EHD3 regulates early-endosome-to-Golgi transport and preserves Golgi morphology.
Depletion of EHD3 affects sorting in endosomes by altering the kinetics and route of receptor recycling to the plasma membrane. Here we demonstrate that siRNA knockdown of EHD3, or its interaction partner rabenosyn-5, causes redistribution of sorting nexin 1 (SNX1) to enlarged early endosomes and disrupts transport of internalized Shiga toxin B subunit (STxB) to the Golgi. Moreover, under these conditions, Golgi morphology appears as a series of highly dispersed and fragmented stacks that maintain characteristics of cis-, medial- and trans-Golgi membranes. Although Arf1 still assembled onto these dispersed Golgi membranes, the level of AP-1 gamma-adaptin recruited to the Golgi was diminished. Whereas VSV-G-secretion from the dispersed Golgi remained largely unaffected, the distribution of mannose 6-phosphate receptor (M6PR) was altered: it remained in peripheral endosomes and did not return to the Golgi. Cathepsin D, a hydrolase that is normally transported to lysosomes via an M6PR-dependent pathway, remained trapped at the Golgi. Our findings support a role for EHD3 in regulating endosome-to-Golgi transport, and as a consequence, lysosomal biosynthetic, but not secretory, transport pathways are also affected. These data also suggest that impaired endosome-to-Golgi transport and the resulting lack of recruitment of AP-1 gamma-adaptin to Golgi membranes affect Golgi morphology.
Naslavsky
Naava
N
Department of Biochemistry and Molecular Biology and Eppley Cancer Center, University of Nebraska Medical Center, Omaha, NE 68198, USA.
McKenzie
Jenna
J
Altan-Bonnet
Nihal
N
Sheff
David
D
Caplan
Steve
S
D020823:ADP-Ribosylation Factor 1 / Q000378:metabolism; D033961:Adaptor Protein Complex 1 / Q000378:metabolism; D001692:Biological Transport / Q000502:physiology; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002402:Cathepsin D / Q000378:metabolism; D011992:Endosomes / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006056:Golgi Apparatus / Q000378:metabolism* / Q000648:ultrastructure; D006367:HeLa Cells; D006801:Humans; D050765:Qa-SNARE Proteins / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000378:metabolism; D017527:Receptor, IGF Type 2; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D022603:Shiga Toxins / Q000378:metabolism; D058305:Sorting Nexins; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism*
D033961:Adaptor Protein Complex 1; D002352:Carrier Proteins; C406447:EHD3 protein, human; D050765:Qa-SNARE Proteins; C416362:RBSN protein, human; D034741:RNA, Small Interfering; D017527:Receptor, IGF Type 2; D018160:Receptors, Cytoplasmic and Nuclear; C513978:SNX1 protein, human; C496151:STX12 protein, human; D022603:Shiga Toxins; D058305:Sorting Nexins; D033921:Vesicular Transport Proteins; C034304:cation-dependent mannose-6-phosphate receptor; C077239:stxB toxin; C533922:CTSD protein, human; D002402:Cathepsin D; D020823:ADP-Ribosylation Factor 1