Proximity interactome mapping furthers a role for spinophilin in protein homeostasis.

Spinophilin is a dendritic spine-enriched scaffolding and protein phosphatase 1-targeting protein. To detail spinophilin-interacting proteins, we created an UltraID and ALFA-tagged spinophilin-encoding construct that permits proximity labeling and orthogonal nanobody pull-down (ID-oPD) of spinophilin-associated protein complexes in heterologous cells. Using this ID-oPD approach, we identified 614 UltraID-specific and 312 UltraID-specific and spinophilin-selective spinophilin-interacting proteins in HEK293 cells and validated a subset of these using orthogonal approaches. Many of these proteins are involved in mRNA processing and translation. In the brain, we determined that spinophilin mRNA is highly neuropil localized and that spinophilin may normally function to limit its own expression but promote the expression of other postsynaptic density (PSD)-associated proteins. Overall, our use of an ID-oPD approach uncovers a putative role for spinophilin in mRNA translation and PSD protein expression.