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== '''Introduction''' == | == '''Introduction''' == | ||
<StructureSection load='2NPS' size='500' side='right' caption='Structure of HMG-CoA reductase (PDB entry [[1dq8]])' scene=''> | |||
Important biological processes, such as synaptic transmission and cellular trafficking in Eukaryotes require '''SNARE proteins''' that are thought to play a crucial role in '''membrane fusion'''.<ref> PMID: 11340056 </ref><ref> PMID: 11252968 </ref><ref> PMID: 12600315 </ref><ref> PMID: 12154365 </ref> To connect membranes and allow their fusion, SNARE (soluble N-ethylmaleimide-sensitive-factor attachment protein receptor) proteins assemble into a '''core-complex of four parallel helices'''.<ref name="anto"> PMID: 11786915 </ref><ref> PMID: 9731768 </ref><ref name="sutton"> PMID: 9759724 </ref> The SNARE complex assembly is mediated by a '''conserved SNARE motif''' consisting of 60-70 amino acids. <ref> PMID: 9096343 </ref> SNAREs can be divided into two categories: the '''v-(vesicle) SNAREs''', which are found in the vesicle membrane and the '''t-(target) SNAREs''', which are anchored in the target membrane.<ref> PMID: 8455717 </ref> | Important biological processes, such as synaptic transmission and cellular trafficking in Eukaryotes require '''SNARE proteins''' that are thought to play a crucial role in '''membrane fusion'''.<ref> PMID: 11340056 </ref><ref> PMID: 11252968 </ref><ref> PMID: 12600315 </ref><ref> PMID: 12154365 </ref> To connect membranes and allow their fusion, SNARE (soluble N-ethylmaleimide-sensitive-factor attachment protein receptor) proteins assemble into a '''core-complex of four parallel helices'''.<ref name="anto"> PMID: 11786915 </ref><ref> PMID: 9731768 </ref><ref name="sutton"> PMID: 9759724 </ref> The SNARE complex assembly is mediated by a '''conserved SNARE motif''' consisting of 60-70 amino acids. <ref> PMID: 9096343 </ref> SNAREs can be divided into two categories: the '''v-(vesicle) SNAREs''', which are found in the vesicle membrane and the '''t-(target) SNAREs''', which are anchored in the target membrane.<ref> PMID: 8455717 </ref> | ||
The best-studied SNAREs are the neuronal and the '''early endosomal SNARE complexes'''. The Neuronal SNARE complex mediates exocytosis of synaptic vesicles in the neurons and includes the vesicle protein synaptobrevin (also called VAMP), the membrane proteins SNAP-25 and syntaxin 1.<ref name="sutton" /> '''The early endosomal SNARE complex''' includes '''syntaxin 6, syntaxin 13, vti1a''' and '''VAMP4''' and is responsible for homotypic fusion of early endosomes.<ref> PMID: 16469845 </ref> It has been shown that the crystal structure of the early endosomal SNARE complex resembles that of the neuronal and late endosomal complexes, but differs in surface side-chain interactions. | The best-studied SNAREs are the neuronal and the '''early endosomal SNARE complexes'''. The Neuronal SNARE complex mediates exocytosis of synaptic vesicles in the neurons and includes the vesicle protein synaptobrevin (also called VAMP), the membrane proteins SNAP-25 and syntaxin 1.<ref name="sutton" /> '''The early endosomal SNARE complex''' includes '''syntaxin 6, syntaxin 13, vti1a''' and '''VAMP4''' and is responsible for homotypic fusion of early endosomes.<ref> PMID: 16469845 </ref> It has been shown that the crystal structure of the early endosomal SNARE complex resembles that of the neuronal and late endosomal complexes, but differs in surface side-chain interactions. | ||
== '''Membrane fusion mechanism''' == | == '''Membrane fusion mechanism''' == | ||
[[Image:Membrane_fusion.jpg|thumbnail||800 px||center| '''Membrane fusion mechanism | [[Image:Membrane_fusion.jpg|thumbnail||800 px||center| '''Membrane fusion mechanism''']] | ||
Membrane Fusion requires the '''assembly of the core complex'''. '''Free t-SNAREs''' that are organized in '''clusters''' first assemble into acceptor complexes thanks to '''SM''' (Sec1/Munc18-related) '''proteins'''. Acceptor complexes can then interact with the '''v-SNAREs''' through the N-terminal domain of the SNARE motif. This enables the formation of '''four-helical trans-complexes''', in which only the N-terminal portions of the SNARE motifs are bound. This binding evolves from a '''loose''' to a '''tight state''', thus leading to the formation of a '''fusion pore'''. During the fusion, the conformation relaxes to a '''cis-configuration'''. Cis-complexes dissociate thanks to '''proteins''' and '''cofactors (SNAPs)'''. T- and v-SNAREs can be separated and recycled.