Abstract only
Naïve T cells activated through acute antigen stimulation expand and contract independently of the presence of the death receptor Fas. However, Fas‐FasL interactions play a critical ...role in TCR‐induced apoptosis of activated CD4
+
T cells. Fas deficiency prompts the accumulation of CD44
+
memory phenotype T cells in mice, suggesting Fas may play a more prominent role in apoptosis of memory T cells. The sensitivity of cells derived from naïve versus memory CD4
+
T cells to Fas‐induced apoptosis has not been addressed. We compared the susceptibility of purified naïve versus memory human CD4
+
T cells, both directly ex vivo and after in vitro activation and expansion, to undergo Fas‐induced apoptosis.
Remarkably, effector memory phenotype (CD27
−
CCR7
−
) cells were the most sensitive to non‐crosslinked anti‐Fas induced cell death. We have previously shown a correlation between lipid raft‐associated Fas and susceptibility to undergo apoptosis. Memory cells have increased lipid raft‐associated Fas surface protein versus naïve cells, which could account for increased sensitivity to Fas. Memory cells have more efficient assembly and activation of proximal Fas signaling components, including FADD and Caspase 8. These results suggest that effector memory cells have more efficient proximal Fas signaling and are intrinsically more sensitive to Fas‐induced apoptosis, possibly due to lipid raft localiztion of Fas.
The death-inducing signaling complex (DISC) formed by the death receptor Fas, the adaptor protein FADD and caspase-8 mediates the extrinsic apoptotic program. Mutations in Fas that disrupt the DISC ...cause autoimmune lymphoproliferative syndrome (ALPS). Here we show that the Fas-FADD death domain (DD) complex forms an asymmetric oligomeric structure composed of 5-7 Fas DD and 5 FADD DD, whose interfaces harbor ALPS-associated mutations. Structure-based mutations disrupt the Fas-FADD interaction in vitro and in living cells; the severity of a mutation correlates with the number of occurrences of a particular interaction in the structure. The highly oligomeric structure explains the requirement for hexameric or membrane-bound FasL in Fas signaling. It also predicts strong dominant negative effects from Fas mutations, which are confirmed by signaling assays. The structure optimally positions the FADD death effector domain (DED) to interact with the caspase-8 DED for caspase recruitment and higher-order aggregation.
Transmembrane metalloproteinases of the disintegrin and metalloproteinase (ADAM) family control cell signaling interactions
via hydrolysis of protein extracellular domains. Prior work has shown that ...the receptor tyrosine kinase, c-Kit (CD117), is
essential for mast cell survival and that serum levels of c-Kit increase in proliferative mast cell disorders, suggesting
the existence of c-Kit shedding pathways in mast cells. In the present work, we report that tumor necrosis factor α-converting
enzyme (TACE; ADAM-17) mediates shedding of c-Kit. Stimulation of transfected cells with phorbol 12-myristate 13-acetate (PMA)
induced metalloproteinase-mediated release of c-Kit ectodomain, which increased further upon TACE overexpression. By contrast,
TACE-deficient fibroblasts did not demonstrate inducible release, thus identifying TACE as the metalloproteinase primarily
responsible for PMA-induced c-Kit shedding. Surface expression of c-Kit by the human mast cell-1 line decreased upon phorbol-induced
shedding, which involved metalloproteinase activity susceptible to inhibition by tissue inhibitor of metalloproteinase (TIMP)-3.
To further explore the role of TACE in shedding of c-Kit from mast cells, we compared the behavior of mast cells derived from
murine embryonic stem cells. In these studies, PMA decreased surface c-Kit levels on mast cells expressing wild-type (+/+)
TACE but not on those expressing an inactive mutant (ÎZn/ÎZn), confirming the role of TACE in PMA-induced c-Kit shedding.
Compared with TACE +/+ cells, TACE ÎZn/ÎZn mast cells also demonstrated decreased constitutive shedding and increased basal surface expression of c-Kit, with diminished
apoptosis in response to c-Kit ligand deprivation. These data suggest that TACE controls mast cell survival by regulating
shedding and surface expression of c-Kit.
Members of the cofilin/ADF family of proteins sever actin filaments, increasing the number of filament ends available for polymerization or depolymerization. Cofilin binds actin filaments with ...positive cooperativity, forming clusters of contiguously bound cofilin along the filament lattice. Filament severing occurs preferentially at boundaries between bare and cofilin-decorated (cofilactin) segments and is biased at 1 side of a cluster. A molecular understanding of cooperative binding and filament severing has been impeded by a lack of structural data describing boundaries. Here, we apply methods for analyzing filament cryo-electron microscopy (cryo-EM) data at the single subunit level to directly investigate the structure of boundaries within partially decorated cofilactin filaments. Subnanometer resolution maps of isolated, bound cofilin molecules and an actin-cofilactin boundary indicate that cofilin-induced actin conformational changes are local and limited to subunits directly contacting bound cofilin. An isolated, bound cofilin compromises longitudinal filament contacts of 1 protofilament, consistent with a single cofilin having filament-severing activity. An individual, bound phosphomimetic (S3D) cofilin with weak severing activity adopts a unique binding mode that does not perturb actin structure. Cofilin clusters disrupt both protofilaments, consistent with a higher severing activity at boundaries compared to single cofilin. Comparison of these structures indicates that this disruption is substantially greater at pointed end sides of cofilactin clusters than at the barbed end. These structures, with the distribution of bound cofilin clusters, suggest that maximum binding cooperativity is achieved when 2 cofilins occupy adjacent sites. These results reveal the structural origins of cooperative cofilin binding and actin filament severing.
