Though myriad tools exist for enacting and reading out perturbations at the DNA and RNA levels, the field of functional genomics has until recently lacked scalable approaches to manipulate proteins, the main effectors of many cellular functions. Here we describe a technology to endogenously tag many genes at one time in a pool of cells, followed by in situ sequencing and computational image analysis, to enable the simultaneous characterization of hundreds of proteins. We then leverage the multifunctional tag to acutely induce protein misfolding in different organelles and characterize compartment-specific protein quality control responses. To enable this work, we first developed a locus-generic and homology-independent approach for endogenous tagging that could be applied at scale. The tag is encoded as a synthetic exon, which is inserted into a Cas9-mediated cut in the intron of the target protein; the tag is consequently transcribed and translated as part of the endogenous coding sequence, thus maintaining near-endogenous regulation of the tagged protein. Unlike tagging methods that require homology-containing reagents, the only locus-specific component is the intron-targeting sgRNA, which confers affordability and scalability to our approach. We adapted this technology to generate cell pools in which each cell harbors a multi-functional HaloTag in one endogenous protein. Treating the cell pool with a fluorescent HaloTag ligand and performing in situ optical pooled sequencing of the intron-targeting sgRNA, which is expressed from the genome of the tagged cell, enabled us to quantify the abundance and specific subcellular localization of every tagged protein. After curating a set of tag sites with high-fidelity localization patterns, we treated a targeted library with a hydrophobic HaloTag ligand to induce compartment-restricted unfolded protein stress. Single-cell RNA-sequencing readouts and additional functional studies allowed us to assign protein quality control roles to previously uncharacterized genes and to determine which protein quality control systems were active in different cell compartments. This work provides the first example of pooled endogenous protein tagging and perturbation as a novel and powerful functional genomics approach to understand the proteome and protein quality control