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Hrs regulates early endosome fusion by inhibiting formation of an endosomal SNARE complex.

Sun W, Yan Q, Vida TA, Bean AJ - J. Cell Biol. (2003)

Bottom Line: We found that the endosome-associated hepatocyte responsive serum phosphoprotein (Hrs) inhibited the homotypic fusion of early endosomes.SNAP-25, syntaxin 13, and VAMP2 were bound from rat brain membranes to the Hrs coiled-coil domain.Hrs inhibited formation of the syntaxin 13-SNAP-25-VAMP2 complex by displacing VAMP2 from the complex.

View Article: PubMed Central - PubMed

Affiliation: The University of Texas Health Science Center, Dept. of Neurobiology and Anatomy, 6431 Fannin Street, MSB 7.208, Houston, TX 77030, USA.

ABSTRACT
Movement through the endocytic pathway occurs principally via a series of membrane fusion and fission reactions that allow sorting of molecules to be recycled from those to be degraded. Endosome fusion is dependent on SNARE proteins, although the nature of the proteins involved and their regulation has not been fully elucidated. We found that the endosome-associated hepatocyte responsive serum phosphoprotein (Hrs) inhibited the homotypic fusion of early endosomes. A region of Hrs predicted to form a coiled coil required for binding the Q-SNARE, SNAP-25, mimicked the inhibition of endosome fusion produced by full-length Hrs, and was sufficient for endosome binding. SNAP-25, syntaxin 13, and VAMP2 were bound from rat brain membranes to the Hrs coiled-coil domain. Syntaxin 13 inhibited early endosomal fusion and botulinum toxin/E inhibition of early endosomal fusion was reversed by addition of SNAP-25(150-206), confirming a role for syntaxin 13, and establishing a role for SNAP-25 in endosomal fusion. Hrs inhibited formation of the syntaxin 13-SNAP-25-VAMP2 complex by displacing VAMP2 from the complex. These data suggest that SNAP-25 is a receptor for Hrs on early endosomal membranes and that the binding of Hrs to SNAP-25 on endosomal membranes inhibits formation of a SNARE complex required for homotypic endosome fusion.

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BoNT/E inhibits early endosomal fusion in a SNAP-25(150–206)–dependent manner. (A) BoNT/E dose dependently and saturably inhibited early endosome fusion. (B) The inhibition of early endosome fusion by BoNT/E is reversed by addition of SNAP-25(150–206) supporting a role for SNAP-25 in endosome fusion.
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fig8: BoNT/E inhibits early endosomal fusion in a SNAP-25(150–206)–dependent manner. (A) BoNT/E dose dependently and saturably inhibited early endosome fusion. (B) The inhibition of early endosome fusion by BoNT/E is reversed by addition of SNAP-25(150–206) supporting a role for SNAP-25 in endosome fusion.

Mentions: Because we detected a SNAP-25–containing SNARE complex on endosomal membranes, we examined the effect of botulinum neurotoxin E (BoNT/E), a zinc endoprotease that cleaves the COOH-terminal 26 aa of SNAP-25. BoNT/E blocks membrane fusion that requires a four-helical–SNARE complex containing SNAP-25 (Banerjee et al., 1996; Chen et al., 1999; Jahn and Sudhof, 1999). We observed that BoNT/E inhibited early endosome fusion in a concentration-dependent manner with a half-maximal inhibition of ∼20 nM (Fig. 8Figure 8.


Hrs regulates early endosome fusion by inhibiting formation of an endosomal SNARE complex.

Sun W, Yan Q, Vida TA, Bean AJ - J. Cell Biol. (2003)

BoNT/E inhibits early endosomal fusion in a SNAP-25(150–206)–dependent manner. (A) BoNT/E dose dependently and saturably inhibited early endosome fusion. (B) The inhibition of early endosome fusion by BoNT/E is reversed by addition of SNAP-25(150–206) supporting a role for SNAP-25 in endosome fusion.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172712&req=5

fig8: BoNT/E inhibits early endosomal fusion in a SNAP-25(150–206)–dependent manner. (A) BoNT/E dose dependently and saturably inhibited early endosome fusion. (B) The inhibition of early endosome fusion by BoNT/E is reversed by addition of SNAP-25(150–206) supporting a role for SNAP-25 in endosome fusion.
Mentions: Because we detected a SNAP-25–containing SNARE complex on endosomal membranes, we examined the effect of botulinum neurotoxin E (BoNT/E), a zinc endoprotease that cleaves the COOH-terminal 26 aa of SNAP-25. BoNT/E blocks membrane fusion that requires a four-helical–SNARE complex containing SNAP-25 (Banerjee et al., 1996; Chen et al., 1999; Jahn and Sudhof, 1999). We observed that BoNT/E inhibited early endosome fusion in a concentration-dependent manner with a half-maximal inhibition of ∼20 nM (Fig. 8Figure 8.

Bottom Line: We found that the endosome-associated hepatocyte responsive serum phosphoprotein (Hrs) inhibited the homotypic fusion of early endosomes.SNAP-25, syntaxin 13, and VAMP2 were bound from rat brain membranes to the Hrs coiled-coil domain.Hrs inhibited formation of the syntaxin 13-SNAP-25-VAMP2 complex by displacing VAMP2 from the complex.

View Article: PubMed Central - PubMed

Affiliation: The University of Texas Health Science Center, Dept. of Neurobiology and Anatomy, 6431 Fannin Street, MSB 7.208, Houston, TX 77030, USA.

ABSTRACT
Movement through the endocytic pathway occurs principally via a series of membrane fusion and fission reactions that allow sorting of molecules to be recycled from those to be degraded. Endosome fusion is dependent on SNARE proteins, although the nature of the proteins involved and their regulation has not been fully elucidated. We found that the endosome-associated hepatocyte responsive serum phosphoprotein (Hrs) inhibited the homotypic fusion of early endosomes. A region of Hrs predicted to form a coiled coil required for binding the Q-SNARE, SNAP-25, mimicked the inhibition of endosome fusion produced by full-length Hrs, and was sufficient for endosome binding. SNAP-25, syntaxin 13, and VAMP2 were bound from rat brain membranes to the Hrs coiled-coil domain. Syntaxin 13 inhibited early endosomal fusion and botulinum toxin/E inhibition of early endosomal fusion was reversed by addition of SNAP-25(150-206), confirming a role for syntaxin 13, and establishing a role for SNAP-25 in endosomal fusion. Hrs inhibited formation of the syntaxin 13-SNAP-25-VAMP2 complex by displacing VAMP2 from the complex. These data suggest that SNAP-25 is a receptor for Hrs on early endosomal membranes and that the binding of Hrs to SNAP-25 on endosomal membranes inhibits formation of a SNARE complex required for homotypic endosome fusion.

Show MeSH