Transcript odd
1. 2. Notch signaling and midline cell development Neuron/glia interactions; nrx Notch signaling is required for glia and the MNB Dl3 / Dl3 Wild- type MP1 cells (odd) MP3 cells (ple) VUM cells (Tbh) MNB (wor) Glia (wrapper) Only MP4 progeny are produced in Dl mutant embryos How does this phenotype arise? Model for early midline development 1. Mesectoderm specification 2. Cell fate specification Models for early Delta phenotype 1. Mesectoderm specification 2. Cell fate specification Single division 3 rounds of division and cell death Models for early Delta phenotype Single division 3 rounds of division and cell death Predictions ~15; 5 each of MP1, MP3, and MP4 All divisions in en masse at s11 # of MPs present at early stages Pattern of division ~3; 1 each of MP1, MP3, and MP4 Multiple divisions from s11 to s14 Experiments: Compare wild type and Dl mutants at the earliest possible stages 1. Fixed samples 2. Live imaging Genetics 1. (f) w; sim-gal4,UAS-tauGFP sim-gal4, UAS-tauGFP/+ X ; (m) Dl[3]/TM6b Dl[3]/+ or TM6b/+ 2. (f,m) sim-gal4, UAS-tauGFP/+ ; Dl[3]/+ sim-gal4, UAS-tauGFP/ sim-gal4, UAS-tauGFP (sim-gal4, UAS-tauGFP/+)x2 +/+ ¾ progeny have at least 1 copy ¼ have 2 copies ; Dl[3]/Dl[3] Dl[3]/+ +/+ ¼ are Dl[3] Odd-skipped identifies multiple MP1s in Dl mutants Odd and Cas identify 3 distinct MP populations in Dl mutants Odd Cas Tentative conclusions: 1. ~5 each of MP1, MP3, and MP4 are specified in Dl mutant embryos. Additional experiments: Go to LSM for stack 1. Look at more segments stained with Odd and Cas 2. Ask if the MP6 is present by staining with Cas and Tkr (likely Cas- Tkr+). 3. Establish pattern of MP division using pH3, Odd, Cas and sim-Gal4 UAS-tauGFP as well as using live imaging. MP1 divisions likely occur in close succession Pros Odd Pros Odd Go to LSM for stack Tentative model 1. Mesectoderm specification Some additional questions: 1. Which cell becomes the MP? What are the txn factors and/or signaling that selects the MP? Candidate genes – odd, cas 2. Cell fate specification equivalence groups 2. How are the MP5, MP6, and MNB specified? 3. How are non-MP cells directed to the AMG or PMG fate? What to do next 1. hedgehog and wingless are doing something, what is it? a. Maybe required to confer anterior and posterior identity (ie. MP1,3,4 vs MP5,6, MNB or AMG vs PMG) b. phenotypic analysis of wg and hh mutants c. misexpression of constitutively active downstream effectors (Tcf.VP16 , Ci.VP16, and others). d. Reporters of activity (nuclear arm, ptc expression) What to do next (cont.) 2. How is glial gene transcription regulated? a. What are the dynamics of glial transcription? i. examine the expression patterns of genes with de novo expression in midline glia (~17 genes) ii. b. What is the role of Notch signaling? i. isolate enhancers for several glial genes and mutate binding sites for Suppressor of Hairless. ii. c. Identify temporal and spatial patterns that may give clues to regulatory hierarchy. Examine expression in Dl mutants. What is the role of single-minded? I. isolate enhancers for several glial genes and mutate binding sites for Sim. II. Examine expression of reporters in sim mutants. 1. 2. Notch signaling and midline cell development Neuron/glia interactions; nrx