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This article addresses the emergence of children and young people as simultaneously sexual/gender non-normative and political subjects during the 2016 Safe Schools controversy. What appeared as a s...

A myriad of transient, nanoscopic lipid- and protein-based interactions confer a steady-state organization of plasma membrane in resting cells that is poised to orchestrate assembly of key signaling components upon reception of an extracellular stimulus. Although difficult to observe directly in live cells, these subtle interactions can be discerned by their impact on the diffusion of membrane constituents. Herein, we quantified the diffusion properties of a panel of structurally distinct lipid-anchored and transmembrane (TM) probes in RBL mast cells by multiplexed Imaging Fluorescence Correlation Spectroscopy. We developed a statistical analysis of data combined from many pixels over multiple cells to characterize differences as small as 10% in diffusion coefficients, which reflect differences in underlying interactions. We found that the distinctive diffusion properties of lipid-anchored probes can be explained by their dynamic partitioning into ordered proteo-lipid nanodomains, which encompass a major fraction of the membrane and whose physical properties are influenced by actin polymerization. Effects on diffusion by functional protein modules in both lipid-anchored and TM probes reflect additional complexity in steady-state membrane organization. The contrast we observe between different probes diffusing through the same membrane milieu represent the dynamic resting steady-state, which serves as a baseline for monitoring plasma membrane remodeling that occurs upon stimulation.

Phosphatidylinositol 4-kinase IIIα (PI4KIIIα) is the major enzyme responsible for generating phosphatidylinositol (4)-phosphate [PI(4)P] at the plasma membrane. This lipid kinase forms two multicomponent complexes, both including a palmitoylated anchor, EFR3. Whereas both PI4KIIIα complexes support production of PI(4)P, the distinct functions of each complex and mechanisms underlying the interplay between them remain unknown. Here, we present roles for differential palmitoylation patterns within a tri-cysteine motif in EFR3B (Cys5, Cys7 and Cys8) in controlling the distribution of PI4KIIIα between these two complexes at the plasma membrane and corresponding functions in phosphoinositide homeostasis. Spacing of palmitoyl groups within three doubly palmitoylated EFR3B ‘lipoforms’ affects both interactions between EFR3B and TMEM150A, a transmembrane protein governing formation of a PI4KIIIα complex functioning in rapid phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P2] resynthesis following phospholipase C signaling, and EFR3B partitioning within liquid-ordered and -disordered regions of the plasma membrane. This work identifies a palmitoylation code involved in controlling protein–protein and protein–lipid interactions that affect a plasma membrane-resident lipid biosynthetic pathway.

Antigen (Ag) crosslinking of immunoglobulin E-receptor (IgE-FceRI) complexes in mast cells stimulates transmembrane (TM) signaling, requiring phosphorylation of the clustered FceRI by lipid-anchored Lyn tyrosine kinase. Previous studies showed that this stimulated coupling between Lyn and FceRI occurs in liquid ordered (Lo)-like nanodomains of the plasma membrane and that Lyn binds directly to cytosolic segments of FceRI that it initially phosphorylates for amplified activity. Net phosphorylation above a nonfunctional threshold is achieved in the stimulated state but not in the resting state, and current evidence supports the hypothesis that this relies on Ag crosslinking to disrupt a balance between Lyn and tyrosine phosphatase activities. However, the structural interactions that underlie the stimulation process remain poorly defined. This study evaluates the relative contributions and functional importance of different types of interactions leading to suprathreshold phosphorylation of Ag-crosslinked IgE-FceRI in live rat basophilic leukemia mast cells. Our high-precision diffusion measurements by imaging fluorescence correlation spectroscopy on multiple structural variants of Lyn and other lipid-anchored probes confirm subtle, stimulated stabilization of the Lo-like nanodomains in the membrane inner leaflet and concomitant sharpening of segregation from liquid disordered (Ld)-like regions. With other structural variants, we determine that lipid-based interactions are essential for access by Lyn, leading to phosphorylation of and protein-based binding to clustered FceRI. By contrast, TM tyrosine phosphatase, PTPα, is excluded from these regions due to its Ld-preference and steric exclusion of TM segments. Overall, we establish a synergy of lipid-based, protein-based, and steric interactions underlying functional TM signaling in mast cells.

