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Pathway sensor-based functional genomics screening identifies modulators of neuronal activity.

Alexander Herholt1,2, Ben Brankatschk2, Nirmal Kannaiyan2

  • 1Department of Psychiatry and Psychotherapy, University Hospital, LMU Munich, Munich, Germany.

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Summary

Researchers developed pathScreener, a novel pooled screening assay for dissecting neuronal signalling pathways. This method identifies key genes regulating neuronal activity and may reveal new targets for brain disorders.

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Area of Science:

  • Neuroscience
  • Genomics
  • Molecular Biology

Background:

  • Neuronal signal transduction is crucial for brain function, and its dysregulation is linked to mental disorders.
  • Existing functional genomics screens are limited for studying neuronal signaling in postmitotic neurons.

Purpose of the Study:

  • To develop a novel pooled screening assay for comprehensive analysis of neuronal signaling pathways.
  • To identify genes modulating neuronal activity using a high-throughput genetic screen.

Main Methods:

  • Developed pathScreener, a pooled screening assay using barcoded activity reporters and pooled genetic perturbation via dual-expression adeno-associated virus (AAV) libraries.
  • Utilized cisProfiler, a barcoded and multiplexed reporter assay, to identify the synaptic-activity responsive element (SARE) as a sensor for neuronal activity.
  • Screened over 4,400 genes for their effects on SARE activity in primary cortical neurons.

Main Results:

  • Identified numerous known genes involved in glutamatergic synapse-to-nucleus signaling with high reproducibility.
  • Validated a subset of identified genes using orthogonal assays.
  • Discovered novel associations between genes like Ptch2 and Ift57 (hedgehog signaling pathway) and neuronal activity regulation.

Conclusions:

  • The pathScreener assay effectively dissects signaling pathways in postmitotic neurons, overcoming limitations of previous methods.
  • This approach enhances the analysis of neuronal signaling and provides a valuable tool for identifying novel therapeutic targets for brain disorders.