Scalable stable comb-to-tone integrated RF photonic drive for superconducting qubits.

The recent advent of quantum computing has the potential to overhaul security, communications, and scientific modeling. Superconducting qubits are a leading platform that is advancing noise-tolerant intermediate-scale quantum processors. The implementation requires scaling to large numbers of superconducting qubits, circuit depths, and gate speeds, wherein high-purity RF signal generation and effective cabling transport are desirable. Fiber photonic-enhanced RF signal generation has demonstrated the principle of addressing both signal generation and transport requirements, supporting intermediate qubit numbers and robust packaging efforts; however, fiber-based approaches to RF signal distribution are often bounded by their phase instability. Here, we present a silicon photonic integrated circuit-based version of a photonic-enhanced RF signal generator that demonstrates the requisite stability, as well as a path towards the necessary signal fidelity.

Comprehensive cancer predisposition testing within the prospective MASTER trial identifies hereditary cancer patients and supports treatment decisions for rare cancers.

BACKGROUND: Germline variant evaluation in precision oncology opens new paths toward the identification of patients with genetic tumor risk syndromes and the exploration of therapeutic relevance. Here, we present the results of germline variant analysis and their clinical implications in a precision oncology study for patients with predominantly rare cancers. PATIENTS AND METHODS: Matched tumor and control genome/exome and RNA sequencing was carried out for 1485 patients with rare cancers (79%) and/or young adults (77% younger than 51 years) in the National Center for Tumor Diseases/German Cancer Consortium (NCT/DKTK) Molecularly Aided Stratification for Tumor Eradication Research (MASTER) trial, a German multicenter, prospective, observational precision oncology study. Clinical and therapeutic relevance of prospective pathogenic germline variant (PGV) evaluation was analyzed and compared to other precision oncology studies. RESULTS: Ten percent of patients (n = 157) harbored PGVs in 35 genes associated with autosomal dominant cancer predisposition, whereof up to 75% were unknown before study participation. Another 5% of patients (n = 75) were heterozygous carriers for recessive genetic tumor risk syndromes. Particularly, high PGV yields were found in patients with gastrointestinal stromal tumors (GISTs) (28%, n = 11/40), and more specifically in wild-type GISTs (50%, n = 10/20), leiomyosarcomas (21%, n = 19/89), and hepatopancreaticobiliary cancers (16%, n = 16/97). Forty-five percent of PGVs (n = 100/221) supported treatment recommendations, and its implementation led to a clinical benefit in 40% of patients (n = 10/25). A comparison of different precision oncology studies revealed variable PGV yields and considerable differences in germline variant analysis workflows. We therefore propose a detailed workflow for germline variant evaluation. CONCLUSIONS: Genetic germline testing in patients with rare cancers can identify the very first patient in a hereditary cancer family and can lead to clinical benefit in a broad range of entities. Its routine implementation in precision oncology accompanied by the harmonization of germline variant evaluation workflows will increase clinical benefit and boost research.

Reduction of the radiative decay of atomic coherence in squeezed vacuum.

Quantum fluctuations of the electromagnetic vacuum are responsible for physical effects such as the Casimir force and the radiative decay of atoms, and set fundamental limits on the sensitivity of measurements. Entanglement between photons can produce correlations that result in a reduction of these fluctuations below the ordinary vacuum level, allowing measurements that surpass the standard quantum limit in sensitivity. The effects of such 'squeezed states' of light on matter were first considered in a prediction of the radiative decay rates of atoms in squeezed vacuum. Despite efforts to demonstrate such effects in experiments with natural atoms, a direct quantitative observation of this prediction has remained elusive. Here we report a twofold reduction of the transverse radiative decay rate of a superconducting artificial atom coupled to continuum squeezed vacuum. The artificial atom is effectively a two-level system formed by the strong interaction between a superconducting circuit and a microwave-frequency cavity. A Josephson parametric amplifier is used to generate quadrature-squeezed electromagnetic vacuum. The observed twofold reduction in the decay rate of the atom allows the transverse coherence time, T2, to exceed the ordinary vacuum decay limit, 2T1. We demonstrate that the measured radiative decay dynamics can be used to reconstruct the Wigner distribution of the itinerant squeezed state. Our results confirm a canonical prediction of quantum optics and should enable new studies of the quantum light-matter interaction.