Daratumumab, a monoclonal CD38 antibody, is approved in the treatment of myeloma, but its efficacy and safety in light-chain (AL) amyloidosis has not been formally studied. This prospective phase 2 ...trial of daratumumab monotherapy for the treatment of AL amyloidosis was designed to determine the safety, tolerability, and hematologic and clinical response. Daratumumab 16 mg/kg was administered by IV infusion once weekly for weeks 1 to 8, every 2 weeks for weeks 9 to 24, and every 4 weeks thereafter until progression or unacceptable toxicity, for up to 24 months. Twenty-two patients with previously treated AL amyloidosis were enrolled. The majority of the patients had received high-dose melphalan and stem cell transplantation and/or treatment with a proteasome inhibitor. The median time between prior therapy and trial enrollment was 9 months (range, 1-180 months). No grade 3-4 infusion-related reactions occurred. The most common grade ≥3 adverse events included respiratory infections (n = 4; 18%) and atrial fibrillation (n = 4, 18%). Hematologic complete and very-good-partial response occurred in 86% of patients. The median time to first and best hematologic response was 4 weeks and 3 months, respectively. Renal response occurred in 10 of 15 patients (67%) with renal involvement and cardiac response occurred in 7 of 14 patients (50%) with cardiac involvement. In summary, daratumumab is well tolerated in patients with relapsed AL amyloidosis and leads to rapid and deep hematologic responses and organ responses. This trial was registered at www.clinicaltrials.gov as #NCT02841033.
•Daratumumab is well tolerated in patients with AL amyloidosis when used with appropriate preinfusion and postinfusion medications.•Daratumumab leads to rapid and deep hematologic responses in previously treated patients with AL amyloidosis.
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Computational and structural studies have been indispensable in investigating the molecular origins of actin filament mechanical properties and modulation by the regulatory severing protein cofilin. ...All-atom molecular dynamics simulations of cofilactin filament structures determined by electron cryomicroscopy reveal how cofilin enhances the bending and twisting compliance of actin filaments. Continuum mechanics models suggest that buckled cofilactin filaments localize elastic energy at boundaries between bare and cofilin-decorated segments because of their nonuniform elasticity, thereby accelerating filament severing. Here, we develop mesoscopic length-scale (cofil)actin filament models and evaluate the effects of compressive and twisting loads on strain energy distribution at specific interprotein interfaces. The models reliably capture the filament bending and torsional rigidities and intersubunit torsional flexibility measured experimentally with purified protein components. Buckling is predicted to enhance cofilactin filament severing with minimal effects on cofilin occupancy, whereas filament twisting enhances cofilin dissociation without compromising filament integrity. Preferential severing at actin-cofilactin boundaries of buckled filaments is more prominent than predicted by continuum models because of the enhanced spatial resolution. The models developed here will be valuable for evaluating the effects of filament shape deformations on filament stability and interactions with regulatory proteins, and analysis of single filament manipulation assays.
Polymorphous low-grade adenocarcinoma (PLGA) is the second most frequent type of malignant tumor of the minor salivary glands. We identified PRKD1 hotspot mutations encoding p.Glu710Asp in 72.9% of ...PLGAs but not in other salivary gland tumors. Functional studies demonstrated that this kinase-activating alteration likely constitutes a driver of PLGA.
Conflicts with Friends Hood, Anthony C.; Cruz, Kevin S.; Bachrach, Daniel G.
Journal of business and psychology,
02/2017, Volume:
32, Issue:
1
Journal Article
Peer reviewed
Drawing on conservation of resources theory, multiplex social networks research, and the emerging conflict involvement perspective, the purpose of this study is to develop and test a multiplex view ...of conflict that explicitly accounts for the nature of the social relationships between those involved in intrateam conflict and how these multiplex relationships differentially impact team performance. Data were collected from 120 teams engaged in a 4-month business simulation. Relationship conflicts occurring among team members who are friends have a negative impact on team performance, whereas those occurring between non-friends have a positive impact on team performance. Although we also find non-friend task conflicts to be beneficial for team performance, friend task conflicts have no impact on team performance. This study highlights the dark side of workplace friendships and admonishes managers to pay close attention not only to conflicts among employees, but also to the relational closeness of those involved in conflict. The current study provides empirical support for the emerging conflict involvement perspective by explicitly assessing the number of individuals involved in conflict as well as the type of relationships between them. We also extend research on multiplex relationships from the individual to the team level of analysis. Finally, we respond to calls for studies of multiplexity that include both positive and negative relationships.
The assembly of actin filaments and filament networks generate forces that drive cell and vesicle movement. These structures and the comprising actin filaments must be mechanically stable to sustain ...these forces and maintain their structural integrity. Filaments in these dynamic structures must also be disassembled to recycle and replenish the pool of actin monomers available for polymerization. Actin-severing proteins such as cofilin and contractile myosin motor proteins fragment these nominally stable structures. We developed a mesoscopic-length-scale actin filament model to investigate force-induced filament fragmentation. We show that fragmentation in our model occurs at curvatures similar to previous measurements of fragmentation within (cofil)actin and actin-cofilactin boundaries. Boundaries between bare and cofilin-decorated segments are brittle and fragment at small bending and twisting deformations. Extending filaments disperses strain uniformly over subunit interfaces, and filaments fragment with no detectable partial rupture or plastic deformation. In contrast, bending or twisting filaments imposes nonuniform interface strain and leads to partial interface rupture, accelerating filament fragmentation. As a result, the rupture force under compressive loads is an order of magnitude lower than under tensile loads. Partial interface rupture may be a primary mechanism of accelerating actin filament fragmentation by other actin-destabilizing proteins.
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