Cortical sinus probing, S1P1-dependent entry and flow-based capture of egressing T cells.

The cellular dynamics of the egress of lymphocytes from lymph nodes are poorly defined. Here we visualized the branched organization of lymph node cortical sinuses and found that after entry, some T cells were retained, whereas others returned to the parenchyma. T cells deficient in sphingosine 1-phosphate receptor type 1 probed the sinus surface but failed to enter the sinuses. In some sinuses, T cells became rounded and moved unidirectionally. T cells traveled from cortical sinuses into macrophage-rich sinus areas. Many T cells flowed from medullary sinuses into the subcapsular space. We propose a multistep model of lymph node egress in which cortical sinus probing is followed by entry dependent on sphingosine 1-phosphate receptor type 1, capture of cells in a sinus region with flow, and transport to medullary sinuses and the efferent lymph

CCL3 Promotes Germinal Center B Cells Sampling by Follicular Regulatory T Cells in Murine Lymph Nodes

Previous studies and our findings suggest upregulated expression of proinflammatory chemokines CCL3/4 in germinal center (GC) centrocytes. However, the role of CCL3/4 for centrocyte interactions with follicular T cells and regulation of humoral immunity is poorly understood. We found that CCL3 promotes chemotaxis of Tfr cells ex vivo. Two-photon imaging revealed that B cells-intrinsic production of CCL3 promotes their probing by follicular regulatory T cells (Tfr) within GCs of murine lymph nodes. Overall this study suggests that CCL3 facilitates direct interactions of foreign antigen-specific GC B cells and their negative regulation with Tfr cells in vivo

Regulation of the Alternative Sigma Factor σ(E) during Initiation, Adaptation, and Shutoff of the Extracytoplasmic Heat Shock Response in Escherichia coli

The alternative sigma factor σ(E) is activated in response to stress in the extracytoplasmic compartment of Escherichia coli. Here we show that σ(E) activity increases upon initiation of the stress response by a shift to an elevated temperature (43°C) and remains at that level for the duration of the stress. When the stress is removed by a temperature downshift, σ(E) activity is strongly repressed and then slowly returns to levels seen in unstressed cells. We provide evidence that information about the state of the cell envelope is communicated to σ(E) primarily through the regulated proteolysis of the inner membrane anti-sigma factor RseA, as the degradation rate of RseA is correlated with the changes in σ(E) activity throughout the stress response. However, the relationship between σ(E) activity and the rate of degradation of RseA is complex, indicating that other factors may cooperate with RseA and serve to fine-tune the response

Transiently antigen primed B cells can generate multiple subsets of memory cells

<div><p>Memory B cells are long-lived cells that generate a more vigorous response upon recognition of antigen (Ag) and T cell help than naïve B cells and ensure maintenance of durable humoral immunity. Functionally distinct subsets of murine memory B cells have been identified based on isotype switching of BCRs and surface expression of the co-stimulatory molecule CD80 and co-inhibitory molecule PD-L2. Memory B cells in a subpopulation with low surface expression of CD80 and PD-L2 are predominantly non-isotype switched and can be efficiently recruited into germinal centers (GCs) in secondary responses. In contrast, a CD80 and PD-L2 positive subset arises predominantly from GCs and can quickly differentiate into antibody-secreting plasma cells (PCs). Here we demonstrate that single transient acquisition of Ag by B cells may be sufficient for their long-term participation in GC responses and for development of various memory B cell subsets including CD80 and PD-L2 positive effector-like memory cells that rapidly differentiate into class-switched PCs during recall responses.</p></div

Design principles of the proteolytic cascade governing the σ(E)-mediated envelope stress response in Escherichia coli: keys to graded, buffered, and rapid signal transduction

Proteolytic cascades often transduce signals between cellular compartments, but the features of these cascades that permit efficient conversion of a biological signal into a transcriptional output are not well elucidated. σ(E) mediates an envelope stress response in Escherichia coli, and its activity is controlled by regulated degradation of RseA, a membrane-spanning anti-σ factor. Examination of the individual steps in this protease cascade reveals that the initial, signal-sensing cleavage step is rate-limiting; that multiple ATP-dependent proteases degrade the cytoplasmic fragment of RseA and that dissociation of σ(E) from RseA is so slow that most free σ(E) must be generated by the active degradation of RseA. As a consequence, the degradation rate of RseA is set by the amount of inducing signal, and insulated from the “load” on and activity of the cytoplasmic proteases. Additionally, changes in RseA degradation rate are rapidly reflected in altered σ(E) activity. These design features are attractive as general components of signal transduction pathways governed by unstable negative regulators

Fine-tuning of the Escherichia coli σ(E) envelope stress response relies on multiple mechanisms to inhibit signal-independent proteolysis of the transmembrane anti-sigma factor, RseA

Proteolytic cascades are widely implicated in signaling between cellular compartments. In Escherichia coli, accumulation of unassembled outer membrane porins (OMPs) in the envelope leads to expression of σ(E)-dependent genes in the cytoplasmic cellular compartment. A proteolytic cascade conveys the OMP signal by regulated proteolysis of RseA, a membrane-spanning anti-sigma factor whose cytoplasmic domain inhibits σ(E)-dependent transcription. Upon activation by OMP C termini, the membrane localized DegS protease cleaves RseA in its periplasmic domain, the membrane-embedded protease RseP (YaeL) cleaves RseA near the inner membrane, and the released cytoplasmic RseA fragment is further degraded. Initiation of RseA degradation by activated DegS makes the system sensitive to a wide range of OMP concentrations and unresponsive to variations in the levels of DegS and RseP proteases. These features rely on the inability of RseP to cleave intact RseA. In the present report, we demonstrate that RseB, which binds to the periplasmic face of RseA, and DegS each independently inhibits RseP cleavage of intact RseA. Thus, the function of RseB, widely conserved among bacteria using the σ(E) pathway, and the second role of DegS (in addition to RseA proteolysis initiation) is to improve the performance characteristics of this signal transduction system

The actin regulator coronin 1A is mutant in a thymic egress-deficient mouse strain and in a patient with severe combined immunodeficiency

Mice carrying the recessive locus for peripheral T cell deficiency (Ptcd) have a block in thymic egress, but the mechanism responsible is undefined. Here we found that Ptcd T cells had an intrinsic migration defect, impaired lymphoid tissue trafficking and irregularly shaped protrusions. Characterization of the Ptcd locus showed a point substitution of lysine for glutamic acid at position 26 in the actin regulator coronin 1A that enhanced its inhibition of the actin regulator Arp2/3 and resulted in its mislocalization from the leading edge of migrating T cells. The discovery of another coronin 1A mutant during an N-ethyl-N-nitrosourea-mutagenesis screen for T cell-lymphopenic mice prompted us to evaluate a T cell-deficient, B cell-sufficient and natural killer cell-sufficient patient with severe combined immunodeficiency, whom we found had mutations in both CORO1A alleles. Our findings establish a function for coronin 1A in T cell egress, identify a surface of coronin involved in Arp2/3 regulation and demonstrate that actin regulation is a biological process defective in human and mouse severe combined immunodeficiency