Antipsychotic plasma levels in the assessment of poor treatment response in schizophrenia.

OBJECTIVE: Treatment resistance is a challenge for the management of schizophrenia. It is not always clear whether inadequate response is secondary to medication ineffectiveness, as opposed to medication underexposure due to non-adherence or pharmacokinetic factors. We investigated the prevalence of subtherapeutic antipsychotic plasma levels in patients identified as treatment-resistant by their treating clinician. METHOD: Between January 2012 and April 2017, antipsychotic plasma levels were measured in 99 individuals provisionally diagnosed with treatment-resistant schizophrenia by their treating clinicians, but not prescribed clozapine. Patients were followed up to determine whether they were subsequently admitted to hospital. RESULTS: Thirty-five per cent of plasma levels were subtherapeutic, and of these, 34% were undetectable. Black ethnicity (P = 0.006) and lower dose (P < 0.001) were significantly associated with subtherapeutic/undetectable plasma levels. Individuals with subtherapeutic/undetectable levels were significantly more likely to be admitted to hospital (P = 0.02). CONCLUSION: A significant proportion of patients considered treatment-resistant have subtherapeutic antipsychotic plasma levels, and this is associated with subsequent admission. The presence of subtherapeutic plasma levels may suggest a need to address adherence or pharmacokinetic factors as opposed to commencing clozapine treatment. While antipsychotic levels are not recommended for the routine adjustment of dosing, they may assist with the assessment of potential treatment resistance in schizophrenia.

Associations between dietary patterns, socio-demographic factors and anthropometric measurements in adult New Zealanders: an analysis of data from the 2008/09 New Zealand Adult Nutrition Survey.

PURPOSE: To investigate associations between dietary patterns, socio-demographic factors and anthropometric measurements in adult New Zealanders. METHODS: Dietary patterns were identified using factor analysis in adults 15 years plus (n = 4657) using 24-h diet recall data from the 2008/09 New Zealand Adult Nutrition Survey. Multivariate regression was used to investigate associations between dietary patterns and age, gender and ethnicity. After controlling for demographic factors, associations between dietary patterns and food insecurity, deprivation, education, and smoking were investigated. Associations between dietary patterns and body mass index and waist circumference were examined adjusting for demographic factors, smoking and energy intake. RESULTS: Two dietary patterns were identified. 'Healthy' was characterised by breakfast cereal, low fat milk, soy and rice milk, soup and stock, yoghurt, bananas, apples, other fruit and tea, and low intakes of pies and pastries, potato chips, white bread, takeaway foods, soft drinks, beer and wine. 'Traditional' was characterised by beef, starchy vegetables, green vegetables, carrots, tomatoes, savoury sauces, regular milk, cream, sugar, tea and coffee, and was low in takeaway foods. The 'healthy' pattern was positively associated with age, female gender, New Zealand European or other ethnicity, and a secondary school qualification, and inversely associated with smoking, food insecurity, area deprivation, BMI and waist circumference. The 'traditional' pattern was positively associated with age, male gender, smoking, food insecurity and inversely associated with a secondary school qualification. CONCLUSIONS: A 'Healthy' dietary pattern was associated with higher socio-economic status and reduced adiposity, while the 'traditional' pattern was associated with lower socio-economic status.

The impact of HLA class I and EBV latency-II antigen-specific CD8(+) T cells on the pathogenesis of EBV(+) Hodgkin lymphoma.

