, and treatment experience. Results: A total of 32 participants were included in the study (mean [range] age, 32 [18-57] years; 17 transgender men, 11 transgender women, and 4 nonbinary participants). Ten participants (31%) self-rated their skin as currently clear or almost clear, 11 (34%) reported mild acne, and 11 (34%) had moderate to severe acne. Participants described experiences of rejection and bullying related to acne and admitted avoiding social interactions in which they anticipated acne-related discrimination, which led to feelings of depression and anxiety. Acne worsened body appearance dissatisfaction. Transgender women reported acne interfering with feminine gender expression. Transgender men often normalized acne development, sometimes viewing acne positively as an early sign of testosterone action. Most participants tried over-the-counter acne treatments and commonly sought acne treatment advice from physicians, peers, online forums, and social media. Barriers to acne treatments included cost, lack of multidisciplinary care, mistrust toward the health care system, and lack of transgender-specific acne care education. Conclusions and Relevance: In this mixed-methods study, transgender and gender-diverse individuals reported experiencing acne-related stigma and facing barriers to acne treatment. Multilevel changes, such as developing strategies to reduce acne stigma, providing transgender-specific acne care education, facilitating multidisciplinary acne care, and expanding transgender-friendly clinical environments, are recommended to reduce the impact of acne in transgender individuals.

Somatic mutations of MLL4/COMPASS induce cytoplasmic localization providing molecular insight into cancer prognosis and treatment.

Cancer genome sequencing consortiums have recently catalogued an abundance of somatic mutations, across a wide range of human cancers, in the chromatin-modifying enzymes that regulate gene expression. Defining the molecular mechanisms underlying the potentially oncogenic functions of these epigenetic mutations could serve as the basis for precision medicine approaches to cancer therapy. MLL4 encoded by the KMT2D gene highly mutated in a large number of human cancers, is a key histone lysine monomethyltransferase within the Complex of Proteins Associated with Set1 (COMPASS) family that regulates gene expression through enhancer function, potentially functioning as a tumor suppressor. We report that the KMT2D mutations which cause MLL4 protein truncation also alter MLL4's subcellular localization, resulting in loss-of-function in the nucleus and gain-of-function in the cytoplasm. We demonstrate that isogenic correction of KMT2D truncation mutation rescues the aberrant localization phenotype and restores multiple regulatory functions of MLL4, including COMPASS integrity/stabilization, histone H3K4 mono-methylation, enhancer activation, and therefore transcriptional regulation. Moreover, isogenic correction diminishes the sensitivity of KMT2D-mutated cancer cells to targeted metabolic inhibition. Using immunohistochemistry, we identified that cytoplasmic MLL4 is unique to the tissue of bladder cancer patients with KMT2D truncation mutations. Using a preclinical carcinogen model of bladder cancer in mouse, we demonstrate that truncated cytoplasmic MLL4 predicts response to targeted metabolic inhibition therapy for bladder cancer and could be developed as a biomarker for KMT2D-mutated cancers. We also highlight the broader potential for prognosis, patient stratification and treatment decision-making based on KMT2D mutation status in MLL4 truncation-relevant diseases, including human cancers and Kabuki Syndrome.

Dermatologic care of patients with differences of sex development.

Background: Differences of sex development (DSD or disorders of sex development) are uncommon congenital conditions, characterized by atypical development of chromosomal, gonadal, or anatomic sex. Objective: Dermatologic care is an important component of the multidisciplinary care needed for individuals with DSD. This article discusses the most common primary dermatologic manifestations of DSD in addition to the cutaneous manifestations of hormonal and surgical therapies in individuals with DSD. Data sources: Published articles including case series and case reports on PubMed. Study selections: Selection was conducted by examining existing literature with a team of multidisciplinary specialists. Methods: Narrative review. Limitations: This article was not conducted as a systematic review. Results: In Klinefelter syndrome, refractory leg ulcers and incontinentia pigmenti have been described. Turner syndrome is associated with lymphatic malformations, halo nevi, dermatitis, and psoriasis. Virilization can be seen in some forms of congenital adrenal hyperplasia, where acne and hirsutism are common. Conclusion: Dermatologists should consider teratogenic risk for treatments of skin conditions in DSD depending on pregnancy potential. Testosterone replacement, commonly used for Klinefelter syndrome, androgen insensitivity syndrome, 5-alpha reductase deficiency, gonadal dysgenesis, or ovotesticular DSD, may cause acne.