<ref> PMID: 16912714 </ref> | Membrane Fusion requires the '''assembly of the core complex'''. '''Free t-SNAREs''' that are organized in '''clusters''' first assemble into acceptor complexes thanks to '''SM''' (Sec1/Munc18-related) '''proteins'''. Acceptor complexes can then interact with the '''v-SNAREs''' through the N-terminal domain of the SNARE motif. This enables the formation of '''four-helical trans-complexes''', in which only the N-terminal portions of the SNARE motifs are bound. This binding evolves from a '''loose''' to a '''tight state''', thus leading to the formation of a '''fusion pore'''. During the fusion, the conformation relaxes to a '''cis-configuration'''. Cis-complexes dissociate thanks to '''proteins''' and '''cofactors (SNAPs)'''. T- and v-SNAREs can be separated and recycled.<ref> PMID: 16912714 </ref> | ||
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=== '' “0”- | === '' “0”-layer'' === | ||
[[Image:Layers.png|thumbnail||1100 px||left| '''Topology of the 16 layers of the SNARE complex | [[Image:Layers.png|thumbnail||1100 px||left| '''Topology of the 16 layers of the SNARE complex''']] | ||
The centre of the four-helix bundle is constituted of '''16 layers'''. These layers are composed of '' | The centre of the four-helix bundle is constituted of '''16 layers'''. These layers are composed of <scene name='56/568021/Hydrophobis_side_chains/2'>hydrophobic side chains</scene>, which are perpendicular to the axis of the four-helix bundle, except for the central '''“0”-layer'''. This last one consists of '''three glutamine (Q)''' and '''one arginine (R)''' highly conserved residues. Those highly conserved residues have led to a new classification of SNAREs into '''Q- and R-SNAREs'''. <ref> PMID: 9861047 </ref> Almost all membrane fusion reactions require one R-SNARE and three Q-SNAREs: Qa, Qb and Qc.<ref> PMID: 11237004 </ref><ref> PMID: 11001046 </ref> In many cases, the R-SNARE is in the vesicle, and the three Q-SNAREs are in the target membrane. For the early endosomal SNARE complex, syntaxin 13, vti1a, syntaxin 6, and VAMP4 were respectively classified as Qa-, Qb-, Qc- and R-SNAREs. | ||
=== ''Structure of the early endosomal SNARE complex 2NPS'' === | === ''Structure of the early endosomal SNARE complex 2NPS'' === | ||
[[Image:Snare_complex.jpg|thumbnail||500 px||right| '''Crystal structure of the early endosomal SNARE complex.''' | |||
[[Image:Snare_complex.jpg|thumbnail|| | |||
(A) Backbone overlay of the early endosomal SNARE complex (in color) and the neuronal SNARE complex (gray). | (A) Backbone overlay of the early endosomal SNARE complex (in color) and the neuronal SNARE complex (gray). | ||
(B) N- to C-terminal view of the 0 layer containing the unusual aspartate in vti1a. (Color, early endosomal SNARE complex; gray, neuronal SNARE complex.) | (B) N- to C-terminal view of the 0 layer containing the unusual aspartate in vti1a. (Color, early endosomal SNARE complex; gray, neuronal SNARE complex.) | ||
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(D) View of the 0 layer, including hydrogen bonds with surface residues and water molecules. | (D) View of the 0 layer, including hydrogen bonds with surface residues and water molecules. | ||
]] | ]] | ||
The early endosomal SNARE complex forms a four-helix bundle with a '''left-handed superhelical twist'''. The position of the Qa-, Qb-, Qc-, and R-SNAREs is the same in the neuronal and late endosomal complexes.<ref name="sutton" /><ref name="anto" /> As mentioned above, an unconventional layer of hydrophilic residues – the “0”-layer – constituted of three glutamines and one arginine, characterizes the center of all SNARE complexes. However, in vti1a an '''aspartate residue''' substitutes the glutamine. The aspartate occupies the '''same position''' as the glutamine in the neuronal and late endosomal complexes, and is involved in a '''similar organization of hydrogen bonds''' with the other layer of amino acids. Moreover, in vti1a, <scene name='56/568021/Asp_156/5'>D156</scene> of the ‘0' layer also interacts with <scene name='56/568021/Asn_152/3'>N152</scene> at the surface of the vti1a helix and with a '''water molecule'''. Also, <scene name='56/568021/Gln_197/4'>sx6 Q197</scene> interacts with the <scene name='56/568021/D156/2'>backbone of vti1a D156</scene> and with <scene name='56/568021/Sx6_e193/2'>sx6 E193</scene> that in turn interacts with <scene name='56/568021/Vti1a_r157/3'>vti1a R157</scene>. Concerning <scene name='56/568021/Vti1a_r157/3'>vti1a R157</scene>, it interacts with <scene name='56/568021/Sx6_e196/2'>sx6 E196</scene>. This network of interactions resembles that observed in the neuronal complex.<ref> PMID: 12496247 </ref> The structure of the other layers is '''generally similar''' to that of the neuronal and late endosomal complexes. Indeed, slightly differences can be noted: in layer 6, the Qc-SNARE syntaxin 6 contains a valine, whereas the late endosomal Qc-SNARE syntaxin 8 contains a glutamate, which is involved in interactions with surface residues. Such interactions are absent in the early endosomal and the neuronal complexes. | |||
