Age at menarche and risk of vasomotor menopausal symptoms: a pooled analysis of six studies.

OBJECTIVE: To examine the association between age at menarche and risk of vasomotor menopausal symptoms (VMS) and whether midlife body mass index (BMI) modified the association. DESIGN: A pooled analysis of six cohort studies. SETTING: The International collaboration on the Life course Approach to reproductive health and Chronic disease Events (InterLACE). POPULATION: 18 555 women from the UK, USA and Australia. METHODS: VMS frequency data (never, rarely, sometimes and often) were harmonised from two studies (n = 13 602); severity data (never, mild, moderate and severe) from the other four studies (n = 4953). Multinominal logistic regression models were used to estimate relative risk ratios (RRRs) and 95% CIs adjusted for confounders and incorporated study as random effects. MAIN OUTCOME MEASURES: Hot flushes and night sweats. RESULTS: Frequency data showed that early menarche ≤11 years was associated with an increased risk of 'often' hot flushes (RRR 1.48, 95% CI 1.24-1.76) and night sweats (RRR 1.59, 95% CI 1.49-1.70) compared with menarche at ≥14 years. Severity data showed similar results, but appeared less conclusive, with RRRs of 1.16 (95% CI 0.94-1.42) and 1.27 (95% CI 1.01-1.58) for 'severe' hot flushes and night sweats, respectively. BMI significantly modified the association as the risk associated with early menarche and 'often' VMS was stronger among women who were overweight or obese than those of normal weight, while this gradient across BMI categories was not as strong with the risk of 'severe' VMS. CONCLUSIONS: Early age at menarche is a risk factor for VMS, particularly for frequent VMS, but midlife BMI may play an important role in modifying this risk. TWEETABLE ABSTRACT: Overweight and obesity exacerbate the risk of vasomotor symptoms associated with early menarche.

Selective Targeting of High-Affinity LFA-1 Does Not Augment Costimulation Blockade in a Nonhuman Primate Renal Transplantation Model.

Costimulation blockade (CoB) via belatacept is a lower-morbidity alternative to calcineurin inhibitor (CNI)-based immunosuppression. However, it has higher rates of early acute rejection. These early rejections are mediated in part by memory T cells, which have reduced dependence on the pathway targeted by belatacept and increased adhesion molecule expression. One such molecule is leukocyte function antigen (LFA)-1. LFA-1 exists in two forms: a commonly expressed, low-affinity form and a transient, high-affinity form, expressed only during activation. We have shown that antibodies reactive with LFA-1 regardless of its configuration are effective in eliminating memory T cells but at the cost of impaired protective immunity. Here we test two novel agents, leukotoxin A and AL-579, each of which targets the high-affinity form of LFA-1, to determine whether this more precise targeting prevents belatacept-resistant rejection. Despite evidence of ex vivo and in vivo ligand-specific activity, neither agent when combined with belatacept proved superior to belatacept monotherapy. Leukotoxin A approached a ceiling of toxicity before efficacy, while AL-579 failed to significantly alter the peripheral immune response. These data, and prior studies, suggest that LFA-1 blockade may not be a suitable adjuvant agent for CoB-resistant rejection.

Anti-Leukocyte Function-Associated Antigen 1 Therapy in a Nonhuman Primate Renal Transplant Model of Costimulation Blockade-Resistant Rejection.

Costimulation blockade with the fusion protein belatacept provides a desirable side effect profile and improvement in renal function compared with calcineurin inhibition in renal transplantation. This comes at the cost of increased rates of early acute rejection. Blockade of the integrin molecule leukocyte function-associated antigen 1 (LFA-1) has been shown to be an effective adjuvant to costimulation blockade in a rigorous nonhuman primate (NHP) model of islet transplantation; therefore, we sought to test this combination in an NHP renal transplant model. Rhesus macaques received belatacept maintenance therapy with or without the addition of LFA-1 blockade, which was achieved using a murine-derived LFA-1-specific antibody TS1/22. Additional experiments were performed using chimeric rhesus IgG1 (TS1/22R1) or IgG4 (TS1/22R4) variants, each engineered to limit antibody clearance. Despite evidence of proper binding to the target molecule and impaired cellular egress from the intravascular space indicative of a therapeutic effect similar to prior islet studies, LFA-1 blockade failed to significantly prolong graft survival. Furthermore, evidence of impaired protective immunity against cytomegalovirus was observed. These data highlight the difficulties in translating treatment regimens between organ models and suggest that the primarily vascularized renal model is more robust with regard to belatacept-resistant rejection than the islet model.

