Alternative splicing of SNAP-25 regulates secretion through nonconservative substitutions in the SNARE domain

TY - JOUR

T1 - Alternative splicing of SNAP-25 regulates secretion through nonconservative substitutions in the SNARE domain

AU - Nagy, Gábor

AU - Milosevic, Ira

AU - Fasshauer, Dirk

AU - Müller, E Matthias

AU - de Groot, Bert L

AU - Lang, Thorsten

AU - Wilson, Michael C

AU - Sørensen, Jakob B

N1 - Keywords: Alternative Splicing; Amino Acid Sequence; Amino Acid Substitution; Animals; Chromaffin Cells; Circular Dichroism; Computer Simulation; Exons; Mice; Mice, Knockout; Molecular Sequence Data; SNARE Proteins; Synaptosomal-Associated Protein 25

PY - 2005

Y1 - 2005

N2 - The essential membrane fusion apparatus in mammalian cells, the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex, consists of four alpha-helices formed by three proteins: SNAP-25, syntaxin 1, and synaptobrevin 2. SNAP-25 contributes two helices to the complex and is targeted to the plasma membrane by palmitoylation of four cysteines in the linker region. It is alternatively spliced into two forms, SNAP-25a and SNAP-25b, differing by nine amino acids substitutions. When expressed in chromaffin cells from SNAP-25 null mice, the isoforms support different levels of secretion. Here, we investigated the basis of that different secretory phenotype. We found that two nonconservative substitutions in the N-terminal SNARE domain and not the different localization of one palmitoylated cysteine cause the functional difference between the isoforms. Biochemical and molecular dynamic simulation experiments revealed that the two substitutions do not regulate secretion by affecting the property of SNARE complex itself, but rather make the SNAP-25b-containing SNARE complex more available for the interaction with accessory factor(s).

AB - The essential membrane fusion apparatus in mammalian cells, the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex, consists of four alpha-helices formed by three proteins: SNAP-25, syntaxin 1, and synaptobrevin 2. SNAP-25 contributes two helices to the complex and is targeted to the plasma membrane by palmitoylation of four cysteines in the linker region. It is alternatively spliced into two forms, SNAP-25a and SNAP-25b, differing by nine amino acids substitutions. When expressed in chromaffin cells from SNAP-25 null mice, the isoforms support different levels of secretion. Here, we investigated the basis of that different secretory phenotype. We found that two nonconservative substitutions in the N-terminal SNARE domain and not the different localization of one palmitoylated cysteine cause the functional difference between the isoforms. Biochemical and molecular dynamic simulation experiments revealed that the two substitutions do not regulate secretion by affecting the property of SNARE complex itself, but rather make the SNAP-25b-containing SNARE complex more available for the interaction with accessory factor(s).

U2 - 10.1091/mbc.E05-07-0595

DO - 10.1091/mbc.E05-07-0595

M3 - Journal article

C2 - 16195346

SN - 1059-1524

VL - 16

SP - 5675

EP - 5685

JO - Molecular Biology of the Cell

JF - Molecular Biology of the Cell

IS - 12

ER -