Bias in the αβ T-cell repertoire: implications for disease pathogenesis and vaccination

The na?ve T-cell repertoire is vast, containing millions of unique T-cell receptor (TCR) structures. Faced with such diversity, the mobilization of TCR structures from this enormous pool was once thought to be a stochastic, even chaotic, process. However, steady and systematic dissection over the last 20 years has revealed that this is not the case. Instead, the TCR repertoire deployed against individual antigens is routinely ordered and biased. Often, identical and near-identical TCR repertoires can be observed across different individuals, suggesting that the system encompasses an element of predictability. This review provides a catalog of αβ TCR bias by disease and by species, and discusses the mechanisms that govern this inherent and widespread phenomenon.     

T-cell development: What does Notch do for T cells?

During their development, T cells are rescued from apoptotic cell death to follow distinct lineage fates. Recent data concerned with the role of the Notch transmembrane receptor in these events are interpreted to show that Notch promotes survival, but contrary to earlier reports has no function in lineage commitment.     

What is the role of cell division in leaf development?

An idea underlying a great deal of research and discussion in plant cell and developmental biology is that the spatial regulation of cell division plays a key role in plant development. In this article, the role of cell division in two aspects of leaf development is analysed: morphogenesis (leaf initiation, growth, and the generation of leaf shape) and histogenesis (the differentiation of leaf cells to form the various cell types that make up a functional leaf). The point of view that emerges from this analysis is that the rate and pattern of cell division is important for leaf development, but does not dictate leaf size, shape, or cell fate.     

A nonredundant role for canonical NF-κB in human myeloid dendritic cell development and function

The plastic role of dendritic cells (DCs) in the regulation of immune responses has made them interesting targets for immunotherapy, but also for pathogens or tumors to evade immunity. Functional alterations of DCs are often ascribed to manipulation of canonical NF-κB activity. However, though this pathway has been linked to murine myeloid DC biology, a detailed analysis of its importance in human myeloid DC differentiation, survival, maturation, and function is lacking. The myeloid DC subsets include interstitial DCs and Langerhans cells. In this study, we investigated the role of canonical NF-κB in human myeloid DCs generated from monocytes (monocyte-derived DCs [mo-DCs]) or CD34(+) progenitors (CD34-derived myeloid DCs [CD34-mDCs]). Inhibition of NF-κB activation during and after mo-DC, CD34-interstitial DC, or CD34-Langerhans cell differentiation resulted in apoptosis induction associated with caspase 3 activation and loss of mitochondrial transmembrane potential. Besides regulating survival, canonical NF-κB activity was required for the acquisition of a DC phenotype. Despite phenotypic differences, however, Ag uptake, costimulatory molecule and CCR7 expression, as well as T cell stimulatory capacity of cells generated under NF-κB inhibition were comparable to control DCs, indicating that canonical NF-κB activity during differentiation is redundant for the development of functional APCs. However, both mo-DC and CD34-mDC functionality were reduced by NF-κB inhibition during activation. In conclusion, canonical NF-κB activity is essential for the development and function of mo-DCs as well as CD34-mDCs. Insight into the role of this pathway may help in understanding how pathogens and tumors escape immunity and aid in developing novel treatment strategies aiming to interfere with human immune responses.     

Suppression of insulin-like3 receptor reveals the role of β-catenin and Notch signaling in gubernaculum development

During male development, the testes move from a high intraabdominal position and descend into the scrotum. The gubernaculum, an inguinoscrotal ligament connecting the testis to the lower abdomen, is believed to play a critical role in this process. The first stage of testicular descent is controlled by insulin like3 hormone (INSL3), produced in testicular Leydig cells. Deletion of Insl3 or its receptor, Rxfp2, in mice causes cryptorchidism. We produced Cre/loxP regulated shRNA transgenic mice targeting RXFP2 expression. We have shown that the transgene was able to reduce Rxfp2 gene expression and thus behaved as a hypomorphic allele of Rxfp2. Variable degrees of uni- and bilateral cryptorchidism was detected in males with the activated shRNA transgene on an Rxfp2+/- background. Conditional suppression of Rxfp2 in the gubernaculum led to cryptorchidism. Gene expression analysis of a mutant cremasteric sac using Illumina microarrays indicated abnormal expression of a significant number of genes in Wnt/β-catenin and Notch pathways. We have demonstrated profound changes in the expression pattern of β-catenin, Notch1, desmin, and androgen receptor (AR), in Rxfp2-/- male embryos, indicating the role of INSL3 in proliferation, differentiation, and survival of specific cellular components of the gubernaculum. We have shown that INSL3/RXFP2 signaling is essential for myogenic differentiation and maintenance of AR-positive cells in the gubernaculum. Males with the deletion of β-catenin or Notch1 in the gubernacular ligament demonstrated abnormal development. Our data indicates that β-catenin and Notch pathways are potential targets of INSL3 signaling during gubernacular development.     

A role for the NF-κB pathway in cell protection from complement-dependent cytotoxicity

Nucleated cells are equipped with several mechanisms that support their resistance to complement-dependent cytotoxicity (CDC). The role of the NF-κB pathway in cell protection from CDC was examined. Elevated sensitivity to CDC was demonstrated in cells lacking the p65 subunit of NF-κB or the IκB kinases IKKα or IKKβ, and in cells treated with p65 small interfering RNA. Pretreatment with the IKK inhibitor PS-1145 also enhanced CDC of wild-type cells (WT) but not of p65(-/-) cells. Furthermore, reconstitution of p65 into p65(-/-) cells and overexpression of p65 in WT cells lowered their sensitivity to CDC. The postulated effect of p65 on the JNK-mediated death-signaling pathway activated by complement was examined. p65 small interfering RNA enhanced CDC in WT cells but not in cells lacking JNK. JNK phosphorylation induced by complement was more pronounced in p65(-/-) cells than in WT cells. The results indicate that the NF-κB pathway mediates cell resistance to CDC, possibly by suppressing JNK-dependent programmed necrotic cell death.     

