Protein liquid-liquid phase separation (LLPS), a pivotal phenomenon intricately linked to cellular processes, is regulated by various other proteins. However, there is still a lack of high-throughput methods for screening protein regulators of LLPS in target proteins. Here, we developed a CRISPR/Cas9-based screening method to identify protein phase separation regulators by integrating bimolecular fluorescence complementation (BiFC) and fluorescence-activated cell sorting (FACS). Using this newly developed method, we screened the RNA-binding proteins that regulate PABPN1 phase separation and identified the tumor suppressor QKI as a promoter of PABPN1 phase separation. Furthermore, QKI exhibits decreased expression levels and diminished nuclear localization in colorectal cancer cells, resulting in reduced PABPN1 phase separation, which, in turn, promotes alternative polyadenylation (APA), cell proliferation, and migration in colorectal cancer.
Humans ; Colorectal Neoplasms/genetics* ; RNA-Binding Proteins/genetics* ; Poly(A)-Binding Protein I/genetics* ; CRISPR-Cas Systems ; Flow Cytometry ; Cell Proliferation ; Cell Line, Tumor ; Cell Movement

Glutamine,as the most abundant free amino acid in the human body,plays a crucial role in various physiological processes.These functions include providing energy to cells,supporting immune system function,maintaining intestinal health,and playing a key role in nitrogen transport.Under normal physiological conditions,muscle tissue is the primary site for the synthesis and release of glutamine.After glutamine is released into the bloodstream,it will be transported to the intestines,immune organs or other tissues and play related roles.However,in a state of illness,especially in the presence of tumors,there are significant changes in the metabolism and function of glutamine.The glutamine metabolism can serve as signaling molecules and participates in regulating various signaling pathways within tumor cells,including key pathways that affect cell growth,survival,and metabolism,such as the mTOR and AMPK pathways.This paper reviews the metabolic steps involving glutamine in tumor cells and the role of glutamine blockade in the tumor microenvironment.