Regarding the SNARE motifs, they are not only connected by the central interacting layers, but also '''by surface interactions''' that often involve side chains with complementary charges. As an example, an interaction between the helices of '''syntaxin 6''' and '''vti1a''' involves a <scene name='56/568021/Aspser/2'>hydrogen bond between E143 (vti1a) and S179 (sx6)</scene>. At a similar position in the late endosomal complex, a '''salt bridge''' is observed between <scene name='56/568021/D157r164/2'>D157 (vti1b) and R164 (sx8)</scene>.<ref name="anto" /> It has to be noted that all these side chains are '''highly conserved''' between the respective SNAREs of ''Drosophila'' and mammals. Many other surface interactions connect the SNARE motifs. However, the role of these surface interactions in the stability of the whole complex is still unknown. | |||
== '''Regulation''' == | == '''Regulation''' == | ||
In addition to SNARE motifs and membrane anchors, SNARE proteins have '''autonomously folding N-terminal domains'''. These autonomously folded domains are able to '''regulate SNARE assembly'''. For example, synthaxin 7 has a '''three-helix-bundle domain''', which with the profiling-like domain Ykt6p of yeast can lightly '''inhibit the SNARE assembly'''.<ref> PMID: 11474112 </ref> Sso1p is the yeast ortholog of syntax in. Ssop has also '''three-helix-bundle domain''' that can bind to the SNARE motif. This binding generates a closed conformation that strongly '''inhibits the entry of Sso1p''' into SNARE complexes.<ref> PMID: 10048921 </ref><ref> PMID: 11017200 </ref><ref> PMID: 11777922 </ref><ref> PMID: 9731774 </ref> The main role of these autonomously folded domains is still under research. However, one of the known functions is the '''recruitment''' of other trafficking proteins thanks to the open/closed equilibrium of some SNAREs. | In addition to SNARE motifs and membrane anchors, SNARE proteins have '''autonomously folding N-terminal domains'''. These autonomously folded domains are able to '''regulate SNARE assembly'''. For example, synthaxin 7 has a '''three-helix-bundle domain''', which with the profiling-like domain Ykt6p of yeast can lightly '''inhibit the SNARE assembly'''.<ref> PMID: 11474112 </ref> Sso1p is the yeast ortholog of syntax in. Ssop has also a '''three-helix-bundle domain''' that can bind to the SNARE motif. This binding generates a closed conformation that strongly '''inhibits the entry of Sso1p''' into SNARE complexes.<ref> PMID: 10048921 </ref><ref> PMID: 11017200 </ref><ref> PMID: 11777922 </ref><ref> PMID: 9731774 </ref> The main role of these autonomously folded domains is still under research. However, one of the known functions is the '''recruitment''' of other trafficking proteins thanks to the open/closed equilibrium of some SNAREs. | ||
== ''' SNAREs and pulmonary hypertension ''' == | == ''' SNAREs and pulmonary hypertension ''' == | ||
'''Pulmonary hypertension''' (PH) corresponds to an '''increase of blood pressure''' in the pulmonary artery, pulmonary vein, or pulmonary capillaries leading to shortness of breath, dizziness, fainting, leg swelling and other symptoms. Pulmonary hypertension can be a '''severe disease''', characterized by a decreased exercise tolerance and heart failure. The origin of the disease seems to be a | '''Pulmonary hypertension''' (PH) corresponds to an '''increase of blood pressure''' in the pulmonary artery, pulmonary vein, or pulmonary capillaries leading to shortness of breath, dizziness, fainting, leg swelling and other symptoms. Pulmonary hypertension can be a '''severe disease''', characterized by a decreased exercise tolerance and heart failure. The origin of the disease seems to be a Golgi-dysfunction. Indeed, dysfunction of Golgi tethers, SNAPs and '''SNAREs''' apparently leads to pulmonary hypertension.<ref> PMID:17416597 </ref> | ||
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Florence HERMAL and Camille ROESCH | Florence HERMAL and Camille ROESCH | ||
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