In 40% of cases of classical Hodgkin lymphoma (cHL), Epstein-Barr virus (EBV) latency-II antigens [EBV nuclear antigen 1 (EBNA1)/latent membrane protein (LMP)1/LMP2A] are present (EBV(+) cHL) in the malignant cells and antigen presentation is intact. Previous studies have shown consistently that HLA-A*02 is protective in EBV(+) cHL, yet its role in disease pathogenesis is unknown. To explore the basis for this observation, gene expression was assessed in 33 cHL nodes. Interestingly, CD8 and LMP2A expression were correlated strongly and, for a given LMP2A level, CD8 was elevated markedly in HLA-A*02(-) versus HLA-A*02(+) EBV(+) cHL patients, suggesting that LMP2A-specific CD8(+) T cell anti-tumoral immunity may be relatively ineffective in HLA-A*02(-) EBV(+) cHL. To ascertain the impact of HLA class I on EBV latency antigen-specific immunodominance, we used a stepwise functional T cell approach. In newly diagnosed EBV(+) cHL, the magnitude of ex-vivo LMP1/2A-specific CD8(+) T cell responses was elevated in HLA-A*02(+) patients. Furthermore, in a controlled in-vitro assay, LMP2A-specific CD8(+) T cells from healthy HLA-A*02 heterozygotes expanded to a greater extent with HLA-A*02-restricted compared to non-HLA-A*02-restricted cell lines. In an extensive analysis of HLA class I-restricted immunity, immunodominant EBNA3A/3B/3C-specific CD8(+) T cell responses were stimulated by numerous HLA class I molecules, whereas the subdominant LMP1/2A-specific responses were confined largely to HLA-A*02. Our results demonstrate that HLA-A*02 mediates a modest, but none the less stronger, EBV-specific CD8(+) T cell response than non-HLA-A*02 alleles, an effect confined to EBV latency-II antigens. Thus, the protective effect of HLA-A*02 against EBV(+) cHL is not a surrogate association, but reflects the impact of HLA class I on EBV latency-II antigen-specific CD8(+) T cell hierarchies.

A distinct immunogenic region of glutamic acid decarboxylase 65 is naturally processed and presented by human islet cells to cytotoxic CD8 T cells.

CD8 T cells specific for islet autoantigens are major effectors of ß cell damage in type 1 diabetes, and measurement of their number and functional characteristics in blood represent potentially important disease biomarkers. CD8 T cell reactivity against glutamic acid decarboxylase 65 (GAD65) in HLA-A*0201 subjects has been reported to focus on an immunogenic region 114-123 (VMNILLQYVV), with studies demonstrating both 114-123 and 114-122 epitopes being targeted. However, the fine specificity of this response is unclear and the key question as to which epitope(s) ß cells naturally process and present and, therefore, the pathogenic potential of CD8 T cells with different specificities within this region has not been addressed. We generated human leucocyte antigen (HLA)-A*0201-restricted CD8 T cell clones recognizing either 114-122 alone or both 114-122 and 114-123. Both clone types show potent and comparable effector functions (cytokine and chemokine secretion) and killing of indicator target cells externally pulsed with cognate peptide. However, only clones recognizing 114-123 kill target cells transfected with HLA-A*0201 and GAD2 and HLA-A*0201(+) human islet cells. We conclude that the endogenous pathway of antigen processing by HLA-A*0201-expressing cells generates GAD65114-123 as the predominant epitope in this region. These studies highlight the importance of understanding ß cell epitope presentation in the design of immune monitoring for potentially pathogenic CD8 T cells.

The practical management of refractory schizophrenia--the Maudsley Treatment REview and Assessment Team service approach.

