Phosphorylation of Cysteine String Protein by Protein Kinase A

Abstract

Cyclic AMP-dependent protein kinase (PKA) enhances regulated exocytosis in neurons and most other secretory cells. To explore the molecular basis of this effect, known exocytotic proteins were screened for PKA substrates. Both cysteine string protein (CSP) and soluble NSF attachment protein-α (α-SNAP) were phosphorylated by PKA in vitro, but immunoprecipitation of cellular α-SNAP failed to detect 32P incorporation. In contrast, endogenous CSP was phosphorylated in synaptosomes, PC12 cells, and chromaffin cells. In-gel kinase assays confirmed PKA to be a cellular CSP kinase, with phosphorylation occurring on Ser10. PKA phosphorylation of CSP reduced its binding to syntaxin by 10-fold but had little effect on its interaction with HSC70 or G-protein subunits. Furthermore, an in vivo role for Ser10 phosphorylation at a late stage of exocytosis is suggested by analysis of chromaffin cells transfected with wild type or non-phosphorylatable mutant CSP. We propose that PKA phosphorylation of CSP could modulate the exocytotic machinery, by selectively altering its availability for protein-protein interactions. Cyclic AMP-dependent protein kinase (PKA) enhances regulated exocytosis in neurons and most other secretory cells. To explore the molecular basis of this effect, known exocytotic proteins were screened for PKA substrates. Both cysteine string protein (CSP) and soluble NSF attachment protein-α (α-SNAP) were phosphorylated by PKA in vitro, but immunoprecipitation of cellular α-SNAP failed to detect 32P incorporation. In contrast, endogenous CSP was phosphorylated in synaptosomes, PC12 cells, and chromaffin cells. In-gel kinase assays confirmed PKA to be a cellular CSP kinase, with phosphorylation occurring on Ser10. PKA phosphorylation of CSP reduced its binding to syntaxin by 10-fold but had little effect on its interaction with HSC70 or G-protein subunits. Furthermore, an in vivo role for Ser10 phosphorylation at a late stage of exocytosis is suggested by analysis of chromaffin cells transfected with wild type or non-phosphorylatable mutant CSP. We propose that PKA phosphorylation of CSP could modulate the exocytotic machinery, by selectively altering its availability for protein-protein interactions. soluble NSF attachment protein cysteine string protein cAMP-dependent protein kinase protein kinase C synaptosome-associated protein of 25 kDa glutathioneS-transferase vesicle-associated membrane protein reversed phase high performance liquid chromatography 4-morpholineethanesulfonic acid Exocytosis is the final stage of the secretory pathway and involves the fusion of secretory vesicles with the plasma membrane in a constitutive or regulated manner (1Morgan A. Essays Biochem. 1995; 30: 77-95PubMed Google Scholar). In regulated exocytosis, vesicles accumulate in the cytoplasm and only fuse with the plasma membrane upon receipt of an appropriate stimulus (usually, but not always, an increase in intracellular free Ca2+). As regulated exocytosis is the basis of chemical transmission in the brain, much research has been devoted to uncovering its molecular mechanism. This has revealed the involvement of a large number of proteins (2Jahn R. Sudhof T.C. Annu. Rev. Biochem. 1999; 68: 863-911Crossref PubMed Scopus (1025) Google Scholar, 3Lin R.C. Scheller R.H. Annu. Rev. Dev. Biol. 2000; 16: 19-49Crossref PubMed Scopus (422) Google Scholar), which can be classified into three groups. The first group, proteins involved in vesicle fusion events in all eukaryotes, includes the SNAP1 receptors, SNAPs, RABs, and the Sec1 family. The second group comprises proteins involved in regulated exocytosis in various cell types and diverse organisms but absent in yeast. This group includes the synaptotagmins and cysteine string proteins (CSP). The third class can be defined as proteins whose role in regulated exocytosis is cell type-specific. An example from this group is the synapsins, which are important modulators of the synaptic vesicle cycle in neurons (4Greengard P. Valtorta F. Czernik A.J. Benfenati F. Science. 