Drugachenok, P. | bioRxiv Science

bioRxiv2025

CRISPR screen identifies CNIH1 as a selective driver of GPCR export

G protein-coupled receptors (GPCRs), the largest family of transmembrane proteins, transduce extracellular stimuli into intracellular signaling cascades to orchestrate human physiology. The transport of newly synthesized receptors from the endoplasmic reticulum (ER) to the plasma membrane (PM) determines cellular responsiveness to incoming ligands, yet the molecular machinery governing GPCR export remains incompletely defined. Here, we combine a synchronized cargo-release assay with a genome-wide CRISPR/Cas9 screen to systematically map regulators of GPCR ER-to-PM transport. Focusing on the {delta}-opioid receptor (DOR), a prototypical class A GPCR, we identify CNIH1 as a dedicated export factor. In the absence of CNIH1, DOR is retained intracellularly with immature glycosylation, and drives reduced PM signaling. CNIH1 localizes to both ER exit sites and the Golgi, promoting the anterograde transport of a subset of class A GPCRs. Opioid receptors directly interact with CNIH1 and require its putative COPII-binding site for export. Distinct from other human cornichon homologs, CNIH1 defines a selective GPCR-sorting receptor that couples GPCR biosynthesis to signaling competence.

cell biology↗

bioRxiv2025

Dopamine selectively regulates pediatric sclera/choroid interactions through the stimulation of exosome-associated retinoic acid

Postnatal eye growth is critical for healthy vision and yet, how this process is regulated at the cellular level is still unclear. The choroid is thought to play a crucial role in relaying signals from the retina to the sclera to modulate eye growth, but which cells are targeted and how they interact is not known. Using primary cultures of pediatric and adult human choroid and sclera stromal fibroblasts, we investigated the effect of choroid-conditioned medium (CCM) on scleral fibroblasts contractile activity. We show that CCM activates pediatric scleral fibroblast contraction, with age and antero-posterior location differences. Upon exposure to dopamine, a known negative regulator of eye growth, pediatric - but not adult - choroid cells lose their ability to stimulate scleral fibroblasts. Using RNA-Sequencing, we show that dopamine stimulates pathways linked to ribosomal activation, translation and exosome release exclusively in pediatric choroid cells. Removing exosomes from the CCM rescued the ability of dopamine-treated choroid cells to stimulate scleral fibroblasts. We further identify retinoic acid as the active exosome-associated compound preventing scleral fibroblast activation. Mechanistically, we show that CCM stimulates actomyosin-mediated protrusive activity in scleral fibroblasts. This is prevented when scleral cells are exposed to dopamine-CCM, and fully rescued upon exosome removal or ALDH1 inhibition. We thus propose that pediatric choroid stromal cells specifically relay dopamine-mediated retinal signals to the sclera during post-natal eye growth, operating through changes in secretome - including the production of exosome-associated retinoic acid - rather than gene expression changes. SIGNIFICANCEEye size is critical to optimal vision but the exact cellular and molecular mechanisms regulating it are still unknown. Failure to properly regulate postnatal eye growth leads to elongated eyes and myopia. Myopia is expected to affect half of the world population by 2050, with many at risk of blinding complications. Eye growth and homeostasis are supported by the sclera and the choroid, but how the two tissues interact to regulate eye growth is unknown. We use here human cells to demonstrate direct, age-specific, interactions between the two cell types and provide a rationale for the known negative effect of dopamine on eye growth, with significant implications for future studies as well as for the understanding and treatment of myopia.

cell biology↗

bioRxiv2024

Functional characterization of the store-operated calcium entry pathway in naked mole-rat cells

Naked mole-rats (NMRs, Heterocephalus glaber) are highly unusual rodents exhibiting remarkable adaptations to their subterranean habitat and resistance to developing various age-related diseases such as those related to abnormal cell proliferation or cancer, neurodegeneration and inflammation. In other rodents, as well as humans, a ubiquitous Ca2+ influx pathway, namely the store-operated Ca2+ entry (SOCE), has been implicated in all these diseases. SOCE is triggered by intracellular Ca2+ store depletion resulting in interaction of Stim proteins with Orai proteins, the putative homologs of which appear be present in the NMR genome, but no characterisation of SOCE in NMRs has yet been conducted. In this study, we provide the first functional and pharmacological characterization of SOCE in NMR cells using both excitable and non-excitable cells.

cell biology↗