TRIM5α-Mediated Ubiquitin Chain Conjugation Is Required for Inhibition of HIV-1 Reverse Transcription and Capsid Destabilization.

UNLABELLED: Rhesus macaque TRIM5α (rhTRIM5α) is a retroviral restriction factor that inhibits HIV-1 infection. Previous studies have revealed that TRIM5α restriction occurs via a two-step process. The first step is restriction factor binding, which is sufficient to inhibit infection. The second step, which is sensitive to proteasome inhibition, prevents the accumulation of reverse transcription products in the target cell. However, because of the pleotropic effects of proteasome inhibitors, the molecular mechanisms underlying the individual steps in the restriction process have remained poorly understood. In this study, we have fused the small catalytic domain of herpes simplex virus UL36 deubiquitinase (DUb) to the N-terminal RING domain of rhTRIM5α, which results in a ubiquitination-resistant protein. Cell lines stably expressing this fusion protein inhibited HIV-1 infection to the same degree as a control fusion to a catalytically inactive DUb. However, reverse transcription products were substantially increased in the DUb-TRIM5α fusion relative to the catalytically inactive control or the wild-type (WT) TRIM5α. Similarly, expression of DUb-rhTRIM5α resulted in the accumulation of viral cores in target cells following infection, while the catalytically inactive control and WT rhTRIM5α induced the abortive disassembly of viral cores, indicating a role for ubiquitin conjugation in rhTRIM5α-mediated destabilization of HIV-1 cores. Finally, DUb-rhTRIM5α failed to activate NF-κB signaling pathways compared to controls, demonstrating that this ubiquitination-dependent activity is separable from the ability to restrict retroviral infection. IMPORTANCE: These studies provide direct evidence that ubiquitin conjugation to rhTRIM5α-containing complexes is required for the second step of HIV-1 restriction. They also provide a novel tool by which the biological activities of TRIM family proteins might be dissected to better understand their function and underlying mechanisms of action.

Human ESCRT-I and ALIX function as scaffolding helical filaments in vivo.

The Endosomal Sorting Complex Required for Transport (ESCRT) is an evolutionarily conserved machinery that performs reverse-topology membrane scission in cells universally required from cytokinesis to budding of enveloped viruses. Upstream acting ESCRT-I and ALIX control these events and link recruitment of viral and cellular partners to late-acting ESCRT-III CHMP4 through incompletely understood mechanisms. Using structure-function analyses combined with super-resolution imaging, we show that ESCRT-I and ALIX function as distinct helical filaments in vivo . Together, they are essential for optimal structural scaffolding of HIV-1 nascent virions, the retention of viral and human genomes through defined functional interfaces, and recruitment of CHMP4 that itself assembles into corkscrew-like filaments intertwined with ESCRT-I or ALIX helices. Disruption of filament assembly or their conformationally clustered RNA binding interfaces in human cells impaired membrane abscission, resulted in major structural instability and leaked nucleic acid from nascent virions and nuclear envelopes. Thus, ESCRT-I and ALIX function as helical filaments in vivo and serve as both nucleic acid-dependent structural scaffolds as well as ESCRT-III assembly templates. Significance statement: When cellular membranes are dissolved or breached, ESCRT is rapidly deployed to repair membranes to restore the integrity of intracellular compartments. Membrane sealing is ensured by ESCRT-III filaments assembled on the inner face of membrane; a mechanism termed inverse topology membrane scission. This mechanism, initiated by ESCRT-I and ALIX, is universally necessary for cytokinesis, wound repair, budding of enveloped viruses, and more. We show ESCRT-I and ALIX individually oligomerize into helical filaments that cluster newly discovered nucleic acid-binding interfaces and scaffold-in genomes within nascent virions and nuclear envelopes. These oligomers additionally appear to serve as ideal templates for ESCRT-III polymerization, as helical filaments of CHMP4B were found intertwined ESCRT-I or ALIX filaments in vivo . Similarly, corkscrew-like filaments of ALIX are also interwoven with ESCRT-I, supporting a model of inverse topology membrane scission that is synergistically reinforced by inward double filament scaffolding.

Breastfeeding and Hormonal Contraception: A Scoping Review of Clinical Guidelines, Professional Association Recommendations, and the Literature.

