Shared as well as distinct roles of EHD proteins revealed by biochemical and functional comparisons in mammalian cells and C. elegans

Manju George, Guo Guang Ying, Mark A. Rainey, Aharon Solomon, Pankit T. Parikh, Qingshen Gao, Vimla Band, Hamid Band

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Abstract

Background: The four highly homologous human EHD proteins (EHD1-4) form a distinct subfamily of the Eps15 homology domain-containing protein family and are thought to regulate endocytic recycling. Certain members of this family have been studied in different cellular contexts; however, a lack of concurrent analyses of all four proteins has impeded an appreciation of their redundant versus distinct functions. Results: Here, we analyzed the four EHD proteins both in mammalian cells and in a cross-species complementation assay using a C. elegans mutant lacking the EHD ortholog RME-1. We show that all human EHD proteins rescue the vacuolated intestinal phenotype of C. elegans rme-1 mutant, are simultaneously expressed in a panel of mammalian cell lines and tissues tested, and variably homo- and hetero-oligomerize and colocalize with each other and Rab11, a recycling endosome marker. Small interfering RNA (siRNA) knock-down of EHD1, 2 and 4, and expression of dominant-negative EH domain deletion mutants showed that loss of EHD1 and 3 (and to a lesser extent EHD4) but not EHD2 function retarded transferrin exit from the endocytic recycling compartment. EH domain deletion mutants of EHD1 and 3 but not 2 or 4, induced a striking perinuclear clustering of co-transfected Rab11. Knock-down analyses indicated that EHD1 and 2 regulate the exit of cargo from the recycling endosome while EHD4, similar to that reported for EHD3 (Naslavsky et al. (2006) Mol. Biol. Cell 17, 163), regulates transport from the early endosome to the recycling endosome. Conclusion: Altogether, our studies suggest that concurrently expressed human EHD proteins perform shared as well as discrete functions in the endocytic recycling pathway and lay a foundation for future studies to identify and characterize the molecular pathways involved.

Original languageEnglish (US)
Article number3
JournalBMC Cell Biology
Volume8
DOIs
StatePublished - Feb 15 2007

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Endosomes
Proteins
Transferrin
Small Interfering RNA
Cluster Analysis
Phenotype
Cell Line

ASJC Scopus subject areas

  • Cell Biology

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Shared as well as distinct roles of EHD proteins revealed by biochemical and functional comparisons in mammalian cells and C. elegans. / George, Manju; Ying, Guo Guang; Rainey, Mark A.; Solomon, Aharon; Parikh, Pankit T.; Gao, Qingshen; Band, Vimla; Band, Hamid.

In: BMC Cell Biology, Vol. 8, 3, 15.02.2007.

Research output: Contribution to journalArticle

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