69 research outputs found
The Role of Protein Kinase CĪ· in T Cell Biology
Protein kinase CĪ· (PKCĪ·) is a member of the novel PKC subfamily, which also includes Ī“, Īµ, and Īø isoforms. Compared to the other novel PKCs, the function of PKCĪ· in the immune system is largely unknown. Several studies have started to reveal the role of PKCĪ·, particularly in T cells. PKCĪ· is highly expressed in T cells, and is upregulated during thymocyte positive selection. Interestingly, like the Īø isoform, PKCĪ· is also recruited to the immunological synapse that is formed between a T cell and an antigen-presenting cell. However, unlike PKCĪø, which becomes concentrated to the central region of the synapse, PKCĪ· remains in a diffuse pattern over the whole area of the synapse, suggesting distinctive roles of these two isoforms in signal transduction. Although PKCĪ· is dispensable for thymocyte development, further analysis of PKCĪ·- or PKCĪø-deficient and double-knockout mice revealed the redundancy of these two isoforms in thymocyte development. In contrast, PKCĪ· rather than PKCĪø, plays an important role for T cell homeostatic proliferation, which requires recognition of self-antigen. Another piece of evidence demonstrating that PKCĪ· and PKCĪø have isoform-specific as well as redundant roles come from the analysis of CD4 to CD8 T cell ratios in the periphery of these knockout mice. Deficiency in PKCĪ· or PKCĪø had opposing effects as PKCĪ· knockout mice had a higher ratio of CD4 to CD8 T cells compared to that of wild-type mice, whereas PKCĪø-deficient mice had a lower ratio. Biochemical studies showed that calcium flux and NFĪŗB translocation is impaired in PKCĪ·-deficient T cells upon TCR crosslinking stimulation, a character shared with PKCĪø-deficient T cells. However, unlike the case with PKCĪø, the mechanistic study of PKCĪ· is at early stage and the signaling pathways involving PKCĪ·, at least in T cells, are essentially unknown. In this review, we will cover the topics mentioned above as well as provide some perspectives for further investigations regarding PKCĪ·
Initiation of TCR Phosphorylation and Signal Transduction
Recent data with CD8+ T cells show that the initial phase of T cell receptor (TCR) binding to MHCāpeptide (MHCp) is quickly followed by a second, stronger, binding phase representing the binding of CD8 to the MHCp. This second phase requires signaling by a Src-family kinase such as Lck. These data point out two aspects of the initial stage of TCR signaling that have not yet been clearly resolved. Firstly, how and by which Src-family kinase, is the initial phosphorylation of CD3Ī¶ accomplished, given that the Lck associated with the co-receptors (CD4 or CD8) is not yet available. Secondly, what is the mechanism by which the co-receptor is brought close to the bound TCR before the co-receptor binds to MHCp
CD8Ī±Ī± and āĪ±Ī² isotypes are equally recruited to the immunological synapse through their ability to bind to MHC class I
In silico Modeling of Itk Activation Kinetics in Thymocytes Suggests Competing Positive and Negative IP4 Mediated Feedbacks Increase Robustness
The inositol-phosphate messenger inositol(1,3,4,5)tetrakisphosphate (IP4) is
essential for thymocyte positive selection by regulating plasma-membrane
association of the protein tyrosine kinase Itk downstream of the T cell
receptor (TCR). IP4 can act as a soluble analog of the phosphoinositide
3-kinase (PI3K) membrane lipid product phosphatidylinositol(3,4,5)trisphosphate
(PIP3). PIP3 recruits signaling proteins such as Itk to cellular membranes by
binding to PH and other domains. In thymocytes, low-dose IP4 binding to the Itk
PH domain surprisingly promoted and high-dose IP4 inhibited PIP3 binding of Itk
PH domains. However, the mechanisms that underlie the regulation of membrane
recruitment of Itk by IP4 and PIP3 remain unclear. The distinct Itk PH domain
ability to oligomerize is consistent with a cooperative-allosteric mode of IP4
action. However, other possibilities cannot be ruled out due to difficulties in
quantitatively measuring the interactions between Itk, IP4 and PIP3, and in
generating non-oligomerizing Itk PH domain mutants. This has hindered a full
mechanistic understanding of how IP4 controls Itk function. By combining
experimentally measured kinetics of PLC{\gamma}1 phosphorylation by Itk with in
silico modeling of multiple Itk signaling circuits and a maximum entropy
(MaxEnt) based computational approach, we show that those in silico models
which are most robust against variations of protein and lipid expression levels
and kinetic rates at the single cell level share a cooperative-allosteric mode
of Itk regulation by IP4 involving oligomeric Itk PH domains at the plasma
membrane. This identifies MaxEnt as an excellent tool for quantifying
robustness for complex TCR signaling circuits and provides testable predictions
to further elucidate a controversial mechanism of PIP3 signaling
Ligand-engaged TCR is triggered by Lck not associated with CD8 coreceptor
ProducciĆ³n CientĆficaThe earliest molecular events in T-cell recognition have not yet been fully described, and the initial T-cell receptor (TCR)-triggering mechanism remains a subject of controversy. Here, using total internal reflection/Forster resonance energy transfer microscopy, we observe a two-stage interaction between TCR, CD8 and major histocompatibility complex (MHC)-peptide. There is an early (within seconds) interaction between CD3Ī¶ and the coreceptor CD8 that is independent of the binding of CD8 to MHC, but that requires CD8 association with Lck. Later (several minutes) CD3Ī¶āCD8 interactions require CD8āMHC binding. Lck can be found free or bound to the coreceptor. This work indicates that the initial TCR-triggering event is induced by free Lck. The early signalling events that trigger initial T-cell receptor signalling are not clearly defined. Here the authors show that this occurs in two stages, the first between the CD8 coreceptor and CD3 requiring Lck association to CD8, while the second interaction requires binding of major histocompatibility molecules