1993; 259: 780-795Crossref PubMed Scopus (1130) Google Scholar). The complex interactions between the numerous proteins of these classes presumably enables sophisticated fine-tuning of exocytosis to suit the particular physiological needs of each cell type. In addition to the cell type-specific repertoire of exocytotic proteins expressed, further control over the exocytotic mechanism can be exerted post-translationally (5Turner K.M. Burgoyne R.D. Morgan A. Trends Neurosci. 1999; 22: 459-464Abstract Full Text Full Text PDF PubMed Scopus (209) Google Scholar). Indeed, a large number of studies have implicated protein kinases in the modulation of regulated exocytosis from many cell types by using cell-permeable inhibitors or activators, including Ca2+/calmodulin-dependent protein kinase II (6Nichols R.A. Sihra T.S. Czernik A.J. Nairn A.C. Greengard P. Nature. 1990; 343: 647-651Crossref PubMed Scopus (226) Google Scholar,7Schweitzer E.S. Sanderson M.J. Wasterlain C.G. J. Cell Sci. 1995; 108: 2619-2628PubMed Google Scholar), mitogen-activated protein kinase (8Cox M.E. Ely C.M. Catling A.D. Weber M.J. Parsons S.J. J. Neurochem. 1996; 66: 1103-1112Crossref PubMed Scopus (41) Google Scholar), cGMP-dependent protein kinase (9Yawo H. J. Neurosci. 1999; 19: 5293-5300Crossref PubMed Google Scholar), and tyrosine kinases (8Cox M.E. Ely C.M. Catling A.D. Weber M.J. Parsons S.J. J. Neurochem. 1996; 66: 1103-1112Crossref PubMed Scopus (41) Google Scholar). However, one shortfall of this approach is that the modulation of exocytosis may be indirect, either by effects on membrane receptor or ion channel phosphorylation or via direct steric inhibition of ion channels (e.g. Ref.10Li G. Hidaka H. Google Scholar). of kinase or inhibitors kinases to cells, and ion channels are is a of a role for protein kinases in the direct of the exocytotic However, a of the only PKA and or an of exocytosis in all secretory for example P. H. J. 1999; PubMed Scopus Google Scholar), chromaffin cells J. Biol. PubMed Scopus Google Scholar, A. Burgoyne R.D. Biochem. J. PubMed Scopus Google Scholar, A. Burgoyne R.D. J. 1993; PubMed Scopus Google Scholar), PC12 cells G. PubMed Scopus Google Scholar), cells J. 1996; 16: Scopus Google Scholar), cells A.J. Biochem. J. PubMed Scopus Google Scholar), cells J. Biol. Full Text PDF PubMed Google Scholar), cells J. PubMed Google Scholar), 1990; PubMed Scopus Google Scholar), and cells R.C. Biochem. J. 1990; PubMed Scopus Google Scholar). of PKA or exocytotic for the direct of exocytosis by To this approach was to known exocytotic proteins for in kinase substrates. that this only be the in the to this and to of in vivo In the have the synaptic vesicle protein CSP as a PKA in and in three cell and the phosphorylation to Ser10 in the of CSP. We that Ser10 phosphorylation can CSP binding to syntaxin but not to HSC70 or G-protein subunits. Furthermore, of Ser10 to a non-phosphorylatable the known effects of CSP on the of exocytotic PKA may exocytosis by the protein-protein interactions of CSP. CSP was as Burgoyne R.D. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). was from and were from was from was from each with an cysteine were from The of these were and is and protein were from was from PKA and all other were from and of and protein were as A. Burgoyne R.D. J. 1995; PubMed Scopus Google Scholar). and were and as H. Scheller R.H. Sci. A. 1996; PubMed Scopus Google Scholar, R. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). and were from of and of was as Burgoyne R.D. J. Biol. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). was a from of of H. Burgoyne R.D. J. J. Cell Sci. 1999; PubMed Google was using the the the are as and in that were to the acid The an which was to mutant was in the to of the only of of chromaffin cells were from each was three with of and and with of type at for The were from the by with of type and at for The cell was to and in by at The cells were a of and the in and The cell was and the chromaffin cells were in and either at 32P or in to by of CSP and as Burgoyne R. J. Neurosci. 2000; PubMed Google Scholar). were in at in a of and for to PC12 cell type and were in as Burgoyne R.D. Biol. PubMed Scopus Google Scholar). cells were in and cells were with CSP and using to the on a were as A. A. Full Text Full Text PDF PubMed Scopus Google Scholar). The protein of the was by Biochem. PubMed Scopus Google Scholar). the of cellular protein chromaffin cells PC12 cells or of were in for or or were for with various or kinase or were and CSP or α-SNAP was using J. Neurosci. 