A pilot trial targeting the ICOS-ICOS-L pathway in nonhuman primate kidney transplantation.

Costimulation blockade with the B7-CD28 pathway-specific agent belatacept is now used in clinical kidney transplantation, but its efficacy remains imperfect. Numerous alternate costimulatory pathways have been proposed as targets to synergize with belatacept, one of which being the inducible costimulator (ICOS)-ICOS ligand (ICOS-L) pathway. Combined ICOS-ICOS-L and CD28-B7 blockade has been shown to prevent rejection in mice, but has not been studied in primates. We therefore tested a novel ICOS-Ig human Fc-fusion protein in a nonhuman primate (NHP) kidney transplant model alone and in combination with belatacept. ICOS-Ig did not prolong rejection-free survival as a monotherapy or in combination with belatacept. In ICOS-Ig alone treated animals, most graft-infiltrating CD4(+) and CD8(+) T cells expressed ICOS, and ICOS(+) T cells were present in peripheral blood to a lesser degree. Adding belatacept reduced the proportion of graft-infiltrating ICOS(+) T cells and virtually eliminated their presence in peripheral blood. Graft-infiltrating T cells in belatacept-resistant rejection were primarily CD8(+) CD28(-) , but importantly, very few CD8(+) CD28(-) T cells expressed ICOS. We conclude that ICOS-Ig, alone or combined with belatacept, does not prolong renal allograft survival in NHPs. This may relate to selective loss of ICOS with CD28 loss.

Prolonged expression of an anti-HIV-1 gp120 minibody to the female rhesus macaque lower genital tract by AAV gene transfer.

Topical microbicides are a leading strategy for prevention of HIV mucosal infection to women; however, numerous pharmacokinetic limitations associated with coitally related dosing strategy have contributed to their limited success. Here we test the hypothesis that adeno-associated virus (AAV) mediated delivery of the b12 human anti-HIV-1 gp120 minibody gene to the lower genital tract of female rhesus macaques (Rh) can provide prolonged expression of b12 minibodies in the cervical-vaginal secretions. Gene transfer studies demonstrated that, of various green fluorescent protein (GFP)-expressing AAV serotypes, AAV-6 most efficiently transduced freshly immortalized and primary genital epithelial cells (PGECs) of female Rh in vitro. In addition, AAV-6-b12 minibody transduction of Rh PGECs led to inhibition of SHIV162p4 transmigration and virus infectivity in vitro. AAV-6-GFP could also successfully transduce vaginal epithelial cells of Rh when applied intravaginally, including p63+ epithelial stem cells. Moreover, intravaginal application of AAV-6-b12 to female Rh resulted in prolonged minibody detection in their vaginal secretions throughout the 79-day study period. These data provide proof of principle that AAV-6-mediated delivery of anti-HIV broadly neutralizing antibody (BnAb) genes to the lower genital tract of female Rh results in persistent minibody detection for several months. This strategy offers promise that an anti-HIV-1 genetic microbicide strategy may be possible in which topical application of AAV vector, with periodic reapplication as needed, may provide sustained local BnAb expression and protection.

Perspectives on global nursing leadership: international experiences from the field.

