Targeted Disruption of an EH‐domain Protein Endocytic Complex, Pan1‐End3

Abstract

Pan1 is a multi-domain scaffold that enables dynamic interactions with both structural and regulatory components of the endocytic pathway. Pan1 is composed of Eps15 Homology (EH) domains which interact with adaptor proteins, a central region that is responsible for its oligomerization and C-terminal binding sites for Arp2/3, F-actin, and type-I myosin motors. In this study, we have characterized the binding sites between Pan1 and its constitutive binding partner End3, another EH domain containing endocytic protein. The C-terminal End3 Repeats of End3 associate with the N-terminal part of Pan1’s central coiled-coil region. These repeats appear to act independently of one another as tandem, redundant binding sites for Pan1. The end3-1 allele was sequenced, and corresponds to a C-terminal truncation lacking the End3 Repeats. Mutations of the End3 Repeats highlight that those residues which are identical between these repeats serve as contact sites for the interaction with Pan1.

A Pan1/End3/Sla1 complex links Arp2/3-mediated actin assembly to sites of clathrin-mediated endocytosis

“…The End3 C-terminus was previously shown to bind directly to the middle of Pan1 in vitro ( Tang et al. , 1997 ; Whitworth et al. , 2014 ; Figure 1C ).…”

The Role of Secretory Pathways in Candida albicans Pathogenesis

“…End3 is an EF-hand Ca 2+ -binding protein that is required for the internalization of the endocytic vesicle and establishes proper actin patch and cell wall chitin localization [281]. The End3 C-terminus is the site of recruitment and binding by Pan1p to form a stable complex, enabling Pan1p phosphoregulation [282] Sc Pan1 is the only endocytic protein that is essential for viability, and contributes to actin cytoskeletal organization [283].…”

Actin and Endocytosis in Budding Yeast

“…Sla1 forms a complex with two EH domain-containing late coat proteins, Pan1 and End3 ( Tang et al 1997 ; Tang et al 2000 ). Recently, the specific domains in Pan1 and End3 mediating their interaction were identified, and it was shown that this interaction is required for Pan1 phosphoregulation ( Whitworth et al 2014 ). This Sla1 – Pan1 – End3 trio may be the yeast functional counterpart to mammalian endocytic scaffold/signaling protein Intersectin, which has EH and SH3 domains in a single polypeptide ( Yamabhai et al 1998 ).…”

“…Pan1 and Ent1 also have predicted Glc7 binding sites ( Gardiner et al 2007 ). Recent work shows that Pan1 dephosphorylation and disappearance from patches depends on Pan1 interactions with End3 and recruitment of Scd5 ( Whitworth et al 2014 ). Thus, there is a critical role for the Scd5 – Glc7 complex in regulating the dynamics and behavior of the endocytic machinery at cortical patches ( Chi et al 2012 ).…”