OBJECTIVE: To describe a practical approach to the community management of treatment-resistant schizophrenia (TRS). METHOD: A descriptive review of an approach to the assessment and management of patients with TRS, including the community titration of clozapine treatment, and a report of the management recommendations for the first one hundred patients assessed by the Treatment REview and Assessment Team (TREAT). RESULTS: The standardized model for the community assessment, management and titration of clozapine is described. To date, 137 patients have been referred to this service and 100 patients (72%) attended for assessment. Of these, 33 have been initiated on clozapine while fifteen have had clozapine recommended but have not wished to undertake clozapine treatment. Other management options recommended have included augmentation strategies and long-acting injectable antipsychotics. CONCLUSION: The service had increased the number of patients receiving community assessment and initiation of clozapine by five-fold relative to the rate prior to the establishment of the service. The large number of referrals and high attendance rate indicates that there is clinical demand for the model. Systematic evaluation is required to determine the clinical and cost-effectiveness of this model and its potential application to other clinical settings.

The gene encoding proline dehydrogenase modulates sensorimotor gating in mice.

Hemizygous cryptic deletions of the q11 band of human chromosome 22 have been associated with a number of psychiatric and behavioural phenotypes, including schizophrenia. Here we report the isolation and characterization of PRODH, a human homologue of Drosophila melanogaster sluggish-A (slgA), which encodes proline dehydrogenase responsible for the behavioural phenotype of the slgA mutant. PRODH is localized at chromosome 22q11 in a region deleted in some psychiatric patients. We also isolated the mouse homologue of slgA (Prodh), identified a mutation in this gene in the Pro/Re hyperprolinaemic mouse strain and found that these mice have a deficit in sensorimotor gating accompanied by regional neurochemical alterations in the brain. Sensorimotor gating is a neural filtering process that allows attention to be focused on a given stimulus, and is affected in patients with neuropsychiatric disorders. Furthermore, several lines of evidence suggest that proline may serve as a modulator of synaptic transmission in the mammalian brain. Our observations, in conjunction with the chromosomal location of PRODH, suggest a potential involvement of this gene in the 22q11-associated psychiatric and behavioural phenotypes.

Dynamics of antibiotic resistance markers and Escherichia coli invasion in riverine heterotrophic biofilms facing increasing heat and flow stagnation.

As motivation to address environmental dissemination of antimicrobial resistance (AMR) is mounting, there is a need to characterize mechanisms by which AMR can propagate under environmental conditions. Here we investigated the effect of temperature and stagnation on the persistence of wastewater-associated antibiotic resistance markers in riverine biofilms and the invasion success of genetically-tagged Escherichia coli. Biofilms grown on glass slides incubated in-situ downstream of a wastewater treatment plant effluent discharge point were transferred to laboratory-scale flumes fed with filtered river water under potentially stressful temperature and flow conditions: recirculation flow at 20 °C, stagnation at 20 °C, and stagnation at 30 °C. After 14 days, quantitative PCR and amplicon sequencing were used to quantify bacteria, biofilms diversity, resistance markers (sul1, sul2, ermB, tetW, tetM, tetB, blaCTX-M-1, intI1) and E. coli. Resistance markers significantly decreased over time regardless of the treatment applied. Although invading E. coli were initially able to colonize the biofilms, its abundance subsequently declined. Stagnation was associated with a shift in biofilm taxonomic composition, but there was no apparent effect of flow conditions or the simulated river-pool warming (30 °C) on AMR persistence or invasion success of E. coli. Results however indicated that antibiotic resistance markers in the riverine biofilms decreased under the experimental conditions in the absence of exposure to external inputs of antibiotics and AMR.

Comparison of the survival of different isolates of SARS-CoV-2 in evaporating aerosols.

