Having identified promising intrinsic and extrinsic molecules and molecular circuits that could be crucial for IM ID and lung homeostasis, we will formally investigate their roles in vivo. Conventional models of conditional gene deletion in IM or niche cells will be used on the one hand, to the extent of the resources available. On the other hand, we are currently developing and implementing a CRISP-seq-based strategy in order to formally and simultaneously test the function of many gene products, alone or in combination. In brief, the CRISP-seq strategy combines CRISPR-based genome editing with a single-cell RNA-sequencing read-out, thus allowing the profiling of the perturbation (i.e. in our case, the IM-specific knock-out of genes coding for transcription factors or receptors binding niche-derived signals) and of the transcriptome in single IM. We have already achieved several milestones in the implementation of CRISP-seq and want to succeed in this innovative method and model that is beyond the state-of-the-art. By the end of the project, we expect to identify the main niche signals activating specific transcription factors in IM subsets, thus endowing them with a particular identity and sustaining lung homeostasis.