PI4KIIIα is the major enzyme responsible for generating the phosphoinositide PI(4)P at the plasma membrane. This lipid kinase forms two multicomponent complexes, both including a palmitoylated anchor, EFR3. Whereas both PI4KIIIα complexes support production of PI(4)P, the distinct functions of each complex and mechanisms underlying the interplay between them remain unknown. Here, we present roles for differential palmitoylation patterns within a tri-Cys motif in EFR3B (Cys5/Cys7/Cys8) in controlling the distribution of PI4KIIIα between these two complexes at the plasma membrane and corresponding functions in phosphoinositide homeostasis. Spacing of palmitoyl groups within three doubly palmitoylated EFR3B “lipoforms” affects both its interactions with TMEM150A, a transmembrane protein governing formation of a PI4KIIIα complex functioning in rapid PI(4,5)P2 resynthesis following PLC signaling, and its partitioning within liquid-ordered and -disordered regions of the plasma membrane. This work identifies a palmitoylation code in controlling protein–protein and protein–lipid interactions affecting a plasma membrane-resident lipid biosynthetic pathway.SUMMARY STATEMENTDifferent palmitoylation patterns on a lipid kinase adaptor protein control partitioning of the kinase between two spatiotemporally and functionally distinct complexes within the plasma membrane.

A large body of feminist scholarship identifies and examines modes of representing abortion, focusing on themes such as stigma and the production of an autonomous foetal subject. In recent years, there has been a proliferation of cultural representations of abortion that do not conform with, and even explicitly reference and rebuke, stigmatising and foetocentric depictions of abortion. Focusing on the Australian context and two sets of online sites—the websites of Australian abortion clinics and widely circulated pro-choice, feminist commentary on abortion—this article provides a critical reading of new, ‘unapologetic’ modes of representing abortion. We argue that they increase the cultural legitimacy of abortion and are therefore incredibly important, but have limitations because of their reification of the neoliberal autonomous subject. Nevertheless, we urge feminist scholars to pay more attention to discourses that presume abortion is unproblematic and beneficial to women. Scholarship on abortion has performative effects, and we hope that identifying and nurturing counter narratives of abortion is one means by which abortion can be normalised.

UNSTRUCTURED Safety issues for researchers conducting and disseminating research on social media have been inadequately addressed in institutional policies and practice globally, despite posing significant challenges to research staff and student well-being. In the context of the COVID-19 pandemic and given the myriad of advantages that web-based platforms offer researchers over traditional recruitment, data collection, and research dissemination methods, developing a comprehensive understanding of and guidance on the safe and effective conduct of research in web-based spaces has never been more pertinent. In this paper, we share our experience of using social media to recruit participants for a study on abortion stigma in Australia, which brought into focus the personal, professional, and institutional risks associated with conducting web-based research that goes viral. The lead researcher (KV), a postgraduate student, experienced a barrage of harassment on and beyond social media. The supportive yet uncoordinated institutional response highlighted gaps in practice, guidance, and policy relating to social media research ethics, researcher safety and well-being, planning for and managing web-based and offline risk, and coordinated organizational responses to adverse events. We call for and provide suggestions to inform the development of training, guidelines, and policies that address practical and ethical aspects of using social media for research, mental and physical health and safety risks and management, and the development of coordinated and evidence-based institutional- and individual-level responses to cyberbullying and harassment. Furthermore, we argue the case for the urgent development of this comprehensive guidance around researcher safety on the web, which would help to ensure that universities have the capacity to maximize the potential of social media for research while better supporting the well-being of their staff and students.