Distinct layers of BRD4-PTEFb reveal bromodomain-independent function in transcriptional regulation.

The BET family protein BRD4, which forms the CDK9-containing BRD4-PTEFb complex, is considered to be a master regulator of RNA polymerase II (Pol II) pause release. Because its tandem bromodomains interact with acetylated histone lysine residues, it has long been thought that BRD4 requires these bromodomains for its recruitment to chromatin and transcriptional regulatory function. Here, using rapid depletion and genetic complementation with domain deletion mutants, we demonstrate that BRD4 bromodomains are dispensable for Pol II pause release. A minimal, bromodomain-less C-terminal BRD4 fragment containing the PTEFb-interacting C-terminal motif (CTM) is instead both necessary and sufficient to mediate Pol II pause release in the absence of full-length BRD4. Although BRD4-PTEFb can associate with chromatin through acetyl recognition, our results indicate that a distinct, active BRD4-PTEFb population functions to regulate transcription independently of bromodomain-mediated chromatin association. These findings may enable more effective pharmaceutical modulation of BRD4-PTEFb activity.

Disability workforce and the NDIS planning process in regional, rural and remote regions of Australia: Scoping review.

BACKGROUND: The Australian geographically rural and remote disability workforce has historically demonstrated difficulties to keep up with the demand for quality services and supports for people with disability. In 2013, the National Disability Insurance Scheme (NDIS) was launched to provide individualised disability support packages to meet people's needs. To receive funding, people with disability are required to develop a NDIS plan. That plan is then funded by the National Disability Insurance Agency (NDIA), the government agency responsible for managing the NDIS. Although the NDIS has been operating for almost 10 years, there is limited research into the planning experiences of the workforce in regional, rural and remote regions of Australia. This review aims to ascertain the level of scholarly investigation into workers' experiences of NDIS planning. METHODOLOGY: Research publication databases were searched using a specific search string to identify publications that included reference to the workforce's experiences of the NDIS planning process in regional, rural and remote regions of Australia. The Mixed Methods Appraisal Tool (MMAT) was adopted to appraise the quality of the research publications. Research publications that focused on those working with Aboriginal and Torres Strait Islander people were also appraised using the Aboriginal and Torres Strait Islander Quality Appraisal Tool developed by the Centre for Excellence in Aboriginal Chronic Disease Knowledge Translation and Exchange. A thematic synthesis of the publications was undertaken to ascertain disability and health workforce experiences of the NDIS planning process. RESULTS: Seven papers met the selection criteria. Two papers were policy reviews and reported the improvements of the NDIS planning process since its inception. These studies reported four reoccurring themes: (1) cultural/socioeconomic and geographical factors; (2) administrative burden and bureaucracy; (3) values, culture and geography; and (4) burden on allied health workers. CONCLUSION: The NDIS planning process has developed and progressed since its rollout in 2013. There are limited research papers available that describe the workforce's experience of the planning process in regional, rural and remote regions. More research in this area is needed to identify the experiences of the disability workforce in relation to the NDIS planning process.

The experiences of people with disability and their families/carers navigating the NDIS planning process in regional, rural and remote regions of Australia: Scoping review.