Background: Postpartum contraceptive use can help prevent short-interval pregnancies, which have been associated with adverse neonatal and maternal health outcomes. Many contraceptive methods are safe for postpartum use, but patients and providers may be confused as to what impact hormonal contraception has on lactation. We performed a scoping review of the most recent U.S.-based guidelines regarding hormonal contraception on lactation to provide synthesis and recommendations to aid providers in counseling their patients. Methods: We conducted a scoping review by identifying the most recent clinical recommendations and guidelines from the Centers for Disease Control and Prevention (CDC) and three maternal and child health professional associations (American College of Obstetricians and Gynecologists [ACOG], Society for Maternal-Fetal Medicine [SMFM], and Academy of Breastfeeding Medicine [ABM]). We also reviewed the citations in these guidelines used in their development. We then conducted an updated literature review to capture studies published since the most recent systematic reviews were conducted. Results: We reviewed 1 clinical guideline from the CDC and 2 systematic reviews cited in its references, 6 professional association recommendations, and 28 publications identified through the updated literature review. Progestin-only contraceptive methods continue to demonstrate safety in breastfeeding patients, while low-quality evidence supports concerns of decreased milk supply with combined hormonal contraception. Discussion: Organizations should consider updating counseling recommendations regarding progestin-only contraceptives and lactation. Further research is needed to examine new contraceptive methods as well as the effect of hormonal contraception on lactation in the setting of preterm birth.

General and reproductive healthcare experiences of formerly incarcerated women in the United States: a qualitative study.

PURPOSE: The US prison population has recently reached an all-time high, with women representing the fastest growing subpopulation. Correctional health-care system in the USA remains fragmented and nonuniform in practice, particularly in women's health care, with poor transitions between incarceration and release. This study aims to examine the qualitative health-care experiences of women while incarcerated and their transition into the community health-care setting. Additionally, this study also examined the experiences of a subset of women who were pregnant while incarcerated. DESIGN/METHODOLOGY/APPROACH: After obtaining institutional review board approval, adult, English-speaking women with a history of incarceration within the past 10 years were interviewed using a semi-structured interview tool. Interview transcripts were analyzed using inductive content analysis. FINDINGS: The authors completed 21 full interviews and identified six themes that were both the most significant and most novel: "feeling stigmatized and insignificant," "care as punishment," "delay in care," "exceptions to the rule," "fragmentation of care" and "obstetric trauma and resilience." ORIGINALITY/VALUE: Women face numerous barriers and hardships when accessing basic and reproductive health-care services while incarcerated. This hardship is particularly challenging for women with substance use disorders. The authors were able to describe for the first time, partially through their own words, novel challenges described by women interacting with incarceration health care. Community providers should understand these barriers and challenges so as to effectively reengage women in care upon release and improve the health-care status of this historically marginalized group.

"Meeting Patients Where They're at": Clinician Perspectives on Integration of Family Planning Services into Office-Based Addiction Treatment.

OBJECTIVES: To study clinician perspectives on the feasibility of incorporating family planning services within office-based addiction treatment (OBAT) clinics. We sought to understand the unique facilitators of and barriers to the integration of contraceptive services within the OBAT model with a goal to support the design and implementation of a program tailored to meet the reproductive health needs of patients with substance use disorder. METHODS: After obtaining institutional review board approval, we conducted qualitative semistructured interviews with OBAT clinicians (registered nurses, advanced practice registered nurses, and physicians) at a tertiary-care safety-net hospital. Interview transcripts were analyzed using deductive codes utilizing key components of the Promoting Action on Research Implementation in Health Services and Ottawa Decision Support Frameworks. RESULTS: We analyzed 20 interviews. Our data noted 3 major themes: (1) evidence to support integration of family planning and OBAT, (2) inherent strengths and facilitative factors of the OBAT model, and (3) barriers and challenges of the OBAT model influencing successful integration. Strengths included the destigmatizing and trust-building OBAT approach to care, common use of patient-centered counseling, and providers' nuanced understanding of substance use disorder-specific impacts on reproductive health. Barriers included time constraints, balancing urgent patient recovery needs, the desire for additional contraception provision training, and concern for potential contraceptive coercion. CONCLUSIONS: Office-based addiction treatment clinics have inherent strengths that may make it a beneficial location for integrated family planning services. Future research should elicit patient perspectives to ensure the implementation of a family planning program in OBAT that supports patients' reproductive goals while avoiding stigma or reproductive coercion.