Antigen (Ag) crosslinking of immunoglobulin E-receptor (IgE-FcεRI) complexes in mast cells stimulates transmembrane (TM) signaling, requiring phosphorylation of the clustered FcεRI by lipid-anchored Lyn tyrosine kinase. Previous studies showed that this stimulated coupling between Lyn and FcεRI occurs in liquid ordered (Lo)-like nanodomains of the plasma membrane and that Lyn binds directly to cytosolic segments of FcεRI that it initially phosphorylates for amplified activity. Net phosphorylation above a non-functional threshold is achieved in the stimulated state, but not in the resting state, and current evidence supports the hypothesis that this relies on disruption by Ag-crosslinking of a balance between Lyn and tyrosine phosphatase activities. However, the structural interactions that underlie the stimulation process remain poorly defined. This study evaluates the relative contributions and functional importance of different types of interactions leading to supra-threshold phosphorylation of Ag-crosslinked IgE-FcεRI in live rat basophilic leukemia (RBL) mast cells. Our high-precision diffusion measurements by Imaging Fluorescence Correlation Spectroscopy (ImFCS) on multiple structural variants of Lyn and other lipid-anchored probes confirm subtle, stimulated stabilization of the Lo-like nanodomains and concomitant sharpening of segregation from liquid-disordered (Ld)-like regions. With other structural variants we determine that lipid-based interactions are essential for access by Lyn leading to phosphorylation of and protein-based binding to clustered FcεRI. By contrast, TM tyrosine phosphatase, PTPα, is excluded from these regions by steric repulsion of TM segments and preference for Ld-like regions. Overall, we establish a synergy of lipid-based, protein-based, and steric interactions underlying functional TM signaling in mast cells.SIGNIFICANCE STATEMENTLipid organization of the plasma membrane is known to be important for facilitating protein interactions in transmembrane signaling. However, the orchestration of these interactions in live cells has been elusive. We employed ImFCS to systemically investigate the interplay of lipids and proteins during signaling in mast cells, initiated as phosphorylation of Ag-crosslinked IgE-FcεRI by lipid-anchored Lyn kinase. We find lipid-based interactions are first required for protein-based phosphorylation of the clustered FcεRI within Lo-like nanodomains. Transmembrane phosphatases must be excluded from these regions, and we find this is mediated by their preference for Ld-like regions and by steric exclusion from the clustered FcεRI proteins. ImFCS provides quantitative characterization of the functional link between features of plasma membrane organization and transmembrane signaling.

Epidermal growth factor receptor (EGFR) dysregulation is observed in many human cancers and is both a cause of oncogenesis and a target for chemotherapy. We previously showed that partial charge neutralization of the juxtamembrane (JX) region of EGFR via the EGFR R1-6 mutant construct induces constitutive receptor activation and transformation of NIH 3T3 cells, both from the plasma membrane and from the ER when combined with the ER-retaining L417H mutation (Bryant et al. in J Biol Chem 288:34930-34942, 2013). Here, we use chemical crosslinking and immunoblotting to show that these mutant constructs form constitutive, phosphorylated dimers in both the plasma membrane and the ER. Furthermore, we combine this electrostatic perturbation with conformationally-restricted receptor mutants to provide evidence that activation of EGFR R1-6 dimers requires functional coupling both between the EGFR extracellular dimerization arms and between intracellular tyrosine kinase domains. These findings provide evidence that the electrostatic charge of the JX region normally serves as a negative regulator of functional dimerization of EGFR.

Stochastic optical reconstruction microscopy (STORM) is an optical super-resolution microscopy (SRM) technique that traditionally requires toxic and non-physiological imaging buffers and setups that are not conducive to live-cell studies. It is observed that ultrasmall (

