egaku: Enhancing the Sketching Process

Architects sketch using a translucent vellum tracing paper with a thick pencil or marker. The translucency of the paper allows architects to employ a layer-drawing technique for the exploration of ideas derived from their basic design. For example, working with a single base layer such as a map of the site, architects can design upwards of hundreds of possible variations. This ultimately leads to a great pile of drawings, which compose the piles of papers typically strewn about an architecture studio. Individually, these “referential” sketches represent small pieces of a much larger design concept [Graves 1977]. Although they are valuable,they are often cumbersome to manage during the ideation process because it interrupts the flow of ideation, and even difficult to understand when a single sketch is taken out of associated sketches

Group 2 Innate Lymphoid Cells Exhibit Tissue-Specific Dynamic Behaviour During Type 2 Immune Responses.

Group 2 innate lymphoid cells (ILC2s) are early effectors of mucosal type 2 immunity, producing cytokines such as interleukin (IL)-13 to mediate responses to helminth infection and allergen-induced inflammation. ILC2s are also present in lymph nodes (LNs) and can express molecules required for antigen presentation, but to date there are limited data on their dynamic behaviour. We used a CD2/IL-13 dual fluorescent reporter mouse for in vivo imaging of ILC2s and Th2 T cells in real time following a type 2 priming helminth infection or egg injection. After helminth challenge, we found that ILC2s were the main source of IL-13 in lymphoid organs (Peyer's patches and peripheral LNs), and were located in T cell areas. Intravital imaging demonstrated an increase in IL-13+ ILC2 size and movement following helminth infection, but reduced duration of interactions with T cells compared with those in homeostasis. In contrast, in the intestinal mucosa, we observed an increase in ILC2-T cell interactions post-infection, including some of prolonged duration, as well as increased IL-13+ ILC2 movement. These data suggest that ILC2 activation enhances cell motility, with the potential to increase the area of distribution of cytokines to optimise the early generation of type 2 responses. The prolonged ILC2 interactions with T cells within the intestinal mucosa are consistent with the conclusion that contact-based T cell activation may occur within inflamed tissues rather than lymphoid organs. Our findings have important implications for our understanding of the in vivo biology of ILC2s and the way in which these cells facilitate adaptive immune responses

Identification of a Thymic Epithelial Cell Subset Sharing Expression of the Class Ib HLA-G Molecule with Fetal Trophoblasts

HLA-G is the only class I determinant of the major histocompatibility complex (MHC) expressed by the trophoblasts, the fetal cells invading the maternal decidua during pregnancy. A unique feature of this nonclassical HLA molecule is its low polymorphism, a property that has been postulated to play an important role in preventing local activation of maternal alloreactive T and natural killer cells against the fetus. Yet, the mechanisms by which fetal HLA-G can be recognized as a self-MHC molecule by the maternal immune system remain unclear. Here we report the novel observation that HLA-G is expressed in the human thymus. Expression is targeted to the cell surface of thymic medullary and subcapsular epithelium. Thymic epithelial cell lines were generated and shown to express three alternatively spliced HLA-G transcripts, previously identified in human trophoblasts. Sequencing of HLA-G1 transcripts revealed a few nucleotide changes resulting in amino acid substitutions, all clustered within exon 3 of HLA-G, encoding for the α2 domain of the molecule. Our findings raise the possibility that maternal unresponsiveness to HLA-G–expressing fetal tissues may be shaped in the thymus by a previously unrecognized central presentation of this MHC molecule on the medullary epithelium