BACKGROUND: Australia's National Disability Insurance Scheme (NDIS) was launched in 2013 to provide financial support packages for people with disability to purchase supports and services to enhance independence. People with disability are required to develop a plan with the National Disability Insurance Agency (NDIA), the government department responsible for managing the NDIS. This scoping review aims to ascertain the level of research into people's experience of the NDIS planning process in these geographic areas. METHODOLOGY: Research publication databases were searched using a specific search string to identify research about people with disability and their families/carer's experiences of the NDIS planning process in regional, rural and remote regions of Australia. The Mixed Methods Appraisal Tool (MMAT) was adopted to appraise the quality of the research publications. Research publications focused on Aboriginal and Torres Strait Islander people were additionally appraised using the Aboriginal and Torres Strait Islander Quality Appraisal Tool developed by the Centre for Excellence in Aboriginal Chronic Disease Knowledge Translation and Exchange. A thematic synthesis of the publications' contents was undertaken to ascertain people with disabilities and carers experience of the NDIS planning process. RESULTS: Ten (N = 10) research papers were found that met the inclusion criteria. Two papers were policy reviews and reported on the improvements of the NDIS planning process since its conception. The analysis found the research archive focused on five themes: (1) healthcare workforce and NDIA staff; (2) NDIS package holders and carers lack of awareness of the NDIS; (3) cultural/socio-economic barriers; (4) travel funding; and (5) emotional burden of the NDIS planning process. CONCLUSION: There are limited papers available that explore people's experiences of the NDIS planning process in regional, rural and remote regions of Australia. This systematic review illuminates the difficulties, barriers and concerns of people with disability and their carers about the planning process.

Adherence to Gender Affirming Hormone Therapy in Transgender Adolescents and Adults: A Retrospective Cohort Study.

CONTEXT: Transgender and gender diverse (TGD) individuals often seek gender-affirming hormone therapy (GAHT). While receipt of GAHT has been associated with improved well-being, the risk of GAHT discontinuation and its reasons are not well known. OBJECTIVE: There were two main objectives: (1) To investigate the proportion of TGD individuals who discontinue therapy after an average of 4 years (maximum 19 years) since GAHT initiation; and (2) to explore reasons for GAHT discontinuation. This was a retrospective cohort study at academic centers providing care to TGD adolescents and adults. TGD individuals prescribed estradiol or testosterone between January 1, 2000, and January 1, 2019, were included. GAHT continuation was ascertained using a 2-phase process. In phase 1, Kaplan-Meier survival analyses were used to examine likelihood of GAHT discontinuation and compare discontinuation rates by age and sex assigned at birth. In phase 2, reasons for stopping GAHT were investigated by reviewing records and by contacting study participants who discontinued therapy. The main outcome measures were incidence and determinants of GAHT discontinuation. RESULTS: Among 385 eligible participants, 231 (60%) were assigned male at birth and 154 (40%) were assigned female at birth. Less than one-third of participants (n = 121) initiated GAHT prior to their 18th birthday, constituting the pediatric cohort (mean age 15 years), and the remaining 264 were included in the adult cohort (mean age 32 years). In phase 1, 6 participants (1.6%) discontinued GAHT during follow-up, and of those only 2 discontinued GAHT permanently (phase 2). CONCLUSION: GAHT discontinuation is uncommon when therapy follows Endocrine Society guidelines. Future research should include prospective studies with long-term follow-up of individuals receiving GAHT.

Therapeutic targeting of metabolic vulnerabilities in cancers with MLL3/4-COMPASS epigenetic regulator mutations.

Epigenetic status-altering mutations in chromatin-modifying enzymes are a feature of human diseases, including many cancers. However, the functional outcomes and cellular dependencies arising from these mutations remain unresolved. In this study, we investigated cellular dependencies, or vulnerabilities, that arise when enhancer function is compromised by loss of the frequently mutated COMPASS family members MLL3 and MLL4. CRISPR dropout screens in MLL3/4-depleted mouse embryonic stem cells (mESCs) revealed synthetic lethality upon suppression of purine and pyrimidine nucleotide synthesis pathways. Consistently, we observed a shift in metabolic activity toward increased purine synthesis in MLL3/4-KO mESCs. These cells also exhibited enhanced sensitivity to the purine synthesis inhibitor lometrexol, which induced a unique gene expression signature. RNA-Seq identified the top MLL3/4 target genes coinciding with suppression of purine metabolism, and tandem mass tag proteomic profiling further confirmed upregulation of purine synthesis in MLL3/4-KO cells. Mechanistically, we demonstrated that compensation by MLL1/COMPASS was underlying these effects. Finally, we demonstrated that tumors with MLL3 and/or MLL4 mutations were highly sensitive to lometrexol in vitro and in vivo, both in culture and in animal models of cancer. Our results depicted a targetable metabolic dependency arising from epigenetic factor deficiency, providing molecular insight to inform therapy for cancers with epigenetic alterations secondary to MLL3/4 COMPASS dysfunction.