Significance Membrane phase behavior in cells permits transient concentration of specific proteins and lipids into dynamic nanoscopic domains. Here, we tested the existence and role of such phase behavior in endoplasmic reticulum (ER) membranes. Employing hypotonic cell swelling, we created large intracellular vesicles (LICVs) from internal organelles. ER LICVs maintained stable interorganelle contacts, with known protein tethers concentrated at the contact sites. Cooled ER LICVs underwent reversible phase separation into microscopically visible domains with different lipid order and membrane fluidity. The phase-separated domains specified sites of contact between the ER and different organelles., The endoplasmic reticulum (ER) is the site of synthesis of secretory and membrane proteins and contacts every organelle of the cell, exchanging lipids and metabolites in a highly regulated manner. How the ER spatially segregates its numerous and diverse functions, including positioning nanoscopic contact sites with other organelles, is unclear. We demonstrate that hypotonic swelling of cells converts the ER and other membrane-bound organelles into micrometer-scale large intracellular vesicles (LICVs) that retain luminal protein content and maintain contact sites with each other through localized organelle tethers. Upon cooling, ER-derived LICVs phase-partition into microscopic domains having different lipid-ordering characteristics, which is reversible upon warming. Ordered ER lipid domains mark contact sites with ER and mitochondria, lipid droplets, endosomes, or plasma membrane, whereas disordered ER lipid domains mark contact sites with lysosomes or peroxisomes. Tethering proteins concentrate at ER–organelle contact sites, allowing time-dependent behavior of lipids and proteins to be studied at these sites. These findings demonstrate that LICVs provide a useful model system for studying the phase behavior and interactive properties of organelles in intact cells.

This article begins a history of twenty-first century LGBTI activism in Australia. It writes a history of the marriage equality campaign in Australia, which began in 2004 and achieved success in 20...

In 1973, the Northern Territory (NT) criminal law relating to abortion was reformed. The NT was one of only two Australian jurisdictions where the 1970s liberalisation of abortion was enabled by le...

This article offers a historical account of the changing politics of abortion in Australia in the late 1960s and early-mid 1970s. The article first focuses on women’s personal stories of se...

Safety issues for researchers conducting and disseminating research on social media have been inadequately addressed in institutional policies and practice globally, despite posing significant challenges to research staff and student well-being. In the context of the COVID-19 pandemic and given the myriad of advantages that web-based platforms offer researchers over traditional recruitment, data collection, and research dissemination methods, developing a comprehensive understanding of and guidance on the safe and effective conduct of research in web-based spaces has never been more pertinent. In this paper, we share our experience of using social media to recruit participants for a study on abortion stigma in Australia, which brought into focus the personal, professional, and institutional risks associated with conducting web-based research that goes viral. The lead researcher (KV), a postgraduate student, experienced a barrage of harassment on and beyond social media. The supportive yet uncoordinated institutional response highlighted gaps in practice, guidance, and policy relating to social media research ethics, researcher safety and well-being, planning for and managing web-based and offline risk, and coordinated organizational responses to adverse events. We call for and provide suggestions to inform the development of training, guidelines, and policies that address practical and ethical aspects of using social media for research, mental and physical health and safety risks and management, and the development of coordinated and evidence-based institutional- and individual-level responses to cyberbullying and harassment. Furthermore, we argue the case for the urgent development of this comprehensive guidance around researcher safety on the web, which would help to ensure that universities have the capacity to maximize the potential of social media for research while better supporting the well-being of their staff and students.

The Marching Dunstans were a group of over 100 people in the annual Pride March in Adelaide, South Australia, in 2015, the 40th anniversary of the decriminalisation of homosexuality in this state. ...

Surface-patterned ligands reveal spatially targeted signaling events mediated by FcεRI on mast cells. Distinctively clustered IgE-FcεRI recruits signaling proteins, including Syk, LAT, and activated PLCγ1. β1- and β3-integrins also colocalize at patterned sites, involving actin polymerization and stimulated trafficking from recycling endosomes., We examined the spatial targeting of early and downstream signaling mediated by the immunoglobulin E (IgE) receptor (FcεRI) in RBL mast cells using surface-patterned 2,4-dinitrophenyl (DNP) ligands. Micron-sized features of DNP are presented as densely immobilized conjugates of bovine serum albumin (DNP-BSA) or mobile in a supported lipid bilayer (DNP-SLB). Although soluble anti-DNP IgE binds uniformly across features for both pattern types, IgE bound to FcεRI on cells shows distinctive distributions: uniform for DNP-SLB and edge concentrated for DNP-BSA. These distributions of IgE-FcεRI propagate to the spatial recruitment of early signaling proteins, including spleen tyrosine kinase (Syk), linker for activation of T-cells (LAT), and activated phospholipase C gamma 1 (PLCγ1), which all localize with engaged receptors. We found stimulated polymerization of F-actin is not required for Syk recruitment but is progressively involved in the recruitment of LAT and PLCγ1. We further found β1- and β3-integrins colocalize with IgE-FcεRI at patterned ligand surfaces as cells spread. This recruitment corresponds to directed exocytosis of recycling endosomes (REs) containing these integrins and their fibronectin ligand. Together our results show targeting of signaling components, including integrins, to regions of clustered IgE-FcεRI in processes that depend on stimulated actin polymerization and outward trafficking of REs.