COVID-19 guidelines for pregnant women and new mothers: A systematic evidence review.

BACKGROUND: Nearly a year after COVID-19 was initially detected, guidance for pregnant and new mothers remains varied. OBJECTIVE: The goal of this systematic review is to summarize recommendations for three areas of maternal and fetal care-breastfeeding, post-partum social distancing, and decontamination. SEARCH STRATEGY: We searched PubMed, Embase and Web of Science spanning from inception to November 9, 2020. SELECTION CRITERIA: Articles were included if they focused on COVID-positive mothers, commented on at least one of the three areas of interest, and were published in English. DATA COLLECTION AND ANALYSIS: Our combined database search yielded 385 articles. After removing duplicates and articles that did not cover the correct populations or subject matter, a total of 74 articles remained in our analysis. MAIN RESULTS: Most articles recommended direct breastfeeding with enhanced precaution measures. Recommendations regarding post-partum social distancing varied, although articles published more recently often recommended keeping the mother and newborn in the same room when possible. Decontamination recommendations emphasized mask wearing, good hand hygiene, and proper cleaning of surfaces. CONCLUSION: In general, there was a focus on shared decision making when approaching topics such as breastfeeding and post-partum social distancing. Guidelines for decontamination were fairly uniform.

Vaginal cuff dehiscence following transvaginal oocyte retrieval: a case report.

BACKGROUND: Vaginal cuff dehiscence (VCD) is a rare but potentially serious complication following hysterectomy with an estimated incidence of 0.14-1.4%. There is a wide range of risk factors thought to contribute to VCD, but due to its rare occurrence, much still remains to be learned about the true impact of risk factors leading to dehiscence. We present here the second known report of VCD to occur in a patient undergoing transvaginal oocyte retrieval during her fertility treatment. This case highlights what may become a more common clinical scenario as more premenopausal women are diagnosed with reproductive tract cancers and access assisted reproductive therapies to preserve fertility. CASE PRESENTATION: Our patient is a 35-year-old G1 P0 A1 who had undergone ovary-sparing total laparoscopic hysterectomy (TLH) following diagnosis of endometrial adenocarcinoma. She underwent two in-vitro fertilization (IVF) cycles after TLH to bank frozen blastocysts, the first vaginal oocyte retrieval (VOR) taking place 12 weeks following hysterectomy. She experienced VCD during her second VOR that occurred 17 weeks after TLH, the second case of VCD to be reported in the literature during fertility preservation treatment following hysterectomy. The patient underwent an emergent and uncomplicated repair of the defect vaginally the same day. CONCLUSIONS: Currently there are no guidelines in place for women who have undergone hysterectomy with regard to when they can begin fertility treatment in the post-operative period. Based on now two case reports, it is worth considering extension of the typical 6-week timeline of avoidance of vaginal procedures to allow for full cuff healing. Infertility providers should also be mindful of limiting transvaginal ultrasounds where possible to reduce force along the cuff.

HIV-1 Nucleocapsid Mimics the Membrane Adaptor Syntenin PDZ to Gain Access to ESCRTs and Promote Virus Budding.

HIV-1 recruits cellular endosomal sorting complexes required for transport (ESCRTs) to bud virions from the membrane. Disruption of the viral nucleocapsid (NC) domain integrity affects HIV-1 budding. However, the molecular mechanisms of NC's involvement in HIV budding remain unclear. We find that NC mimics the PDZ domains of syntenin, a membrane-binding adaptor involved in cell-to-cell contact/communication, to capture the Bro1 domain of ALIX, which is an ESCRTs recruiting cellular adaptor. NC binds membranes via basic residues in either the distal or proximal zinc fingers, and NC-membrane binding is essential for Bro1 capture and HIV-1 budding. Removal of RNA enhances NC membrane binding, suggesting a dynamic competition between membrane lipids and RNA for the same binding sites in NC. Remarkably, syntenin PDZ can substitute for NC function in HIV-1 budding. Thus, NC mimics syntenin PDZs to function as a membrane-binding adaptor critical for HIV-1 budding at specific microdomains of the membrane.