AIM: Nursing leaders from six countries engaged in a year-long discussion on global leadership development. The purpose of these dialogues was to strengthen individual and collective capacity as nursing leaders in a global society. Field experiences in practice and education were shared. Perspectives on global leadership can strengthen nurses' contributions to practice, workplace and policy issues worldwide. BACKGROUND: Transformational leadership empowers nurses' increasing confidence. Mentoring is needed to stimulate leadership development but this is lacking in many settings where nurses practice, teach and influence policy. Organizations with global mission provide opportunity for nurses' professional growth in leadership through international dialogues. PROCEDURES: Dialogues among participants were held monthly by conference calls or videoconferences. Example stories from each participant illustrated nursing leadership in action. From these exemplars, concepts were chosen to create a framework. Emerging perspectives and leadership themes represented all contexts of practice, education, research and policy. The cultural context of each country was reflected in the examples. RESULTS: Themes emerged that crossed global regions and countries. Themes were creativity, change, collaboration, community, context and courage. IMPLICATIONS FOR NURSING AND HEALTH POLICY: Relationships initially formed in professional organizations can be extended to intentionally facilitate global nursing leadership development. Exemplars from the dialogues demonstrated nursing leadership in health policy development within each cultural context. Recommendations are given for infrastructure development in organizations to enhance future collaborations.

Inhibition of αvß6 promotes acute renal allograft rejection in nonhuman primates.

The integrin αvß6 activates latent transforming growth factor-ß (TGF-ß) within the kidney and may be a target for the prevention of chronic allograft fibrosis after kidney transplantation. However, TGF-ß also has known immunosuppressive properties that are exploited by calcineurin inhibitors (CNIs); thus, the net benefit of αvß6 inhibition remains undetermined. To assess the acute impact of interference with αvß6 on acute rejection, we tested a humanized αvß6-specific monoclonal antibody (STX-100) in a randomized, double-blinded, placebo-controlled nonhuman primate renal transplantation study to evaluate whether αvß6 blockade alters the risk of acute rejection during CNI-based immunosuppression. Rhesus monkeys underwent renal allotransplantation under standard CNI-based maintenance immunosuppression; 10 biopsy-confirmed rejection-free animals were randomized to receive weekly STX-100 or placebo. Animals treated with STX-100 experienced significantly decreased rejection-free survival compared to placebo animals (p = 0.049). Immunohistochemical analysis confirmed αvß6 ligand presence, and αvß6 staining intensity was lower in STX-100-treated animals (p = 0.055), indicating an apparent blockade effect of STX-100. LAP, LTBP-1 and TGF-ß were all decreased in animals that rejected on STX-100 compared to those that rejected on standard immunosuppression alone, suggesting a relevant effect of αvß6 blockade on local TGF-ß. These data caution against the use of αvß6 blockade to achieve TGF-ß inhibition in kidney transplantation.

Belatacept and sirolimus prolong nonhuman primate renal allograft survival without a requirement for memory T cell depletion.

Belatacept is an inhibitor of CD28/B7 costimulation that is clinically indicated as a calcineurin inhibitor (CNI) alternative in combination with mycophenolate mofetil and steroids after renal transplantation. We sought to develop a clinically translatable, nonlymphocyte depleting, belatacept-based regimen that could obviate the need for both CNIs and steroids. Thus, based on murine data showing synergy between costimulation blockade and mTOR inhibition, we studied rhesus monkeys undergoing MHC-mismatched renal allotransplants treated with belatacept and the mTOR inhibitor, sirolimus. To extend prior work on costimulation blockade-resistant rejection, some animals also received CD2 blockade with alefacept (LFA3-Ig). Belatacept and sirolimus therapy successfully prevented rejection in all animals. Tolerance was not induced, as animals rejected after withdrawal of therapy. The regimen did not deplete T cells. Alefecept did not add a survival benefit to the optimized belatacept and sirolimus regimen, despite causing an intended depletion of memory T cells, and caused a marked reduction in regulatory T cells. Furthermore, alefacept-treated animals had a significantly increased incidence of CMV reactivation, suggesting that this combination overly compromised protective immunity. These data support belatacept and sirolimus as a clinically translatable, nondepleting, CNI-free, steroid-sparing immunomodulatory regimen that promotes sustained rejection-free allograft survival after renal transplantation.