Numerous variants of SARS-CoV-2 with increased transmissibility have emerged over the course of the pandemic. Potential explanations for the increased transmissibility of these variants include increased shedding from infected individuals, increased environmental stability, and/or a lower infectious dose. Upon exhalation of a respiratory particle into the environment, water present in the particle is rapidly lost through evaporation, resulting in a decrease in particle size. The aim of the present study was to compare the losses of infectivity of different isolates of SARS-CoV-2 during the rapid evaporation of aerosol particles that occurs immediately post-generation to assess if there are differences suggestive of increased survival, and ultimately greater transmissibility, for more recent variants. Losses of infectivity of several isolates of SARS-CoV-2 suspended in viral culture media were assessed following aerosolization and evaporation in a flowing chamber. The results demonstrate that losses of infectivity measured post-evaporation were similar for three different isolates of SARS-CoV-2, including isolates from the more recent Delta and Omicron lineages. The average loss in infectivity across all three isolates was 61 ± 15% (-0.46 ± 0.17 log10 TCID50/L-air) at a relative humidity <30%. These results, together with those from several previous studies, suggest that it is unlikely that an increase in environmental stability contributes to the observed increases in transmissibility observed with more recent variants of SARS-CoV-2.

Organization and function of the cytoskeleton in polarized epithelial cells: a component of the protein sorting machinery.

Development and maintenance of cell-surface polarity in epithelial cells requires specialized localization of proteins to functionally and structurally distinct plasma membrane domains. The organization of these domains is dependent upon targeted delivery of transport vesicles between different membrane compartments, and upon protein sorting in the membranes of the Golgi complex and cell surface. Increasing evidence has been gathered in recent years that cytoskeletal components facilitate these processes.

Transmission of classical scrapie to wild-type mice: the influence of the ovine PrP sequence on lesion profiles.

Susceptibility of sheep to classical scrapie is determined by polymorphisms in the coding region of the prion protein gene (PRNP), mainly at codons 136, 154 and 171. It has recently been shown that lesion profiles from classical field scrapie isolates that transmitted to RIII mice can be classified into different groups. There was also strong, but not absolute, association between the different groups and codon 136. Here, we examine the hypothesis that additional polymorphisms in the open reading frame sequence of the ovine PRNP may account for the different groups of lesion profiles observed following transmission to mice.

Variability in pulmonary function following rapid altitude ascent to the Amundsen-Scott South Pole station.

The impact of acute altitude exposure on pulmonary function is variable. A large inter-individual variability in the changes in forced expiratory flows (FEFs) is reported with acute exposure to altitude, which is suggested to represent an interaction between several factors influencing bronchial tone such as changes in gas density, catecholamine stimulation, and mild interstitial edema. This study examined the association between FEF variability, acute mountain sickness (AMS) and various blood markers affecting bronchial tone (endothelin-1, vascular endothelial growth factor (VEGF), catecholamines, angiotensin II) in 102 individuals rapidly transported to the South Pole (2835 m). The mean FEF between 25 and 75% (FEF(25-75)) and blood markers were recorded at sea level and after the second night at altitude. AMS was assessed using Lake Louise questionnaires. FEF(25-75) increased by an average of 12% with changes ranging from -26 to +59% from sea level to altitude. On the second day, AMS incidence was 36% and was higher in individuals with increases in FEF(25-75) (41 vs. 22%, P = 0.05). Ascent to altitude induced an increase in endothelin-1 levels, with greater levels observed in individuals with decreased FEF(25-75). Epinephrine levels increased with ascent to altitude and the response was six times larger in individuals with decreased FEF(25-75). Greater levels of endothelin-1 in individuals with decreased FEF(25-75) suggest a response consistent with pulmonary hypertension and/or mild interstitial edema, while epinephrine may be upregulated in these individuals to clear lung fluid through stimulation of ß(2)-adrenergic receptors.

Evidence of prophylaxis treatment in adult haemophiliacs. Need of this treatment strategy?

In contrast to children with haemophilia the scientific evidence of prophylaxis treatment in adults is not yet proven. Existing studies are of observational character and mostly retrospectively designed. Therefore, opinion leaders in this field postulate prospectively designed, randomized, controlled and multicentric studies to set up urgently needed guidelines. Evidence according to the Canadian task force ranking is assessed as level III with a recommendation grade C by the authors. The recognition of benefits of health care providers in accordance with the German Federal Joint Committee generally demands a Grade-Ia to Ib evidence. As long as the actual evidence of prophylaxis in adult haemophiliacs does not meet the postulated criteria of the German Federal Joint Committee, prophylactic replacement therapy of the individual case has to be well documented and reasonably explained.

