Recombinant Drosophila melanogaster Protein trapped in endoderm-1 (Tre1)
Product Specs
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Note: Our proteins are typically shipped with standard blue ice packs. If you require dry ice shipping, please inform us in advance, as additional fees will apply.
Generally, the shelf life of liquid form is 6 months at -20°C/-80°C. The shelf life of lyophilized form is 12 months at -20°C/-80°C.
Target Background
- Bifurcation of the Tre1 G protein-coupled receptor signaling response via G protein-dependent and independent branches enables distinct spatiotemporal regulation of germ cell migration. PMID: 28687666
- Feminizing the Tre1-expressing neurons in male flies leads to rapid courtship initiation, a phenotype that contributes to increased reproductive success. Tre1 is expressed in a sexually dimorphic pattern in olfactory sensory neurons and the central nervous system. PMID: 26721856
- An investigation of the role of the arginine within the NRY motif in Tre1 suggests that a salt bridge may form between this critical arginine and an aspartic acid in transmembrane alpha-helix (TM6) in Tre1. PMID: 24044607
- The scattershot allele, characterized by an in-frame deletion of 8 amino acids at the junction of the third transmembrane domain and the second intracellular loop of Tre1, significantly impairs the function of this GPCR in germ cell migration. PMID: 20676220
- Tre1 plays a significant role in regulating germ cell migration, polarity, cell adhesion, and cadherin binding. (review) PMID: 20058705
- Tre1 serves as a G protein-coupled receptor that directs transepithelial migration of Drosophila germ cells. PMID: 14691551
- Down-regulation of E-cadherin leads to germ cell dispersal but is insufficient for transepithelial migration in the absence of Tre1, suggesting a novel mechanism for Tre1 GPCR function that connects cell polarity, modulation of cell adhesion, and invasion. PMID: 18824569
Q&A
What is the role of Tre1 in Drosophila germ cell migration, and how is it experimentally validated?
Tre1 orchestrates GC navigation by polarizing actin dynamics through localized PI(4,5)P₂ production and Wiskott-Aldrich syndrome protein (WASP)-mediated actin polymerization. Key experimental approaches include:
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CRISPR/Cas9-generated null alleles: Complete Tre1 knockout (KO) alleles, such as Tre1ᴷᴼ, exhibit ~90% GC migration failure due to misorientation of F-actin protrusions .
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Live-cell imaging: Tracking GC trajectories in Tre1ᴷᴼ mutants reveals random motility patterns, contrasting with directed migration in wild-type (WT) embryos .
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Localization studies: Tre1 co-localizes with F-actin, PI(4,5)P₂, and dPIP5K at migration fronts, validated via immunofluorescence and tagged constructs (e.g., Tre1-GFP) .
Table 1: Phenotypic Outcomes of Tre1 Mutants
Which signaling pathways interact with Tre1, and how are these interactions assayed?
Tre1 integrates guidance cues from Hh signaling through Smo. Methodologies to map these interactions include:
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Co-immunoprecipitation (Co-IP): Demonstrates physical association between Tre1 and Smo in GC lysates .
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Pharmacological inhibition: Cyclopamine (Smo inhibitor) reduces Tre1 membrane localization, impairing F-actin polarization .
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Genetic epistasis: Smo RNAi rescues Tre1ᴷᴼ migration defects, confirming functional interdependence .
How can researchers resolve contradictions in Tre1’s reported roles in cell adhesion vs. guidance?
Conflicting studies attribute Tre1 mutant GC dispersal to either failed adhesion dissolution or misguided motility . A tripartite analysis (TriPA) framework is recommended:
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Case-by-case integration: Compare adhesion molecule (e.g., E-cadherin) dynamics in Tre1ᴷᴼ vs. WT GCs using time-lapse microscopy .
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Quantitative-qualitative merging: Quantify E-cadherin fluorescence intensity and correlate with GC trajectory data .
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Pattern recognition: Identify whether adhesion loss precedes or follows motility defects .
Table 2: Conflicting Interpretations of Tre1 Function
| Study | Proposed Mechanism | Key Evidence |
|---|---|---|
| Kunwar et al. (2003) | Adhesion dissolution | E-cadherin downregulation in Tre1 mutants |
| Lin et al. (2020) | Guidance cue sensing | Random Rho1/MyoII oscillations in mutants |
| Kim et al. (2021) | PI(4,5)P₂-dependent polarization | Tre1-dPIP5K co-localization at protrusions |
What methodologies optimize detection of Tre1 in vivo, given its low expression and rapid turnover?
Tre1’s transient membrane localization necessitates:
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Stabilized tagged constructs: Tre1-GFP (half-life >6 hrs) outperforms Tre1-FLAG/HA in pulse-chase assays .
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Cycloheximide treatment: Confirms Tre1-GFP resistance to degradation in S2R+ cells .
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Super-resolution microscopy: Resolve Tre1 nanodomains at protrusions using STED or PALM .
How can multi-omics data be integrated to map Tre1’s regulatory network?
A multi-tiered approach is critical:
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Transcriptomics: RNA-seq of Tre1ᴷᴼ GCs identifies downstream effectors (e.g., dWIP, Rho1) .
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Lipidomics: MALDI-TOF detects PI(4,5)P₂ spatial gradients in migrating GCs .
What controls are essential for Tre1 loss-of-function studies?
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Maternal-zygotic mutants: Distinguish maternal vs. zygotic Tre1 contributions .
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Rescue assays: Express tagged Tre1 in mutants to confirm phenotype reversibility .
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Off-target controls: Use dual sgRNAs in CRISPR to exclude false positives .
How should researchers address partial penetrance in Tre1 mutant phenotypes?
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Quantitative motility metrics: Track GC speed, directionality, and protrusion dynamics using Fiji/ImageJ .
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Environmental standardization: Control embryo staging and incubation temperature to reduce variability .
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Statistical modeling: Apply mixed-effects models to account for clutch-to-clutch variation .
