GRIF1 binds Hrs and is a new regulator of endosomal trafficking.
Endosomal sorting of internalized cell surface receptors to the lysosomal pathway plays a crucial role in the control of cell signaling and function. Here we report the identification of GABA(A) receptor interacting factor-1 (GRIF1), a recently discovered protein of unknown function, as a new regulator of endosome-to-lysosome trafficking. Yeast two-hybrid screen and co-immunoprecipitation analysis reveal that GRIF1 interacts with hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs), an essential component of the endosomal sorting machinery. We have mapped the binding domains of GRIF1 and Hrs that mediate their association and shown the colocalization of GRIF1 with Hrs on early endosomes. Like Hrs, both overexpression and siRNA-mediated depletion of GRIF1 inhibit the degradation of internalized epidermal growth factor receptors and block the trafficking of the receptors from early endosomes to the lysosomal pathway. Our results indicate, for the first time, a functional role for GRIF1 in the regulation of endosomal trafficking. Interestingly, overexpression of full-length GRIF1, but not the Hrs- or kinesin-interacting GRIF1 deletion mutants, causes a perinuclear clustering of early endosomes. Our findings suggest that GRIF1 may also participate in microtubule-based transport of early endosomes by acting as an adaptor linking Hrs-containing endosomes to kinesin.
Kirk
Elizabeth
E
Department of Pharmacology, Emory University School of Medicine, Atlanta, GA 30322, USA.
Chin
Lih-Shen
LS
Li
Lian
L
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D003001:Cloning, Molecular; D056827:Endosomal Sorting Complexes Required for Transport; D011992:Endosomes / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008247:Lysosomes / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008870:Microtubules / Q000378:metabolism; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D009928:Organ Specificity; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques; D033921:Vesicular Transport Proteins / Q000378:metabolism
D002352:Carrier Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010750:Phosphoproteins; C517505:Trak2 protein, rat; D033921:Vesicular Transport Proteins; C093846:early endosome antigen 1; D066246:ErbB Receptors; D000251:Adenosine Triphosphatases; C115505:Hgs protein, rat