Light chain C-terminal region reinforces the stability of clathrin heavy chain trimers.
The self-assembly of clathrin into lattices relies on the ability of heavy chain legs to form a three-legged pinwheel structure. We investigated the role of light chains in clathrin trimerization by challenging recombinant hub (plus and minus light chain) with an anionic detergent. The binding of light chain increases the amount of detergent needed to induce detrimerization, suggesting light chains reinforced hub trimers. We also show that light chain C-terminal residues are important for enhancing the in vitro assembly of hub at low pH. We assessed how much the C-terminus of light chain contributed to the stability of the trimerization domain by adding full-length and truncated light chains to trimer-defective hub mutants, C1573S and C1573A. Adding full-length LCb to C1573S caused some retrimerization, but little activity was restored, suggesting the majority of oligomeric C1573S was nonnative. A larger percentage of monomeric C1573A could be retrimerized into an assembly-competent form by adding intact LCb. We also discovered that C-terminally deleted light chains produced a heterogeneous population of hubs that were smaller than native hubs, but were assembly active. We propose a model showing how light chains reinforce the puckered clathrin triskelion. Finally, the ability of light chains to retrimerize C1573A hub suggests that the structural role of light chain may be conserved in yeast and mammals.
Ybe
Joel A
JA
Department of Biology, Indiana University, Bloomington, IN 47405, USA. jybe@indiana.edu
Perez-Miller
Samantha
S
Niu
Qian
Q
Coates
David A
DA
Drazer
Michael W
MW
Clegg
Mary E
ME
D019943:Amino Acid Substitution; D000818:Animals; D002417:Cattle; D002966:Clathrin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D003902:Detergents; D020836:Protein Structure, Quaternary / Q000502:physiology*; D017434:Protein Structure, Tertiary / Q000502:physiology; D012521:Sarcosine / Q000031:analogs & derivatives
D002966:Clathrin; D003902:Detergents; C025231:sarkosyl; D012521:Sarcosine