Single-molecule digital sizing of proteins in solution.

The physical characterization of proteins in terms of their sizes, interactions, and assembly states is key to understanding their biological function and dysfunction. However, this has remained a difficult task because proteins are often highly polydisperse and present as multicomponent mixtures. Here, we address this challenge by introducing single-molecule microfluidic diffusional sizing (smMDS). This approach measures the hydrodynamic radius of single proteins and protein assemblies in microchannels using single-molecule fluorescence detection. smMDS allows for ultrasensitive sizing of proteins down to femtomolar concentrations and enables affinity profiling of protein interactions at the single-molecule level. We show that smMDS is effective in resolving the assembly states of protein oligomers and in characterizing the size of protein species within complex mixtures, including fibrillar protein aggregates and nanoscale condensate clusters. Overall, smMDS is a highly sensitive method for the analysis of proteins in solution, with wide-ranging applications in drug discovery, diagnostics, and nanobiotechnology.

Clinical Implications of HIV Treatment and Prevention for Polygamous Families in Kenya and Uganda: "My Co-Wife Is the One Who Used to Encourage Me".

Polygamy is the practice of marriage to multiple partners. Approximately 6-11% of households in Uganda and 4-11% of households in Kenya are polygamous. The complex families produced by polygamous marriage customs give rise to additional considerations for healthcare providers and public health messaging around HIV care. Using 27 in-depth, semi-structured qualitative interviews with participants in two studies in rural Kenya and Uganda, we analysed challenges and opportunities that polygamous families presented in the diagnosis, treatment and prevention of HIV, and provider roles in improving HIV outcomes in these families. Overall, prevention methods seemed more justifiable to families where co-wives live far apart than when all members live in the same household. In treatment, diagnosis of one member did not always lead to disclosure to other members, creating an adverse home environment; but sometimes diagnosis of one wife led not only to diagnosis of the other, but also to greater household support.

Clinical implications of HIV treatment and prevention for polygamous families in Kenya and UgandaPolygamy is the practice of marriage to multiple partners. Approximately 6-11% of households in Uganda and 4-11% of households in Kenya are polygamous. The complex families produced by polygamous marriage customs give rise to additional considerations for healthcare providers and public health messaging around HIV care.

eIF4F is a thermo-sensing regulatory node in the translational heat shock response.

Heat-shocked cells prioritize the translation of heat shock (HS) mRNAs, but the underlying mechanism is unclear. We report that HS in budding yeast induces the disassembly of the eIF4F complex, where eIF4G and eIF4E assemble into translationally arrested mRNA ribonucleoprotein particles (mRNPs) and HS granules (HSGs), whereas eIF4A promotes HS translation. Using in vitro reconstitution biochemistry, we show that a conformational rearrangement of the thermo-sensing eIF4A-binding domain of eIF4G dissociates eIF4A and promotes the assembly with mRNA into HS-mRNPs, which recruit additional translation factors, including Pab1p and eIF4E, to form multi-component condensates. Using extracts and cellular experiments, we demonstrate that HS-mRNPs and condensates repress the translation of associated mRNA and deplete translation factors that are required for housekeeping translation, whereas HS mRNAs can be efficiently translated by eIF4A. We conclude that the eIF4F complex is a thermo-sensing node that regulates translation during HS.

PARP1-DNA co-condensation drives DNA repair site assembly to prevent disjunction of broken DNA ends.

DNA double-strand breaks (DSBs) are repaired at DSB sites. How DSB sites assemble and how broken DNA is prevented from separating is not understood. Here we uncover that the synapsis of broken DNA is mediated by the DSB sensor protein poly(ADP-ribose) (PAR) polymerase 1 (PARP1). Using bottom-up biochemistry, we reconstitute functional DSB sites and show that DSB sites form through co-condensation of PARP1 multimers with DNA. The co-condensates exert mechanical forces to keep DNA ends together and become enzymatically active for PAR synthesis. PARylation promotes release of PARP1 from DNA ends and the recruitment of effectors, such as Fused in Sarcoma, which stabilizes broken DNA ends against separation, revealing a finely orchestrated order of events that primes broken DNA for repair. We provide a comprehensive model for the hierarchical assembly of DSB condensates to explain DNA end synapsis and the recruitment of effector proteins for DNA damage repair.

Intra-condensate demixing of TDP-43 inside stress granules generates pathological aggregates.