Hydrophobicity and Charge Shape Cellular Metabolite Concentrations

What governs the concentrations of metabolites within living cells? Beyond specific metabolic and enzymatic considerations, are there global trends that affect their values? We hypothesize that the physico-chemical properties of metabolites considerably affect their in-vivo concentrations. The recently achieved experimental capability to measure the concentrations of many metabolites simultaneously has made the testing of this hypothesis possible. Here, we analyze such recently available data sets of metabolite concentrations within E. coli, S. cerevisiae, B. subtilis and human. Overall, these data sets encompass more than twenty conditions, each containing dozens (28-108) of simultaneously measured metabolites. We test for correlations with various physico-chemical properties and find that the number of charged atoms, non-polar surface area, lipophilicity and solubility consistently correlate with concentration. In most data sets, a change in one of these properties elicits a ∼100 fold increase in metabolite concentrations. We find that the non-polar surface area and number of charged atoms account for almost half of the variation in concentrations in the most reliable and comprehensive data set. Analyzing specific groups of metabolites, such as amino-acids or phosphorylated nucleotides, reveals even a higher dependence of concentration on hydrophobicity. We suggest that these findings can be explained by evolutionary constraints imposed on metabolite concentrations and discuss possible selective pressures that can account for them. These include the reduction of solute leakage through the lipid membrane, avoidance of deleterious aggregates and reduction of non-specific hydrophobic binding. By highlighting the global constraints imposed on metabolic pathways, future research could shed light onto aspects of biochemical evolution and the chemical constraints that bound metabolic engineering efforts

Tropomyosin 1: multiple roles in the developing heart and in the formation of congenital heart defects

Tropomyosin 1 (TPM1) is an essential sarcomeric component, stabilising the thin filament and facilitating actin's interaction with myosin. A number of sarcomeric proteins, such as alpha myosin heavy chain, play crucial roles in cardiac development. Mutations in these genes have been linked to congenital heart defects (CHDs), occurring in approximately 1 in 145 live births. To date, TPM1 has not been associated with isolated CHDs. Analysis of 380 CHD cases revealed three novel mutations in the TPM1 gene; IVS1 + 2T > C, I130V, S229F and a polyadenylation signal site variant GATAAA/AATAAA. Analysis of IVS1 + 2T > C revealed aberrant pre-mRNA splicing. In addition, abnormal structural properties were found in hearts transfected with TPM1 carrying I130V and S229F mutations. Phenotypic analysis of TPM1 morpholino-treated embryos revealed roles for TPM1 in cardiac looping, atrial septation and ventricular trabeculae formation and increased apoptosis was seen within the heart. In addition, sarcomere assembly was affected and altered action potentials were exhibited. This study demonstrated that sarcomeric TPM1 plays vital roles in cardiogenesis and is a suitable candidate gene for screening individuals with isolated CHDs

Genetic analysis reveals a hierarchy of interactions between polycystin-encoding genes and genes controlling cilia function during left-right determination

During mammalian development, left-right (L-R) asymmetry is established by a cilia-driven leftward fluid flow within a midline embryonic cavity called the node. This ‘nodal flow’ is detected by peripherally-located crown cells that each assemble a primary cilium which contain the putative Ca2+ channel PKD2. The interaction of flow and crown cell cilia promotes left side-specific expression of Nodal in the lateral plate mesoderm (LPM). Whilst the PKD2-interacting protein PKD1L1 has also been implicated in L-R patterning, the underlying mechanism by which flow is detected and the genetic relationship between Polycystin function and asymmetric gene expression remains unknown. Here, we characterize a Pkd1l1 mutant line in which Nodal is activated bilaterally, suggesting that PKD1L1 is not required for LPM Nodal pathway activation per se, but rather to restrict Nodal to the left side downstream of nodal flow. Epistasis analysis shows that Pkd1l1 acts as an upstream genetic repressor of Pkd2. This study therefore provides a genetic pathway for the early stages of L-R determination. Moreover, using a system in which cultured cells are supplied artificial flow, we demonstrate that PKD1L1 is sufficient to mediate a Ca2+ signaling response after flow stimulation. Finally, we show that an extracellular PKD domain within PKD1L1 is crucial for PKD1L1 function; as such, destabilizing the domain causes L-R defects in the mouse. Our demonstration that PKD1L1 protein can mediate a response to flow coheres with a mechanosensation model of flow sensation in which the force of fluid flow drives asymmetric gene expression in the embryo