NRSF/REST is required in vivo for repression of multiple neuronal target genes during embryogenesis.

The neuron-restrictive silencer factor NRSF (also known as REST and XBR) can silence transcription from neuronal promoters in non-neuronal cell lines, but its function during normal development is unknown. In mice, a targeted mutation of Rest, the gene encoding NRSF, caused derepression of neuron-specific tubulin in a subset of non-neural tissues and embryonic lethality. Mosaic inhibition of NRSF in chicken embryos, using a dominant-negative form of NRSF, also caused derepression of neuronal tubulin, as well as of several other neuronal target genes, in both non-neural tissues and central nervous system neuronal progenitors. These results indicate that NRSF is required to repress neuronal gene expression in vivo, in both extra-neural and undifferentiated neural tissue.

Genomic imprinting of Mash2, a mouse gene required for trophoblast development.

The mouse gene Mash2 encodes a transcription factor required for development of trophoblast progenitors. Mash2-homozygous mutant embryos die at 10 days postcoitum from placental failure. Here we show that Mash2 is genomically imprinted. First, Mash2+/- embryos inheriting a wild-type allele from their father die at the same stage as -/- embryos, with a similar placental phenotype. Second, the Mash2 paternal allele is initially expressed by groups of trophoblast cells at 6.5 and 7.5 days post-coitum, but appears almost completely repressed by 8.5 days post-coitum. Finally, we have genetically and physically mapped Mash2 to the distal region of chromosome 7, within a cluster of imprinted genes, including insulin-2, insulin-like growth factor-2 and H19.

Protocol of trans-Tasman feasibility randomised controlled trial of the Younger Women's Wellness After Breast Cancer (YWWACP) lifestyle intervention.

BACKGROUND: Younger women (defined as those < 50 years who are likely pre-menopausal at time of diagnosis) with breast cancer often experience persistent treatment-related side effects that adversely affect their physical and psychological wellbeing. The Women's Wellness After Cancer Program (WWACP) was adapted and piloted in Australia to address these outcomes in younger women. The aims of this feasibility study are to determine (1) the potential to translate the Younger WWACP (YWWACP) intervention to a broader population base in Aotearoa/New Zealand and Australia, and (2) the potential for success of a larger, international, phase ΙΙΙ, randomised controlled trial. METHODS: This bi-national, randomised, single-blinded controlled trial involves two main study sites in Aotearoa/New Zealand (Kowhai study) and Australia (EMERALD study). Young women aged 18 to 50 years who completed intensive treatment (surgery, chemotherapy, and/or radiotherapy) for breast cancer in the previous 24 months are eligible. The potential to translate the YWWACP to women in these two populations will be assessed according to several feasibility outcomes. These include examining intervention accessibility, acceptability and uptake; intervention sustainability and adherence; the prevalence components of the intervention in the control group; intervention efficacy; participants' perception of measurement burden; the effectiveness of planned recruitment strategies; and trial methods and procedures. The studies collectively aim to enrol 60 participants in the intervention group and 60 participants in the control group (total = 120 participants). DISCUSSION: Ethical approval has been received from the Southern Health and Disability Ethics Committee (Kowhai ref: 19/STH/215), and UnitingCare Human Research Ethics Committee (EMERALD ref: 202103). This study will provide important data on the feasibility of the refined YWWACP in the trans-Tasman context. This study will account for and harmonise cross-country differences to ensure the success of a proposed international grant application for a phase ΙΙΙ randomised controlled trial of this program to improve outcomes in younger women living with breast cancer. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry (ANZCTR): Kowhai ACTRN12620000260921 , registered on 27 February 2020. EMERALD ACTRN12621000447853 , registered on 19 April 2021.

Mechanisms of global diversification in the brown booby (Sula leucogaster) revealed by uniting statistical phylogeographic and multilocus phylogenetic methods.

