Multicellular organisms are equipped with an array of G-protein-coupled receptors (GPCRs) that mediate cell–cell signaling allowing them to adapt to environmental cues and ultimately survive. This is mechanistically possible through complex intracellular GPCR machinery that encompasses a vast network of proteins. Within this network, there is a group called scaffolding proteins that facilitate proper localization of signaling proteins for a quick and robust GPCR response. One protein family within this scaffolding group is the PSD-95/Dlg/ZO-1 (PDZ) family which is important for GPCR localization, internalization, recycling, and downstream signaling. Although the PDZ family of proteins regulate the functions of several receptors, this chapter focuses on a subfamily within the PDZ protein family called the Na+/H+ exchanger regulatory factors (NHERFs). Here we extensively review the predominantly characterized roles of NHERFs in renal phosphate absorption, intestinal ion regulation, cancer progression, and immune cell functions. Finally, we discuss the future perspectives and possible clinical application of targeting NHERFs in several disorders.