1999; PubMed Scopus Google Scholar). The were by or to by of the to a and for CSP or were with a of and was using phosphorylation were at in a of of and with the addition of of and to a final of of the and of phosphorylation of CSP or protein was with PKA or for to the of phosphorylated and CSP for the phosphorylation syntaxin and HSC70 of CSP was in the or of PKA for of proteins was by the with the on and liquid the CSP In the of phosphorylation for the and were or were at for with PKA or these the of was with and were by by in acid and of was by liquid were by of is and is of the of the in PKA phosphorylation of phosphorylated and CSP were as CSP or was reduced and in the of of the to a of of was and was to for at The was a at a of The were by with a of in acid over a of was at an of The the phosphorylated was by of the and by of the This was in was to a on a and the acid acid of the by The was to membrane using an of the The membrane was in the and to as but each cycle the acid by was to a using and by the of the CSP phosphorylation in PC12 cells, PC12 cells Burgoyne R.D. Biol. PubMed Scopus Google were and the CSP was as The CSP from cells was on a of CSP that had been phosphorylated in by The was and to a to CSP. The PC12 CSP and CSP were and to by The were as by and the were for The kinase was as H. Biochem. PubMed Scopus Google with R.A. Hidaka H. J. Biol. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). cellular proteins in from brain, PC12 cells, synaptosomes, and chromaffin cells were on with and CSP protein to the The were in with for in for the proteins were with for The proteins were by at in with were in kinase for to the phosphorylation were for at in kinase with and The was by the for in The were to a for of The binding of and CSP to was as M.E. Burgoyne R.D. J. Cell Sci. PubMed Google Scholar). the has a and a of CSP or were in the binding a was for CSP of was by to a and The of HSC70 by CSP was by a as Burgoyne R.D. Biochem. J. PubMed Scopus Google Scholar). cells with CSP wild type or mutant and were with and with free were and as Burgoyne R. J. Neurosci. 2000; PubMed Google Scholar). The phosphorylation by PKA and of proteins known to have a direct or role in the late of exocytosis was that were not phosphorylated by either kinase were syntaxin and The phosphorylation of and confirmed J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar, Greengard P. Czernik A.J. J. Neurochem. 1999; PubMed Scopus Google Scholar, A. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). However, and α-SNAP were substrates. this an of that are for the of exocytosis, of which have been J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar, Greengard P. Czernik A.J. J. Neurochem. 1999; PubMed Scopus Google Scholar, A. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar, R. Scheller R.H. J. Neurosci. 2000; PubMed Google Scholar, J. Neurochem. 1999; PubMed Scopus Google Scholar). In to for PKA phosphorylation with H. Scheller R.H. Sci. A. 1996; PubMed Scopus Google that exocytotic proteins are PKA substrates. We that α-SNAP is phosphorylated by in with H. Scheller R.H. Sci. A. 1996; PubMed Scopus Google Scholar, and that CSP is a PKA is a of a role for PKA in the modulation of exocytosis, the of exocytotic PKA is We the of CSP and α-SNAP phosphorylation by that CSP and α-SNAP are in for for was that phosphorylation of CSP by PKA at at a of of of protein α-SNAP by was phosphorylated to a by PKA with a of only of of protein the of in phosphorylation of a protein is not a of in vivo phosphorylation on to CSP and α-SNAP phosphorylation in To endogenous CSP and α-SNAP are phosphorylated in for regulated exocytosis, chromaffin cells, and The cells or were with and to with a or or the cell-permeable PKA and CSP and α-SNAP were from the with to and to 32P was by of the protein on the membrane was confirmed by with the analysis of CSP that was phosphorylated in in chromaffin cells, phosphorylation of CSP was by or α-SNAP was not phosphorylated in either