The impacts of intensive mining on terrestrial and aquatic ecosystems: A case of sediment pollution and calcium decline in cool temperate Tasmania, Australia.

Mining causes extensive damage to aquatic ecosystems via acidification, heavy metal pollution, sediment loading, and Ca decline. Yet little is known about the effects of mining on freshwater systems in the Southern Hemisphere. A case in point is the region of western Tasmania, Australia, an area extensively mined in the 19th century, resulting in severe environmental contamination. In order to assess the impacts of mining on aquatic ecosystems in this region, we present a multiproxy investigation of the lacustrine sediments from Owen Tarn, Tasmania. This study includes a combination of radiometric dating (14C and 210Pb), sediment geochemistry (XRF and ICP-MS), pollen, charcoal and diatoms. Generalised additive mixed models were used to test if changes in the aquatic ecosystem can be explained by other covariates. Results from this record found four key impact phases: (1) Pre-mining, (2) Early mining, (3) Intense mining, and (4) Post-mining. Before mining, low heavy metal concentrations, slow sedimentation, low fire activity, and high biomass indicate pre-impact conditions. The aquatic environment at this time was oligotrophic and dystrophic with sufficient light availability, typical of western Tasmanian lakes during the Holocene. Prosperous mining resulted in increased burning, a decrease in landscape biomass and an increase in sedimentation resulting in decreased light availability of the aquatic environment. Extensive mining at Mount Lyell in the 1930s resulted in peak heavy metal pollutants (Pb, Cu and Co) and a further increase in inorganic inputs resulted in a disturbed low light lake environment (dominated by Hantzschia amphioxys and Pinnularia divergentissima). Following the closure of the Mount Lyell Co. in 1994 CE, Ca declined to below pre-mining levels resulting in a new diatom assemblage and deformed diatom valves. Therefore, the Owen Tarn record demonstrates severe sediment pollution and continued impacts of mining long after mining has stopped at Mt. Lyell Mining Co.

Correction to: Practical issues and limitations of brain attenuation correction on a simultaneous PET-MR scanner.

An amendment to this paper has been published and can be accessed via the original article.

Practical issues and limitations of brain attenuation correction on a simultaneous PET-MR scanner.

BACKGROUND: Despite the advent of clinical PET-MR imaging for routine use in 2011 and the development of several methods to address the problem of attenuation correction, some challenges remain. We have identified and investigated several issues that might affect the reliability and accuracy of current attenuation correction methods when these are implemented for clinical and research studies of the brain. These are (1) the accuracy of converting CT Hounsfield units, obtained from an independently acquired CT scan, to 511 keV linear attenuation coefficients; (2) the effect of padding used in the MR head coil; (3) the presence of close-packed hair; (4) the effect of headphones. For each of these, we have examined the effect on reconstructed PET images and evaluated practical mitigating measures. RESULTS: Our major findings were (1) for both Siemens and GE PET-MR systems, CT data from either a Siemens or a GE PET-CT scanner may be used, provided the conversion to 511 keV µ-map is performed by the PET-MR vendor's own method, as implemented on their PET-CT scanner; (2) the effect of the head coil pads is minimal; (3) the effect of dense hair in the field of view is marked (> 10% error in reconstructed PET images); and (4) using headphones and not including them in the attenuation map causes significant errors in reconstructed PET images, but the risk of scanning without them may be acceptable following sound level measurements. CONCLUSIONS: It is important that the limitations of attenuation correction in PET-MR are considered when designing research and clinical PET-MR protocols in order to enable accurate quantification of brain PET scans. Whilst the effect of pads is not significant, dense hair, the use of headphones and the use of an independently acquired CT-scan can all lead to non-negligible effects on PET quantification. Although seemingly trivial, these effects add complications to setting up protocols for clinical and research PET-MR studies that do not occur with PET-CT. In the absence of more sophisticated PET-MR brain attenuation correction, the effect of all of the issues above can be minimised if the pragmatic approaches presented in this work are followed.