Re-thinking Stepfathers' Contributions: Fathers, Stepfathers, and Child Wellbeing.

Using data from a contemporary cohort of children, we revisit the question of whether children benefit from being close to and engaging in activities with a stepfather. We deploy the Fragile Families and Child Wellbeing Study, a birth cohort study of nearly 5000 children born in US cities in 1998-2000, with a large oversample of nonmarital births. We explore the relationships between stepfathers' closeness and active engagement and youth's internalizing and externalizing behaviors and school connectedness at ages 9 and 15 for between 550 and 740 children (depending on the wave) with stepfathers. We find that the emotional tenor of the relationship and level of active engagement between youth and their stepfathers are associated with reduced internalizing behaviors and higher school connectedness. Our findings suggest that stepfathers' roles seem to have evolved in ways that are more beneficial to their adolescent stepchildren than was previously the case.

Incidence and Factors Associated With Acne in Transgender Adolescents on Testosterone: A Retrospective Cohort Study.

OBJECTIVE: This retrospective cohort study aimed to assess incidence and predictors of acne among transgender adolescents receiving testosterone. METHODS: We analyzed records of patients aged <18 years, assigned female at birth, seen at Children's Healthcare of Atlanta Pediatric Endocrinology clinic for testosterone initiation between January 1, 2016, and January 1, 2019, with at least 1-year follow-up documented. Bivariable analyses to determine the association of clinical and demographic factors with new acne diagnosis were performed. RESULTS: Of 60 patients, 46 (77%) did not have baseline acne, but of those 46 patients, 25 (54%) developed acne within 1 year of testosterone initiation. Overall incidence proportion was 70% at 2 years; patients who used progestin prior to or during follow-up were more likely to develop acne than nonusers (92% vs 33%, P <.001). CONCLUSION: Transgender adolescents starting testosterone, particularly those taking progestin, should be monitored for acne development and treated proactively by hormone providers and dermatologists.

Identification of an immunogenic epitope and protective antibody against the furin cleavage site of SARS-CoV-2.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the global coronavirus disease 2019 (COVID-19) pandemic, contains a unique, four amino acid (aa) "PRRA" insertion in the spike (S) protein that creates a transmembrane protease serine 2 (TMPRSS2)/furin cleavage site and enhances viral infectivity. More research into immunogenic epitopes and protective antibodies against this SARS-CoV-2 furin cleavage site is needed. METHODS: Combining computational and experimental methods, we identified and characterized an immunogenic epitope overlapping the furin cleavage site that detects antibodies in COVID-19 patients and elicits strong antibody responses in immunized mice. We also identified a high-affinity monoclonal antibody from COVID-19 patient peripheral blood mononuclear cells; the antibody directly binds the furin cleavage site and protects against SARS-CoV-2 infection in a mouse model. FINDINGS: The presence of "PRRA" amino acids in the S protein of SARS-CoV-2 not only creates a furin cleavage site but also generates an immunogenic epitope that elicits an antibody response in COVID-19 patients. An antibody against this epitope protected against SARS-CoV-2 infection in mice. INTERPRETATION: The immunogenic epitope and protective antibody we have identified may augment our strategy in handling COVID-19 epidemic. FUNDING: The National Natural Science Foundation of China (82102371, 91542201, 81925025, 82073181, and 81802870), the Chinese Academy of Medical Sciences Initiative for Innovative Medicine (2021-I2M-1-047 and 2022-I2M-2-004), the Non-profit Central Research Institute Fund of the Chinese Academy of Medical Sciences (2020-PT310-006, 2019XK310002, and 2018TX31001), the National Key Research and Development Project of China (2020YFC0841700), US National Institute of Health (NIH) funds grant AI158154, University of California Los Angeles (UCLA) AI and Charity Treks, and UCLA DGSOM BSCRC COVID-19 Award Program. H.Y. is supported by Natural Science Foundation of Jiangsu Province (BK20211554 andBE2022728).

