Assembly of an endocytic clathrin-coated pit (CCP) at the plasma membrane depends on the recruitment of the AP-2 adaptor protein complex and clathrin triskelions to the lipid bilayer (reviewed in McMahon and Boucrot, 2011; Robinson, 2015). Transient interactions between the plasma membrane-enriched lipid phosphatidlyinositol 4,5-bisphosphate (PI(4,5)P2) and AP-2 initiate coated pit formation (Beck et al, 1991; Honing et al, 2005; Loerke et al, 2009; Cocucci et al, 2012). A proportion of the transient complexes between AP-2, clathrin and the plasma membrane are rapidly stabilized by the recruitment of a number of proteins, including FCHo proteins, intersectins (ITSNs), EPS15 and SGIP1 among others (Henne et al, 2010; Stimpson et al, 2009; Reider et al, 2009; Cocucci et al, 2012; reviewed in McMahon and Boucrot, 2011). Many of these early players in CCP formation bind both to the plasma membrane and to the AP-2 complex and/or clathrin.
CCP formation is a highly heterogeneous and dynamic process and includes abortive initiation of nearly half of nascent CCPs (Loerke et al, 2009; Aguet et al, 2013). Heterogeneity is in part the result of the widely varied cargo proteins, which compete for a limited number of interaction hubs on AP-2 and clatrhin and influence the other protein components of the CCPs. Heterogeneity may also be partly stochastic, or be influenced by the presence of CCP 'hot spots' in the plasma membrane (Taylor et al, 2011; Antonescu et al, 2011; Gaidarov et al, 1999; Ehrlich et al, 2004; Saffarian et al, 2009; Nunez et al, 2011). It is important to note that although events in this pathway are depicted as occuring sequentially in a defined order, in reality the assembly of a clathrin-coated vesicle may be highly variable and the temporal boundaries are likely less clearly defined. Moreover, not every CCP will have all of the proteins indicated in this pathway.