Coreceptor affinity for MHC defines peptide specificity requirements for TCR interaction with coagonist peptide-MHC
Recent work has demonstrated that nonstimulatory endogenous peptides can enhance T cell recognition of antigen, but MHCI- and MHCII-restricted systems have generated very different results. MHCII-restricted TCRs need to interact with the nonstimulatory peptideāMHC (pMHC), showing peptide specificity for activation enhancers or coagonists. In contrast, the MHCI-restricted cells studied to date show no such peptide specificity for coagonists, suggesting that CD8 binding to noncognate MHCI is more important. Here we show how this dichotomy can be resolved by varying CD8 and TCR binding to agonist and coagonists coupled with computer simulations, and we identify two distinct mechanisms by which CD8 influences the peptide specificity of coagonism. Mechanism 1 identifies the requirement of CD8 binding to noncognate ligand and suggests a direct relationship between the magnitude of coagonism and CD8 affinity for coagonist pMHCI. Mechanism 2 describes how the affinity of CD8 for agonist pMHCI changes the requirement for specific coagonist peptides. MHCs that bind CD8 strongly were tolerant of all or most peptides as coagonists, but weaker CD8-binding MHCs required stronger TCR binding to coagonist, limiting the potential coagonist peptides. These findings in MHCI systems also explain peptide-specific coagonism in MHCII-restricted cells, as CD4āMHCII interaction is generally weaker than CD8āMHCI.National Institutes of Health (U.S.). Pioneer Awar
Nonstimulatory peptide?MHC enhances human T-cell antigen-specific responses by amplifying proximal TCR signaling
Self-love and sociability: the ārudiments of commerceā in the state of nature
Istvan Hontās classic work on the theoretical links between the seventeenth-century natural jurists Hugo Grotius and Samuel Pufendorf and the eighteenth-century Scottish political economists remains a popular trope among intellectual and economic historians of various stamps. Despite this, a common criticism levelled at Hont remains his relative lack of engagement with the relationship between religion and economics in the early modern period. This paper challenges this aspect of Hontās narrative by drawing attention to an alternative, albeit complementary, assessment of the natural jurisprudential heritage of eighteenth-century British political economy. Specifically, the article attempts to map on to Hontās thesis the Christian Stoic interpretation of Grotius and Pufendorf which has gained greater currency in recent years. In doing so, the paper argues that Grotius and Pufendorfās contributions to the āunsocial sociabilityā debate do not necessarily lead directly to the Scottish school of political economists, as is commonly assumed. Instead, it contends that a reconsideration of Grotius and Pufendorf as neo-Stoic theorists, particularly via scrutiny of their respective adaptations of the traditional Stoic theory of oikeiosis, steers us towards the heart of the early English āclericalā Enlightenment
A novel canine kidney cell line model for the evaluation of neoplastic development: karyotype evolution associated with spontaneous immortalization and tumorigenicity
The molecular mechanisms underlying spontaneous neoplastic transformation in cultured mammalian cells remain poorly understood, confounding recognition of parallels with the biology of naturally occurring cancer. The broad use of tumorigenic canine cell lines as research tools, coupled with the accumulation of cytogenomic data from naturally occurring canine cancers, makes the domestic dog an ideal system in which to investigate these relationships. We developed a canine kidney cell line, CKB1-3T7, which allows prospective examination of the onset of spontaneous immortalization and tumorigenicity. We documented the accumulation of cytogenomic aberrations in CKB1-3T7 over 24Ā months in continuous culture. The majority of aberrations emerged in parallel with key phenotypic changes in cell morphology, growth kinetics, and tumor incidence and latency. Focal deletion of CDKN2A/B emerged first, preceding the onset and progression of tumorigenic potential, and progressed to a homozygous deletion across the cell population during extended culture. Interestingly, CKB1-3T7 demonstrated a tumorigenic phenotype in vivo prior to exhibiting loss of contact inhibition in vitro. We also performed the first genome-wide characterization of the canine tumorigenic cell line MDCK, which also exhibited CDKN2A/B deletion. MDCK and CKB1-3T7 cells shared several additional aberrations that we have reported previously as being highly recurrent in spontaneous canine cancers, many of which, as with CDKN2A/B deletion, are evolutionarily conserved in their human counterparts. The conservation of these molecular events across multiple species, in vitro and in vivo, despite their contrasting karyotypic architecture, is a powerful indicator of a common mechanism underlying emerging neoplastic activity. Through integrated cytogenomic and phenotypic characterization of serial passages of CKB1-3T7 from initiation to development of a tumorigenic phenotype, we present a robust and readily accessible model (to be made available through the American Type Culture Collection) of spontaneous neoplastic transformation that overcomes many of the limitations of earlier studies. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s10577-015-9474-8) contains supplementary material, which is available to authorized users
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