We characterized phenotypes in RBL-2H3 mast cells transfected with human alpha synuclein (a-syn) using stimulated exocytosis of recycling endosomes as a proxy for similar activities of synaptic vesicles in neurons. We found that low expression of a-syn inhibits stimulated exocytosis and that higher expression causes slight enhancement. NMR measurements of membrane interactions correlate with these functional effects: they are eliminated differentially by mutants that perturb helical structure in the helix 1 (A30P) or NAC/helix-2 (V70P) regions of membrane-bound a-syn, but not by other PD-associated mutants or C-terminal truncation. We further found that a-syn (but not A30P or V70P mutants) associates weakly with mitochondria, but this association increases markedly under conditions of cellular stress. These results highlight the importance of specific structural features of a-syn in regulating vesicle release, and point to a potential role for a-syn in perturbing mitochondrial function under pathological conditions.

Stimulated exocytic events provide a means for physiological communication and are a hallmark of the mast cell-mediated allergic response. In mast cells these processes are triggered by antigen crosslinking of IgE bound to its high-affinity receptor, FcϵRI, on the cell surface. Here we use the endosomal v-SNARE VAMP8, and the lysosomal hydrolase β-hexosaminidase (β-Hex), each C-terminally fused to super-ecliptic pHluorin, to monitor stimulated exocytosis. Using these pHluorin-tagged constructs, we monitor stimulated exocytosis by fluorimetry and visualize individual exocytic events with total internal reflection (TIRF) microscopy. Similar to constitutive recycling endosome (RE) trafficking, we find that stimulated RE exocytosis, monitored by VAMP8, is attenuated by expression of dominant negative (S25N) Rab11. Stimulated β-Hex exocytosis is also reduced in the presence of S25N Rab11, suggesting that expression of this mutant broadly impacts exocytosis. Interestingly, pretreatment with inhibitors of actin polymerization, cytochalasin D or latrunculin A, substantially restores both RE and lysosome exocytosis in cells expressing S25N Rab11. Conversely, stabilizing F-actin with jasplakinolide inhibits antigen-stimulated exocytosis but is not additive with S25N Rab11-mediated inhibition, suggesting that these reagents inhibit related processes. Together, our results suggest that Rab11 participates in the regulation necessary for depolymerization of the actin cytoskeleton during stimulated exocytosis in mast cells.

Ca2+ mobilization in response to cross-linking of IgE bound to its high affinity receptor, FcεRI, on mast cells is central to immune allergic responses. Stimulated tyrosine phosphorylation caused by this cross-linking activates store-operated Ca2+ entry that results in sustained Ca2+ oscillations dependent on Rho family GTPases and phosphoinositide synthesis. Coupling of the endoplasmic reticulum (ER) Ca2+ sensor, stromal interaction molecule 1 (STIM1), to the Ca2+-selective channel, Orai1, is regulated by these elements and depends on membrane organization, both at the plasma membrane and at the ER. Mitochondria also contribute to the regulation of Ca2+ mobilization, and we describe recent evidence that the ER membrane protein vesicle-associated membrane protein-associated protein (VAP) plays a significant role in the coupling between ER and mitochondria in this process. In addition to granule exocytosis, Ca2+ mobilization in these cells also contributes to stimulated outward trafficking of recycling endosomes and to antigen-stimulated chemotaxis, and it is pathologically regulated by protozoan parasitic invasion.