Managing Dermatologic Effects of Gender-Affirming Therapy in Transgender Adolescents.

Transgender adolescents seek gender-affirming medical care to address gender identity and incongruence. Improved understanding of the dermatologic impact of gender-affirming medical care such as pubertal suppression, hormone therapy, and surgeries can enhance patient outcome. Pubertal suppression treats dysphoria associated with development of secondary sex characteristics, including androgen-driven acne. Gender-affirming hormone therapy influences acne and hair development in transgender adolescents. Dermatologists can help manage skin effects associated with chest binding and gender-affirming hormone therapy and surgery. Provision of patient-centered gender-affirming care in dermatologic and multidisciplinary settings is essential to improve skin and overall outcomes of gender-affirming therapy.

Stepping back: Impact of pass-fail Step 1 scoring on dermatology applicants.

The United States Medical Licensing Examination (USMLE) Step 1 recently transitioned to a pass-fail system from their three-digit scoring system. The implications of this change will ultimately influence medical students, residency programs, and, most importantly, patients. This article seeks to address some of the varying ethical and practical effects of this conversion to a pass/fail examination.

Membrane tension propagation couples axon growth and collateral branching.

Neuronal axons must navigate a mechanically heterogeneous environment to reach their targets, but the biophysical mechanisms coupling mechanosensation, growth, and branching are not fully understood. Here, we show that local changes in membrane tension propagate along axons at approximately 20 µm/s, more than 1000-fold faster than in most other nonmotile cells where this property has been measured. Local perturbations to tension decay along the axon with a length constant of approximately 41 µm. This rapid and long-range mechanical signaling mediates bidirectional competition between axonal branch initiation and growth cone extension. Our data suggest a mechanism by which mechanical cues at one part of a growing axon can affect growth dynamics remotely.

The RNA pseudoknots in foot-and-mouth disease virus are dispensable for genome replication, but essential for the production of infectious virus.

Non-coding regions of viral RNA (vRNA) genomes are critically important in the regulation of gene expression. In particular, pseudoknot (PK) structures, which are present in a wide range of RNA molecules, have a variety of roles. The 5' untranslated region (5' UTR) of foot-and-mouth disease virus (FMDV) vRNA is considerably longer than in other viruses from the picornavirus family and consists of a number of distinctive structural motifs that includes multiple (2, 3 or 4 depending on the virus strain) putative PKs linked in tandem. The role(s) of the PKs in the FMDV infection are not fully understood. Here, using bioinformatics, sub-genomic replicons and recombinant viruses we have investigated the structural conservation and importance of the PKs in the FMDV lifecycle. Our results show that despite the conservation of two or more PKs across all FMDVs, a replicon lacking PKs was replication competent, albeit at reduced levels. Furthermore, in competition experiments, GFP FMDV replicons with less than two (0 or 1) PK structures were outcompeted by a mCherry FMDV wt replicon that had 4 PKs, whereas GFP replicons with 2 or 4 PKs were not. This apparent replicative advantage offered by the additional PKs correlates with the maintenance of at least two PKs in the genomes of FMDV field isolates. Despite a replicon lacking any PKs retaining the ability to replicate, viruses completely lacking PK were not viable and at least one PK was essential for recovery of infections virus, suggesting a role for the PKs in virion assembly. Thus, our study points to roles for the PKs in both vRNA replication and virion assembly, thereby improving understanding the molecular biology of FMDV replication and the wider roles of PK in RNA functions.