Cytoneme-mediated signaling essential for tumorigenesis.

Communication between neoplastic cells and cells of their microenvironment is critical to cancer progression. To investigate the role of cytoneme-mediated signaling as a mechanism for distributing growth factor signaling proteins between tumor and tumor-associated cells, we analyzed EGFR and RET Drosophila tumor models and tested several genetic loss-of-function conditions that impair cytoneme-mediated signaling. Neuroglian, capricious, Irk2, SCAR, and diaphanous are genes that cytonemes require during normal development. Neuroglian and Capricious are cell adhesion proteins, Irk2 is a potassium channel, and SCAR and Diaphanous are actin-binding proteins, and the only process to which they are known to contribute jointly is cytoneme-mediated signaling. We observed that diminished function of any one of these genes suppressed tumor growth and increased organism survival. We also noted that EGFR-expressing tumor discs have abnormally extensive tracheation (respiratory tubes) and ectopically express Branchless (Bnl, a FGF) and FGFR. Bnl is a known inducer of tracheation that signals by a cytoneme-mediated process in other contexts, and we determined that exogenous over-expression of dominant negative FGFR suppressed tumor growth. Our results are consistent with the idea that cytonemes move signaling proteins between tumor and stromal cells and that cytoneme-mediated signaling is required for tumor growth and malignancy

Expression profiling of Drosophila imaginal discs

BACKGROUND: In the Drosophila larva, imaginal discs are programmed to produce adult structures at metamorphosis. Although their fate is precisely determined, these organs remain largely undifferentiated in the larva. To identify genes that establish and express the different states of determination in discs and larval tissues, we used DNA microarrays to analyze mRNAs isolated from single imaginal discs. RESULTS: Linear amplification protocols were used to generate hybridization probes for microarray analysis from poly(A)(+) RNA from single imaginal discs containing between 10,000 and 60,000 cells. Probe reproducibility and degree of representation were tested using microarrays with approximately 6,000 different cDNAs. Hybridizations with probes that had been prepared separately from the same starting RNA pool had a correlation coefficient of 0.97. Expression-profile comparisons of the left and right wing imaginal discs from the same larva correlated with a coefficient of 0.99, indicating a high degree of reproducibility of independent amplifications. Using this method, we identified genes with preferential expression in the different imaginal discs using pairwise comparisons of discs and larval organs. Whereas disc-to-disc comparisons revealed only moderate differences, profiles differed substantially between imaginal discs and larval tissues, such as larval endodermal midgut and mesodermal fat body. CONCLUSION: The combination of linear RNA amplification and DNA microarray hybridization allowed us to determine the expression profiles of individual imaginal discs and larval tissues and to identify genes expressed in tissue-specific patterns. These methods should be widely applicable to comparisons of expression profiles for tissues or parts of tissues that are available only in small amounts

An Experiment in Visual Ethnography

This paper is an output from my attendance at an ESRC-funded ‘Live Sociology’ course at Goldsmiths College, London in 2006. The material here is based on a photography exercise and on discussions that took place during the workshop sessions.This paper discusses a one-day exercise in visual ethnography using a digital camera to take photographs of Deptford in South East London. For me, this represented an experiment in using photography in social research.ESR

Broadly conserved roles of TMEM131 family proteins in intracellular collagen assembly and secretory cargo trafficking.

Collagen is the most abundant protein in animals. Its dysregulation contributes to aging and many human disorders, including pathological tissue fibrosis in major organs. How premature collagen proteins in the endoplasmic reticulum (ER) assemble and route for secretion remains molecularly undefined. From an RNA interference screen, we identified an uncharacterized Caenorhabditis elegans gene tmem-131, deficiency of which impairs collagen production and activates ER stress response. We find that amino termini of human TMEM131 contain bacterial PapD chaperone-like domains, which recruit premature collagen monomers for proper assembly and secretion. Carboxy termini of TMEM131 interact with TRAPPC8, a component of the TRAPP tethering complex, to drive collagen cargo trafficking from ER to the Golgi. We provide evidence that previously undescribed roles of TMEM131 in collagen recruitment and secretion are evolutionarily conserved in C. elegans, Drosophila, and humans

Constraints, Histones, and the 30 Nanometer Spiral

We investigate the mechanical stability of a segment of DNA wrapped around a histone in the nucleosome configuration. The assumption underlying this investigation is that the proper model for this packaging arrangement is that of an elastic rod that is free to twist and that writhes subject to mechanical constraints. We find that the number of constraints required to stabilize the nuclesome configuration is determined by the length of the segment, the number of times the DNA wraps around the histone spool, and the specific constraints utilized. While it can be shown that four constraints suffice, in principle, to insure stability of the nucleosome, a proper choice must be made to guarantee the effectiveness of this minimal number. The optimal choice of constraints appears to bear a relation to the existence of a spiral ridge on the surface of the histone octamer. The particular configuration that we investigate is related to the 30 nanometer spiral, a higher-order organization of DNA in chromatin.Comment: ReVTeX, 15 pages, 18 figure