Oligomerization of the Ca2+ sensor, STIM1, in the endoplasmic reticulum (ER) membrane, caused by depletion of ER Ca2+ stores, results in STIM1 coupling to the plasma membrane Ca2+ channel protein, Orai1, to activate Ca2+ influx in a process known as store-operated Ca2+ entry. We use fluorimetry-based fluorescence resonance energy transfer (FRET) to monitor changes in STIM1 oligomerization in COS7 cells transfected with STIM1 constructs containing selected truncations, deletions, and point mutations, and labeled with donor and acceptor fluorescent proteins at either the luminal (N-terminal) or the cytoplasmic (C-terminal) ends. Our results with sequential truncations of STIM1 from the C-terminus support previous evidence that the CRAC activation domain (CAD/SOAR, human sequence 342–448) is an oligomer-promoting segment of STIM1, and they show that truncation just after CAD/SOAR (1–448) causes significantly elevated basal cytoplasmic Ca2+ and spontaneous STIM1 clustering. We find that a 14 amino acid sequence just C-terminal of CAD/SOAR (449–462) prevents spontaneous clustering and activation of STIM1 in COS7 cells. In response to store depletion, C-terminally labeled STIM1 without CAD/SOAR clusters together with CAD/SOAR-containing STIM1 constructs. However, these donor-acceptor pairs do not undergo a stimulated increase in FRET, exhibiting instead a decrease in FRET consistent with a stimulated conformational extension in full length STIM1. We find that the 14 amino acid sequence plays a regulatory role in this process. Overall, our FRET results provide evidence in live cells that Ca2+ store depletion stimulates a conformational extension in the cytoplasmic segment of STIM1 that accompanies its oligomerization.

This article provides a brief overview of the state of discourse, politics and provision of abortion in the Anglophone West, including developments in the wake of the COVID-19 pandemic. It then surveys three promising directions for feminist abortion scholarship. The first is work inspired by the Reproductive Justice Movement, that points to the intersectional axes of inequality that shape abortion discourse and position us in relation to reproductive choice and access issues. The second is work that examines the particularity of the constitution of the aborting body, reflecting the particularity of the pregnant body. This is a specific body, with a specific history; abortion discourse draws from and makes a significant contribution to the meaning and lived experience of this body. The third area of scholarship we highlight is that which seeks to amplify the meaning of abortion as a social good. Much abortion scholarship is attuned to a critique of negative aspects of abortion-from its representation in popular culture to restrictive law and access issues. This is critical work but/and the performative nature of abortion scholarship, like all discourse, means that it can amplify the association of negativity with abortion. The article concludes by introducing the articles contained in the special section of Women's Studies International Forum, 'Abortion at the edges: Politics, practices, performances'.

Lipid phase heterogeneity in plasma membranes is thought to play a key role in targeting cellular signaling, but efforts to test lipid raft and related hypotheses are limited by the spatially dynamic nature of these phase-based structures in cells and by experimental characterization tools. We suggest that perturbation of plasma membrane structure by lipid derivatives offers a general method for assessing functional roles for ordered lipid regions in membrane and cell biology. We previously reported that short chain ceramides with either C2 or C6 acyl chains inhibit antigen-stimulated Ca2+ mobilization (Gidwani et al., 2003). We now show that these short chain ceramides inhibit liquid order (Lo)-liquid disorder (Ld) phase separation in giant plasma membrane vesicles that normally occurs at low temperatures. Furthermore, they are effective inhibitors of tyrosine phosphorylation stimulated by antigen, as well as store-operated Ca2+ entry. In Jurkat T cells, C6-ceramide is also effective at inhibiting Ca2+ mobilization stimulated by either anti-TCR or thapsigargin, consistent with the view that these short chain ceramides effectively interfere with functional responses that depend on ordered lipid regions in the plasma membrane., Summary: Our manuscript describes how perturbation of plasma membrane structure by short chain ceramides offers a general method for assessing functional roles for ordered lipid regions in membrane and cell biology.