Recent theoretical and empirical research suggests that statistical models based on coalescent theory can improve both phylogeographic and phylogenetic inference. An approach that involves elements of both statistical phylogeography (e.g. Isolation with Migration analyses) and multilocus phylogenetic inference (e.g. *beast) may be particularly useful when applied to populations with relatively old divergence times. Here, we use such an approach in the globally distributed brown booby (Sula leucogaster). We sampled 215 individuals from all major breeding areas and genotyped them at eight microsatellite and three nuclear intron loci. We found that brown booby populations were highly differentiated and that colonies can be grouped into four major genetic populations (Caribbean Sea, Central Atlantic Ocean, Indo-Central Pacific and Eastern Pacific). These populations apparently diverged in the absence of gene flow and, with one exception, currently exchange few to no migrants. The Eastern Pacific population diverged from all other populations approximately one million years ago [90% highest posterior density: 330,000-2,000,000 years ago] and exhibits a distinct male plumage, relative to other populations. However, recent gene flow from the Indo-Central Pacific into the Eastern Pacific appears to have occurred, suggesting that approximately one million years of genetic isolation and divergence in male plumage colour are not sufficient to prevent interbreeding. Gene flow following secondary contact of the Indo-Central Pacific and Eastern Pacific populations was not detected in previous mitochondrial DNA (mtDNA) studies, and the contrast between the mtDNA results and our current results highlights the advantage of a multilocus phylogeographic approach.

Transtracheal jet ventilation in 50 patients with severe airway compromise and stridor.

BACKGROUND: Management of the airway is difficult in patients with pharyngeal or laryngeal pathology caused by malignancy, extensive surgery, or radiotherapy scarring, particularly when undergoing pharyngolaryngeal surgery. Tracheal intubation, with or without fibreoptic guidance, is often impractical because of the severe glottic stenosis and primary tracheostomy under local anaesthesia has been the preferred technique. However, complication rates as high as 30% have been reported after primary tracheostomy and there is the potential for long-term morbidity. High-frequency jet ventilation (HFJV) has several advantages over other techniques in the management of the difficult airway and can be delivered by supraglottic and infraglottic routes. To date, no large series has described the use of transtracheal HFJV (TTHFJV) in adult patients with stridor and critical airway obstruction. METHODS: We report a prospective, descriptive audit of the safe use of TTHFJV in patients with severe airway compromise and stridor undergoing pharyngolaryngeal surgery (50 consecutive procedures in 44 patients). RESULTS: TTHFJV was successful in all 50 cases. There were no major complications and the incidence of minor complications was 20% with no subsequent morbidity. CONCLUSIONS: We attribute this low incidence to the use of an automated jet ventilator with airway pressure monitoring and control, and the alteration of ventilator parameters by an experienced anaesthetist.

Randomized controlled trial of the A.P. Advance, McGrath, and Macintosh laryngoscopes in normal and difficult intubation scenarios: a manikin study.

BACKGROUND: Several videolaryngoscopes are available which may facilitate tracheal intubation in difficult airways. We compared the McGrath(®) Series 5 and the Venner™ A.P. Advance™ (APA) videolaryngoscopes with a Macintosh laryngoscope by studying the performance of experienced anaesthetists using manikins in normal and difficult airway scenarios. METHODS: We recruited 48 anaesthetists into a randomized trial. Each performed tracheal intubation with each laryngoscope in one easy and one difficult laryngoscopy scenario. The primary endpoint was time to intubation. Other endpoints were time to best glottic visualization, grade of view, and number of glottic advances. RESULTS: There were no dropouts. In the easy scenario, the time to intubation was greater using the McGrath [median time 40.7 s, inter-quartile range (IQR) 31.0, 57.4, P<0.001] than the other devices. In the difficult scenario, the time to intubation using the APA with Difficult Airway Blade (DAB) was less (median time 23.2 s, IQR 19.8, 29.0, P<0.001) than the other devices. Time to glottic visualization was reduced using the McGrath and the APA with DAB. Glottic advances were fewer using the APA with DAB. CONCLUSIONS: Experienced anaesthetists required a longer time for intubation in a standard manikin using a McGrath compared with other laryngoscopes, but a shorter time for intubation in a difficult manikin using an APA with DAB, and with fewer glottic advances, compared with other laryngoscopes.