Attachment to an endogenous laminin-like protein initiates sprouting by leech neurons.

Leech neurons in culture sprout rapidly when attached to extracts from connective tissue surrounding the nervous system. Laminin-like molecules that promote sprouting have now been isolated from this extracellular matrix. Two mAbs have been prepared that react on immunoblots with a approximately equal to 220- and a approximately equal to 340-kD polypeptide, respectively. These antibodies have been used to purify molecules with cross-shaped structures in the electron microscope. The molecules, of approximately equal to 10(3) kD on nonreducing SDS gels, have subunits of approximately equal to 340, 220, and 160-180 kD. Attachment to the laminin-like molecules was sufficient to initiate sprouting by single isolated leech neurons in defined medium. This demonstrates directly a function for a laminin-related invertebrate protein. The mAbs directed against the approximately equal to 220-kD chains of the laminin-like leech molecule labeled basement membrane extracellular matrix in leech ganglia and nerves. A polyclonal antiserum against the approximately equal to 220-kD polypeptide inhibited neurite outgrowth. Vertebrate laminin did not mediate the sprouting of leech neurons; similarly, the leech molecule was an inert substrate for vertebrate neurons. Although some traits of structure, function, and distribution are conserved between vertebrate laminin and the invertebrate molecule, our results suggest that the functional domains differ.

Clathrin assembly protein AP-2 induces aggregation of membrane vesicles: a possible role for AP-2 in endosome formation.

We have examined the in vitro behavior of clathrin-coated vesicles that have been stripped of their surface coats such that the majority of the clathrin is removed but substantial amounts of clathrin assembly proteins (AP) remain membrane-associated. Aggregation of these stripped coated vesicles (s-CV) is observed when they are placed under conditions that approximate the pH and ionic strength of the cell interior (pH 7.2, approximately 100 mM salt). This s-CV aggregation reaction is rapid (t1/2 < or = 0.5 min), independent of temperature within a range of 4-37 degrees C, and unaffected by ATP, guanosine-5'-O-(3-thiophosphate), and in particular EGTA, distinguishing it from Ca(2+)-dependent membrane aggregation reactions. The process is driven by the action of membrane-associated AP molecules since partial proteolysis results in a full loss of activity and since aggregation is abolished by pretreatment of the s-CVs with a monoclonal antibody that reacts with the alpha subunit of AP-2. However, vesicle aggregation is not inhibited by PPPi, indicating that the previously characterized polyphosphate-sensitive AP-2 self-association is not responsible for the reaction. The vesicle aggregation reaction can be reconstituted: liposomes of phospholipid composition approximating that found on the cytoplasmic surfaces of the plasma membrane and of coated vesicles (70% L-alpha-phosphatidylethanolamine (type I-A), 15% L-alpha-phosphatidyl-L-serine, and 15% L-alpha-phosphatidylinositol) aggregated after addition of AP-2, but not of AP-1, AP-3 (AP180), or pure clathrin triskelions. Aggregation of liposomes is abolished by limited proteolysis of AP-2 with trypsin. In addition, a highly purified AP-2 alpha preparation devoid of beta causes liposome aggregation, whereas pure beta subunit does not, consistent with results obtained in the s-CV assay which also indicate the involvement of the alpha subunit. Using a fluorescence energy transfer assay we show that AP-2 does not cause fusion of liposomes under physiological solution conditions. However, since the fusion of membranes necessarily requires the close opposition of the two participating bilayers, the AP-2-dependent vesicle aggregation events that we have identified may represent an initial step in the formation and fusion of endosomes that occur subsequent to endocytosis and clathrin uncoating in vivo.