Lipid phase heterogeneity in the plasma membrane is thought to be crucial for many aspects of cell signaling, but the physical basis of participating membrane domains such as "lipid rafts" remains controversial. Here we consider a lattice model yielding a phase diagram that includes several states proposed to be relevant for the cell membrane, including microemulsion - which can be related to membrane curvature - and Ising critical behavior. Using a neural network-based machine learning approach, we compute the full phase diagram of this lattice model. We analyze selected regions of this phase diagram in the context of a signaling initiation event in mast cells: recruitment of the membrane-anchored tyrosine kinase Lyn to a cluster of transmembrane of IgE-Fc{\epsilon}RI receptors. We find that model membrane systems in microemulsion and Ising critical states can mediate roughly equal levels of kinase recruitment (binding energy ~ -0.6 kBT), whereas a membrane near a tricritical point can mediate much stronger kinase recruitment (-1.7 kBT). By comparing several models for lipid heterogeneity within a single theoretical framework, this work points to testable differences between existing models. We also suggest the tricritical point as a new possibility for the basis of membrane domains that facilitate preferential partitioning of signaling components., Comment: 33 pages, 7 figures, 16 supplementary pages, 10 supplementary figures

The high-affinity receptor for IgE expressed on the surface of mast cells and basophils interacts with antigens, via bound IgE antibody, and triggers secretion of inflammatory mediators that contribute to allergic reactions. To understand how past inputs (memory) influence future inflammatory responses in mast cells, a microfluidic device was used to precisely control exposure of cells to alternating stimulatory and non-stimulatory inputs. We determined that the response to subsequent stimulation depends on the interval of signaling quiescence. For shorter intervals of signaling quiescence, the second response is blunted relative to the first response, whereas longer intervals of quiescence induce an enhanced second response. Through an iterative process of computational modeling and experimental tests, we found that these memory-like phenomena arise from a confluence of rapid, short-lived positive signals driven by the protein tyrosine kinase Syk; slow, long-lived negative signals driven by the lipid phosphatase Ship1; and slower degradation of Ship1 co-factors. This work advances our understanding of mast cell signaling and represents a generalizable approach for investigating the dynamics of signaling systems.

This article foregrounds the public voices of children of gay and lesbian parents in Australia. In doing so, it contributes to scholarship about these children but its focus is one that has been only partially explored. It begins by showing how children's welfare continues to be the ground on which both homophobic and pro-gay public discourse stakes territory. The article documents the emergence in Australia, since the early 2000s, of a new self-defined subject position, evident in a range of Internet sources. The article analyzes the ways in which the voices which speak from this position are both constituted by the pro-gay discourse and also speak back to it. Three themes are identified and analyzed: resistance to the binary opposition of the child at risk versus the prospering child, responses to heteronormativity and homophobia, and the “love makes a family” discourse. While the children of gay and lesbian parents are not immune to the public discourses that create them as objects of debate, attention...

Decreased luminal endoplasmic reticulum (ER) Ca2+ concentration triggers oligomerization and clustering of the ER Ca2+ sensor STIM1 to promote its association with plasma membrane Orai1 Ca2+ channels leading to increased Ca2+ influx. A key step in STIM1 activation is the release of its SOAR domain from an intramolecular clamp formed with the STIM1 first coiled-coil (CC1) region. Using a truncated STIM1(1-343) molecule that captures or releases the isolated SOAR domain depending on luminal ER Ca2+ concentrations, we analyzed the early molecular events that control the intramolecular clamp formed between the CC1 and SOAR domains. We found that STIM1 forms constitutive dimers, and its CC1 domain can bind the SOAR domain of another STIM1 molecule in trans. Artificial oligomerization failed to liberate the SOAR domain or activate STIM1 unless the luminal Ca2+-sensing domains were removed. We propose that the release of SOAR from its CC1 interaction is controlled by changes in the orientation of the two CC1 domains in STIM1 dimers. Ca2+ unbinding in the STIM1 luminal domains initiates the conformational change allowing SOAR domain liberation and clustering, leading to Orai1 channel activation.