Younger and older women's concerns about menopause after breast cancer.

A number of treatments for breast cancer induce menopause. This study's aim was to explore women's perceptions and beliefs about menopausal symptoms and their management following breast cancer, and to compare younger and older women's experiences. Data were collected via semi-structured focus groups from women who had undergone treatment for breast cancer, and who were currently experiencing menopausal symptoms. Data were interpreted by way of simple inductive thematic analysis. The women experienced a range of menopausal symptoms that they were not prepared for and found difficult to manage. The central themes related to their lack of knowledge of how to manage menopausal symptoms, and the distress and helplessness that arose from this. Women who were diagnosed prior to 40 years of age reported additional menopausal issues than women who were older at diagnosis. The women in this study expressed a thirst for information related to menopause after breast cancer. The women identified that their needs with regard to menopause after breast cancer were not being met, either through their own lack of knowledge or via conflicting or absent support and management. The importance of enabling women to deal with menopausal symptoms was a central theme to emerge from the data.

Comparative phylogeography of brown (Sula leucogaster) and red-footed boobies (S. sula): the influence of physical barriers and habitat preference on gene flow in pelagic seabirds.

To test the hypothesis that both physical and ecological barriers to gene flow drive population differentiation in tropical seabirds, we surveyed mitochondrial control region variation in 242 brown boobies (Sula leucogaster), which prefer inshore habitat, and 271 red-footed boobies (S. sula), which prefer pelagic habitat. To determine the relative influence of isolation and gene flow on population structure, we used both traditional methods and a recently developed statistical method based on coalescent theory and Bayesian inference (Isolation with Migration). We found that global population genetic structure was high in both species, and that female-mediated gene flow among ocean basins apparently has been restricted by major physical barriers including the Isthmus of Panama, and the periodic emergence of the Sunda and Sahul Shelves in Southeast Asia. In contrast, the evolutionary history of populations within ocean basins differed markedly between the two species. In brown boobies, we found high levels of population genetic differentiation and limited gene flow among colonies, even at spatial scales as small as 500km. Although red-footed booby colonies were also genetically differentiated within ocean basins, coalescent analyses indicated that populations have either diverged in the face of ongoing gene flow, or diverged without gene flow but recently made secondary contact. Regardless, gene flow among red-footed booby populations was higher than among brown booby populations. We suggest that these contrasting patterns of gene flow within ocean basins may be explained by the different habitat preferences of brown and red-footed boobies.

Liquid biopsy for cancer diagnosis using vibrational spectroscopy: systematic review.

BACKGROUND: Vibrational spectroscopy (VS) is a minimally invasive tool for analysing biological material to detect disease. This study aimed to review its application to human blood for cancer diagnosis. METHODS: A systematic review was undertaken using a keyword electronic database search (MEDLINE, Embase, PubMed, TRIP and Cochrane Library), with all original English-language manuscripts examining the use of vibrational spectral analysis of human blood for cancer detection. Studies involving fewer than 75 patients in the cancer or control group, animal studies, or where the primary analyte was not blood were excluded. RESULTS: From 1446 results, six studies (published in 2010-2018) examining brain, bladder, oral, breast, oesophageal and hepatic cancer met the criteria for inclusion, with a total population of 2392 (1316 cancer, 1076 control; 1476 men, 916 women). For cancer detection, reported mean sensitivities in each included study ranged from 79·3 to 98 per cent, with specificities of 82·8-95 per cent and accuracies between 81·1 and 97·1 per cent. Heterogeneity in reporting strategies, methods and outcome measures made meta-analysis inappropriate. CONCLUSION: VS shows high potential for cancer diagnosis, but until there is agreement on uniform standard reporting methods and studies with adequate sample size for valid classification models have been performed, its value in clinical practice will remain uncertain.