A role for γS-crystallin in the organization of actin and fiber cell maturation in the mouse lens

γS-crystallin (γS) is a highly conserved component of the eye lens. To gain insights into the functional role(s) of this protein, the mouse gene (Crygs) was deleted. Although mutations in γS can cause severe cataracts, loss of function of γS in knockout (KO) mice produced no obvious lens opacity, but was associated with focusing defects. Electron microscopy showed no major differences in lens cell organization, suggesting that the optical defects are primarily cytoplasmic in origin. KO lenses were also grossly normal by light microscopy but showed evidence of incomplete clearance of cellular organelles in maturing fiber cells. Phalloidin labeling showed an unusual distribution of F-actin in a band of mature fiber cells in KO lenses, suggesting a defect in the organization or processing of the actin cytoskeleton. Indeed, in wild-type lenses, γS and F-actin colocalize along the fiber cell plasma membrane. Relative levels of F-actin and G-actin in wild-type and KO lenses were estimated from fluorescent staining profiles and from isolation of actin fractions from whole lenses. Both methods showed a two-fold reduction in the F-actin/G-actin ratio in KO lenses, whereas no difference in tubulin organization was detected. In vitro experiments showed that recombinant mouse γS can directly stabilize F-actin. This suggests that γS may have a functional role related to actin, perhaps in 'shepherding' filaments to maintain the optical properties of the lens cytoplasm and normal fiber cell maturation.     

The thymus and T-cell commitment: the right niche for Notch?

The current dogma is that the thymus is colonized by progenitors that retain the capacity to generate both T cells and B cells, and that intrathymic Notch signalling determines lineage choice so that T cells, rather than B cells, develop in the thymus. However, evidence is now accumulating to indicate that, at least during fetal life, this is not the case. Rather, it now seems that the fetal thymus is colonized by progenitors that have already made the T-cell versus B-cell lineage choice. We propose an alternative role for Notch signalling in the thymus, which is not to mediate this choice but instead to reveal it by supporting further T-cell differentiation in the thymic microenvironment.     

Temporal predisposition to αβ and γδ T cell fates in the thymus

How T cell progenitors engage into the γδ or αβ T cell lineages is a matter of intense debate. In this study, we analyzed the differentiation potential of single thymocytes from wild-type and TCRγδ-transgenic mice at two sequential early developmental stages. Double-negative (DN) 3 progenitors from both wild-type and transgenic mice retain the capacity to engage into both pathways, indicating that full commitment is only completed after this stage. More importantly, DN2 and DN3 progenitors from TCRγδ transgenic mice have strong biases for opposite fates, indicating that developmentally regulated changes, other than the production of a functional TCR, altered their likelihood to become a γδ or an αβ T cell. Thus, unlike the differentiation in other hematopoietic lineages, T cell progenitors did not restrict, but rather switch their differentiation potential as they developed.     

A mitotic role for GSK-3β kinase in Drosophila

Glycogen synthase kinase-3β (GSK-3β) is involved in a wide variety of cellular processes, and implicated in a growing list of human diseases. Recent drug inhibition studies have suggested a role for GSK-3β in mitosis in animals. Here, we take an alternative approach to understanding GSK-3β function in mitosis by genetic mutational analysis in Drosophila. GSK-3β function is well conserved between Drosophila (Zw3) and humans, frequently operating similarly in pathways, as diverse as the Wnt signaling and circadian rhythm pathways, and sharing a key role in the development of the neuromuscular junction. Unlike drug inhibitor studies, we find that loss of function mutations of zw3 result in markedly curved, or bent, metaphase spindles that exhibit metaphase delay. These defects do not routinely result in mitotic catastrophe, and argue that Zw3 plays a role in the maintenance of the mitotic spindle, rather than an essential role in spindle morphogenesis. Consistent with a mitotic function, we observe a complex and dynamic localization of Zw3 during cell division. These studies provide genetic data that validate and extend drug inhibition studies on a novel mitotic role for glycogen synthase kinase in the maintenance of the mitotic spindle.     

A role for polyamines in stimulus-secretion coupling in the pancreatic β-cell

Measurement of the content of polyamines in pancreatic islets indicated that no significant change in their concentration took place during glucose-stimulated insulin release. The finding, together with the absence of any effect of -difluoromethylornithine on glucosestimulated insulin release suggested that rapid synthesis of polyamines is not involved in stimulus-secretion coupling in the -cell. The concentration of polyamines found in islets were high enough for them to act as substrates for the Ca²⁺-dependent islet transglutaminase during insulin release. This was further demonstrated by the ability of islet transglutaminase to incorporate [¹⁴C]putrescine into proteins from islet homogenates and by the demonstration of an increase in the covalent incorporation of [¹⁴C]putrescine into the proteins of intact islets following their challenge with glucose. Unlike monoamine substrates of transglutaminase, putrescine failed to effectively inhibit insulin release when its intracellular concentration was increased. A role for polyamines in the secretory process through their incorporation into islet proteins is suggested.