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The identification of specific genetic loci that contribute to inflammatory and autoimmune diseases has proved difficult due to the contribution of multiple interacting genes, the inherent genetic heterogeneity present in human populations, and a lack of new mouse mutants. By using N-ethyl-N-nitrosourea (ENU) mutagenesis to discover new immune regulators, we identified a point mutation in the murine phospholipase Cg2 (Plcg2) gene that leads to severe spontaneous inflammation and autoimmunity. The disease is composed of an autoimmune component mediated by autoantibody immune complexes and B and T cell independent inflammation. The underlying mechanism is a gain-of-function mutation in Plcg2, which leads to hyperreactive external calcium entry in B cells and expansion of innate inflammatory cells. This mutant identifies Plcg2 as a key regulator in an autoimmune and inflammatory disease mediated by B cells and non-B, non-T haematopoietic cells and emphasizes that by distinct genetic modulation, a single point mutation can lead to a complex immunological phenotype.

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Cell motility is a critical event in many processes and is underlined by complex signalling interactions. Although many components have been implicated in different forms of cell migration, identification of early key mediators of these events has proved difficult. One potential signalling intermediate, PLCγ1, has previously been implicated in growth-factor-mediated chemotaxis but its position and roles in more-complex motility events remain poorly understood. This study links PLCγ1 to early, integrin-regulated changes leading to cell motility. The key role of PLCγ1 was supported by findings that specific depletion of PLCγ1 by small interfering (si)RNA, or by pharmacological inhibition, or the absence of this isoform in PLCγ1–/– cells resulted in the failure to form cell protrusions and undergo cell spreading and elongation in response to integrin engagement. This integrin-PLCγ1 pathway was shown to underlie motility processes involved in morphogenesis of endothelial cells on basement membranes and invasion of cancer cells into such three-dimensional matrices. By combining cellular and biochemical approaches, we have further characterized this signalling pathway. Upstream of PLCγ1 activity, β1 integrin and Src kinase are demonstrated to be essential for phosphorylation of PLCγ1, formation of protein complexes and accumulation of intracellular calcium. Cancer cell invasion and the early morphological changes associated with cell motility were abolished by inhibition of β1 integrin or Src. Our findings establish PLCγ1 as a key player in integrin-mediated cell motility processes and identify other critical components of the signalling pathway involved in establishing a motile phenotype. This suggests a more general role for PLCγ1 in cell motility, functioning as a mediator of both growth factor and integrin-initiated signals.

Phospholipase C␥ (PLC␥) isoforms are regulated through activation of tyrosine kinase-linked receptors. The importance of growth factor-stimulated phosphorylation of specific tyrosine residues has been documented for PLC␥1; however, despite the critical importance of PLC␥2 in B-cell signal transduction, neither the tyrosine kinase(s) that directly phosphorylate PLC␥2 nor the sites in PLC␥2 that become phosphorylated after stimulation are known. By measuring the ability of human PLC␥2 to restore calcium responses to the B-cell receptor stimulation or oxidative stress in a B-cell line (DT40) deficient in PLC␥2, we have demonstrated that two tyrosine residues, Tyr 753 and Tyr 759 , were important for the PLC␥2 signaling function. Furthermore, the double mutation Y753F/Y759F in PLC␥2 resulted in a loss of tyrosine phosphorylation in stimulated DT40 cells. Of the two kinases that previously have been proposed to phosphorylate PLC␥2, Btk, and Syk, purified Btk had much greater ability to phosphorylate recombinant PLC␥2 in vitro, whereas Syk efficiently phosphorylated adapter protein BLNK. Using purified proteins to analyze the formation of complexes, we suggest that function of Syk is to phosphorylate BLNK, providing binding sites for PLC␥2. Further analysis of PLC␥2 tyrosine residues phosphorylated by Btk and several kinases from the Src family has suggested multiple sites of phosphorylation and, in the context of a peptide incorporating residues Tyr 753 and Tyr 759 , shown preferential phosphorylation of Tyr 753 .

Plasticity of cancer metabolism can be a major obstacle to efficient targeting of tumour-specific metabolic vulnerabilities. Here, we identify the compensatory mechanisms following the inhibition of major pathways of central carbon metabolism in cMYC-induced liver tumours. We find that, while inhibition of both glutaminase isoforms (Gls1 and Gls2) in tumours significantly delays tumourigenesis, glutamine catabolism continues due to the action of amidotransferases. Synergistic inhibition of both glutaminases and compensatory amidotransferases is required to block glutamine catabolism and proliferation of mouse and human tumour cells in vitro and in vivo. Gls1 deletion is also compensated by glycolysis. Thus, co-inhibition of Gls1 and hexokinase *

Treatments for severe depression have moderate success rates, often take many weeks to yield responses, and are often followed by relapse or recurrence. Neurobehavioral interventions address these limitations by targeting mechanisms of cognitive and emotional dysregulation directly. This study extends data and observations from a pilot study examining effects of 2 weeks (6 sessions) of adjunctive cognitive control training exercises added to medication and psychotherapy in severely depressed patients. We examined acute effects and predictors of change in rumination, and long-term effects on service utilization. Compared with treatment as usual, exercises were associated with decreases in rumination and decreased use of intensive outpatient services in the following year. Responses were strongest among patients who displayed physiological indicators (pupillary oscillations at the task frequency) of task engagement before the intervention. These indices changed following intervention, suggesting that the intervention required capitalization on relevant attentional mechanisms and addressed fundamental emotional processes through their cognitive substrates.