la espectroscopia vibracional (vibrational spectroscopy, VS) es un dispositivo mínimamente invasivo para analizar material biológico y detectar enfermedad. Este estudio se propuso revisar su aplicación en la sangre humana para el diagnóstico de cáncer. MÉTODOS: Se llevó a cabo una revisión sistemática utilizando una búsqueda con palabras claves en bases de datos electrónicas (MEDLINE, EMBASE, PubMed, TRIP, Cochrane Library), de todos los manuscritos originales publicados en inglés que examinaban la utilización del análisis espectral vibracional de la sangre humana para la detección del cáncer. Se excluyeron estudios que incluían menos de 75 pacientes en los grupos cáncer/control, estudios en animales o cuando la muestra principal no fuera la sangre. RESULTADOS: De los 1.446 resultados, 6 estudios publicados en 2010-2018, y que examinaban cánceres del cerebro, vejiga, oral, mama, esófago e hígado cumplieron los criterios de inclusión, con una población total de 2.506 casos (1.316 cánceres, 1.076 controles; 1.476 varones, 915 mujeres). Para la detección del cáncer, las sensibilidades medias descritas en cada uno de los estudios incluidos variaron entre 79,3-98,0%, con especificidades entre 82,8-95,0%, y exactitudes diagnósticas entre 81,1% y 97,1%. La heterogeneidad en las estrategias descritas, métodos y medidas de resultados hicieron que el metaanálisis fuera inapropiado. CONCLUSIÓN: La VS muestra un alto potencial para el diagnóstico del cáncer, pero hasta que no se llegue a un acuerdo en los métodos para informar de los resultados y se hayan efectuado estudios con un tamaño muestral adecuado para una clasificación válida de los modelos, su valor en la práctica clínica sigue siendo incierto.

Signal recognition protein is required for the integration of acetylcholine receptor delta subunit, a transmembrane glycoprotein, into the endoplasmic reticulum membrane.

Purified Signal Recognition Protein (SRP) has previously been shown to be required for the translocation of secretory proteins across the microsomal membrane (Walter and Blobel, 1980. Proc. Natl. Acad. Sci. U. S. A. 77:7, 112-7, 116) and to function in the early events of this process (Walter and Blobel, 1981. J. Cell Biol. 91:557-561). We demonstrate here that the delta subunit of acetylcholine receptor (AChR-delta), a transmembrane glycoprotein, likewise requires SRP for its asymmetric integration into microsomal membranes. We further demonstrate by partial sequence analysis that AChR-delta is synthesized with a transient NH2-terminal signal sequence of 21 residues that is cleaved off during integration into microsomal membranes. Integration of AChR-delta into the microsomal membrane vesicles proceeded asymmetrically, yielding a large (44 kdalton) core-glycosylated domain, inaccessible to externally added proteolytic enzymes and a smaller (approximately 16 kdalton) domain exposed on the outside of the vesicles and accessible to externally added proteolytic enzymes. The NH2 terminus of the molecule is contained in the 44-kdalton domain.

Comparative analysis of the major polypeptides from liver gap junctions and lens fiber junctions.

Gap junctions from rat liver and fiber junctions from bovine lens have similar septilaminar profiles when examined by thin-section electron microscopy and differ only slightly with respect to the packing of intramembrane particles in freeze-fracture images. These similarities have often led to lens fiber junctions being referred to as gap junctions. Junctions from both sources were isolated as enriched subcellular fractions and their major polypeptide components compared biochemically and immunochemically. The major liver gap junction polypeptide has an apparent molecular weight of 27,000, while a 25,000-dalton polypeptide is the major component of lens fiber junctions. The two polypeptides are not homologous when compared by partial peptide mapping in SDS. In addition, there is not detectable antigenic similarity between the two polypeptides by immunochemical criteria using antibodies to the 25,000-dalton lens fiber junction polypeptide. Thus, in spite of the ultrastructural similarities, the gap junction and the lens fiber junction are comprised of distinctly different polypeptides, suggesting that the lens fiber junction contains a unique gene product and potentially different physiological properties.