Cytosolic aggregation of the nuclear protein TDP-43 is associated with many neurodegenerative diseases, but the triggers for TDP-43 aggregation are still debated. Here, we demonstrate that TDP-43 aggregation requires a double event. One is up-concentration in stress granules beyond a threshold, and the other is oxidative stress. These two events collectively induce intra-condensate demixing, giving rise to a dynamic TDP-43 enriched phase within stress granules, which subsequently transitions into pathological aggregates. Mechanistically, intra-condensate demixing is triggered by local unfolding of the RRM1 domain for intermolecular disulfide bond formation and by increased hydrophobic patch interactions in the C-terminal domain. By engineering TDP-43 variants resistant to intra-condensate demixing, we successfully eliminate pathological TDP-43 aggregates in cells. We conclude that up-concentration inside condensates and simultaneous exposure to environmental stress could be a general pathway for protein aggregation, with intra-condensate demixing constituting a key intermediate step.

Ground penetrating radar observations of the contact between the western delta and the crater floor of Jezero crater, Mars.

The delta deposits in Jezero crater contain sedimentary records of potentially habitable conditions on Mars. NASA's Perseverance rover is exploring the Jezero western delta with a suite of instruments that include the RIMFAX ground penetrating radar, which provides continuous subsurface images that probe up to 20 meters below the rover. As Perseverance traversed across the contact between the Jezero crater floor and the delta, RIMFAX detected a distinct discontinuity in the subsurface layer structure. Below the contact boundary are older crater floor units exhibiting discontinuous inclined layering. Above the contact boundary are younger basal delta units exhibiting regular horizontal layering. At one location, there is a clear unconformity between the crater floor and delta layers, which implies that the crater floor experienced a period of erosion before the deposition of the overlying delta strata. The regularity and horizontality of the basal delta sediments observed in the radar cross sections indicate that they were deposited in a low-energy lake environment.

Improving patient care by virtual case discussion between plastic surgeons and residents of Uganda and the Netherlands.

INTRODUCTION: Traditional on-site missions of plastic surgeons from "high-income countries" in "low- and middle-income countries" are often limited in time and lack proper follow-up. Regular digital collaboration could lead to a more impactful and durable exchange of knowledge for plastic surgeons and residents in both settings. AIMS: The aim of this study was to evaluate the satisfaction of the first twelve months of weekly digital meetings, explore advantages/disadvantages, and to provide tools for similar initiatives. METHODS: Weekly meetings started from August 2021. An encrypted digital connection allowed residents and plastic surgeons from Uganda and the Netherlands to discuss cases for educational purposes, where treatment options were considered. After twelve months, a survey was sent to participants from both countries to indicate the meetings' strengths, weaknesses, and possible improvements. RESULTS: A total of 18 participants responded to the questionnaire (ten plastic surgeons, six residents, and two researchers). The strengths of the meetings were the accessibility of the meetings, knowledge exchange and practice for residents' final exams. Possible improvements included having a clear format for patient discussion, a session moderator and better internet connectivity. Moreover, a database to assess the impact of the given intervention on the patient cases by evaluating postoperatively (e.g. three months), could further improve clinical care. CONCLUSIONS: Virtual patient discussions subjectively contributed to medical education at both locations. Improved digital infrastructure and a collaborative database could further maximize learning capacity. Furthermore, digital proctoring is a promising way to establish sustainable collaborations between high- and low-resource countries.

Smart surgical glasses for orofacial cleft surgery training in high- and low-income countries: A proof of concept.

INTRODUCTION: The coronavirus 2019 pandemic urged us to find alternatives for education through remote proctoring and international surgical collaborations among high-, middle-, and low-income countries. Smart surgical glasses are promising for remote surgical education and international surgical collaborations. AIMS: This study aimed to assess the usability of smart surgical glasses during cleft surgery and explore their potential in remote surgical education and collaboration. METHODS: Six plastic surgical cases were randomly selected and recorded using the RODS&CONES glasses in 4K (3840 × 2160p). A 23-point questionnaire was sent to one plastic surgeon, one plastic surgery resident, and eight doctors who were not trained to critically appraise the video and audio quality of the smart surgical glasses and their applicability for remote surgical education. RESULTS: The participants indicated that the smart glasses had several significant advantages over conventional on-site education, such as facilitating a better view of the surgical field and providing possibilities for remote interaction. The audio quality was considered excellent. The main limitations were image stabilization issues and loss of video connection due to weak wireless fidelity. CONCLUSIONS: All participants appreciated the use of smart glasses for remote education and considered them a promising tool for enhancing the quality of surgical education. The glasses can enable remote assistance and education of local surgical residents and may facilitate sustainable surgical collaborations among high-, middle-, and low-income countries.