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Updated: Jul 11, 2025
Interactome-Seq: A Protocol for Domainome Library Construction, Validation and Selection by Phage Display and Next Generation Sequencing
Published on: October 3, 2018
Combined Multiplexed Phage Display, High-Throughput Sequencing, and Functional Assays as a Platform for Identifying
Anielka Zehnaker1, Amandine Vallet1, Juliette Gourdon1
1Physiologie de la Reproduction et des Comportements (PRC), Institut National de Recherche Pour l'Agriculture, l'Alimentation et l'Environnement (INRAE), Centre National de la Recherche Scientifique (CNRS), Université de Tours, 37380 Nouzilly, France.
Researchers developed novel antibodies (VHHs) targeting the follicle-stimulating hormone receptor (FSHR). A new selection strategy rapidly identified PRC1, a VHH that positively modulates FSHR activity, offering a potential tool for reproductive research.
Area of Science:
- Biochemistry
- Molecular Biology
- Immunology
Background:
- Developing modulatory antibodies for G protein-coupled receptors (GPCRs), such as the follicle-stimulating hormone receptor (FSHR), presents significant challenges.
- FSHR is a key regulator of reproductive functions.
Purpose of the Study:
- To develop a novel strategy for identifying functional modulatory antibodies targeting the FSHR.
- To isolate and characterize novel variable domains of heavy chain-only antibodies (VHHs) that modulate FSHR activity.
Main Methods:
- Construction and screening of two immune VHH libraries using multiplexed phage display.
- Next-generation sequencing for identifying enriched VHH sequences.
- Functional assessment of VHHs using a cAMP response element (CRE)-dependent reporter gene assay.
Main Results:
- Identification of 34 clusters of specifically enriched VHH sequences.
- 23 VHHs demonstrated significant positive or negative modulation of FSH-induced responses.
- The VHH cluster PRC1 specifically binds to human FSHR and potentiates FSH-induced cAMP production and Gs recruitment.
Conclusions:
- The improved selection strategy is effective for rapidly identifying functionally active VHHs against challenging membrane receptors.
- PRC1 is the first identified positive modulator VHH for the human FSHR, with potential applications in reproductive medicine.