Generalized myoclonic epilepsy with photosensitivity in juvenile dogs caused by a defective DIRAS family GTPase 1

The clinical and electroencephalographic features of a canine generalized myoclonic epilepsy with photosensitivity and onset in young Rhodesian Ridgeback dogs (6 wk to 18 mo) are described. A fully penetrant recessive 4-bp deletion was identified in the DIRAS family GTPase 1 (DIRAS1) gene with an altered expression pattern of DIRAS1 protein in the affected brain. This neuronal DIRAS1 gene with a proposed role in cholinergic transmission provides not only a candidate for human myoclonic epilepsy but also insights into the disease etiology, while establishing a spontaneous model for future intervention studies and functional characterization

Different forms of TFIIH for transcription and DNA repair: Holo-TFIIH and a nucleotide excision repairosome

AbstractYeast TFIIH that is active in transcription can be dissociated into three components: a 5-subunit core, the SSL2 gene product, and a complex of 47 kDa, 45 kDa, and 33 kDa polypeptides that possesses protein kinase activity directed towards the C-terminal repeat domain of RNA polymerase II. These three components can reconstitute fully functional TFIIH, and all three are required for transcription in vitro. By contrast, TFIIH that is highly active in nucleotide excision repair (NER) lacks the kinase complex and instead contains the products of all other genes known to be required for NER in yeast: RAD1, RAD2, RAD4, RAD10, and RAD14. This repairosome is not active in reconstituted transcription in vitro and is significantly more active than any of the constituent polypeptides in correcting defective repair in extracts from strains mutated in NER genes

A translational signature for nucleosome positioning in vivo

In vivo nucleosomes often occupy well-defined preferred positions on genomic DNA. An important question is to what extent these preferred positions are directly encoded by the DNA sequence itself. We derive here from in vivo positions, accurately mapped by partial micrococcal nuclease digestion, a translational positioning signal that identifies the approximate midpoint of DNA bound by a histone octamer. This midpoint is, on average, highly A/T rich (∼73%) and, in particular, the dinucleotide TpA occurs preferentially at this and other outward-facing minor grooves. We conclude that in this set of sequences the sequence code for DNA bending and nucleosome positioning differs from the other described sets and we suggest that the enrichment of AT-containing dinucleotides at the centre is required for local untwisting. We show that this signature is preferentially associated with nucleosomes flanking promoter regions and suggest that it contributes to the establishment of gene-specific nucleosome arrays

Clinical Course and Diagnostic Findings of Biopsy Controlled Presumed Immune-Mediated Polyneuropathy in 70 European Cats

There is a paucity of information on the clinical course and outcome of young cats with polyneuropathy. The aim of the study was to describe the clinical features, diagnostic investigations, and outcome of a large cohort of cats with inflammatory polyneuropathy from several European countries. Seventy cats with inflammatory infiltrates in intramuscular nerves and/or peripheral nerve biopsies were retrospectively included. Information from medical records and follow up were acquired via questionnaires filled by veterinary neurologists who had submitted muscle and nerve biopsies (2011–2019). Median age at onset was 10 months (range: 4–120 months). The most common breed was British short hair (25.7%), followed by Domestic short hair (24.3%), Bengal cat (11.4%), Maine Coon (8.6%) and Persian cat (5.7%), and 14 other breeds. Male cats were predominantly affected (64.3%). Clinical signs were weakness (98.6%) and tetraparesis (75.7%) in association with decreased withdrawal reflexes (83.6%) and, less commonly, cranial nerve signs (17.1%), spinal pain/hyperesthesia (12.9%), and micturition/defecation problems (14.3%). Onset was sudden (30.1%) or insidious (69.1%), and an initial progressive phase was reported in 74.3%. Characteristic findings on electrodiagnostic examination were presence of generalized spontaneous electric muscle activity (89.6%), decreased motor nerve conduction velocity (52.3%), abnormal F-wave studies (72.4%), pattern of temporal dispersion (26.1%) and unremarkable sensory tests. The clinical course was mainly described as remittent (49.2%) or remittent-relapsing (34.9%), while stagnation, progressive course or waxing and waning were less frequently reported. Relapses were common and occurred in 35.7% of the cats' population. An overall favorable outcome was reported in 79.4% of patients. In conclusion, young age at the time of diagnosis and sudden onset of clinical signs were significantly associated with recovery (p < 0.05). Clinical and electrodiagnostic features and the remittent-relapsing clinical course resembles juvenile chronic inflammatory demyelinating polyneuropathy (CIDP), as seen in human (children/adolescents), in many aspects