chromaffin cells or The direct phosphorylation of CSP by PKA in with the of CSP phosphorylation by in vivo that PKA is a cellular CSP We an kinase to that PKA from cell and could CSP. from synaptosomes, PC12 cells, and chromaffin cells and PKA were on an with or CSP in the and of the the were in a kinase and to a kinase was in the control In the the of PKA an of kDa of this to the on the of the of kDa was in all of the of the molecular as the of PKA of the PKA A. J. Biol. 1990; Full Text PDF PubMed Google to the kinase for a phosphorylation of the in all that the in was that CSP is a in vivo for to the phosphorylation by using of phosphorylated in by PKA with and in the of was and the proteins were with and the by and was that the for CSP an by in with the for the protein of were to and only the in the of the from the were by and was that the phosphorylated had the as an by in and in the and phosphorylated and to with the this and the phosphorylated by PKA could not be to the of in the was to a membrane and to The acid were for and all of the in was in To further that PKA only not either of the or in PKA phosphorylation of a to and with either an or at the an PKA J. Biol. Full Text PDF PubMed Google Scholar), was in for for the was phosphorylated with with that of However, the and and were not phosphorylated at of to The phosphorylation of CSP by PKA in is to Ser10 are not phosphorylated by PKA in phosphorylated at phosphorylated at each with an cysteine were phosphorylated with PKA in the of of for at were by of and of a known for was in a each with an cysteine were phosphorylated with PKA in the of of for at were by of and of a known for was To endogenous CSP is phosphorylated on Ser10 in to a large of CSP from cells for analysis by and CSP a PC12 cell that Burgoyne R.D. Biol. PubMed Scopus Google Scholar). As in CSP is in PC12 cells Burgoyne R.D. Biol. PubMed Scopus Google with wild type cells, as Burgoyne R.D. Biol. PubMed Scopus Google Scholar). We first confirmed that CSP in PC12 cells was to phosphorylation as for chromaffin cells and cells were with a or and and CSP was CSP phosphorylation is control and not by or kinase to that in the constitutive CSP phosphorylation in control cells was high and the kinase suggested PKA is the only PC12 CSP kinase to control PC12 cells for the in vivo of CSP. PC12 cells were for with of and of the CSP from the PC12 and of that had been phosphorylated in by PKA were by The were and for The in vivo CSP only a of that to the in the phosphorylated CSP in PC12 cells CSP is phosphorylated in the as that in that PKA is a CSP kinase in the cell and the of PKA for Ser10 phosphorylation is that Ser10 is the CSP phosphorylation in of the effects of protein phosphorylation is to the of interaction of a protein with its binding CSP has been to vivo with and the and of R. P. J. Neurosci. 1999; 19: PubMed Google Scholar, 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, Sudhof T.C. J. Cell Biol. 1999; PubMed Scopus Google Scholar, 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, P. 30: Full Text Full Text PDF PubMed Scopus Google Scholar). To effect phosphorylation of CSP by PKA have upon its its interactions with these We a with or and of syntaxin were from the all not CSP protein was by which the of a that of CSP protein the of of CSP to and this was from the binding to to the of CSP to syntaxin CSP in a manner with a of CSP with to M.E. Burgoyne R.D. J. Cell Sci. PubMed Google Scholar, R. P. J. Neurosci. 1999; 19: PubMed Google Scholar, 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). of CSP by PKA in a in the of CSP for syntaxin a effect of CSP phosphorylation upon regulated exocytosis modulation of protein to CSP in vivo is P. 30: Full Text Full Text PDF PubMed Scopus Google Scholar). The of HSC70 by CSP Burgoyne R.D. Biochem. J. PubMed Scopus Google Scholar, Scheller R.H. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google a for in the binding of CSP to or was at a of in an with and by HSC70 was by a Burgoyne R.D. Biochem. J. PubMed Scopus Google Scholar). An of was at the of CSP Burgoyne R.D. Biochem. J. PubMed Scopus Google Scholar, Scheller R.H. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). The phosphorylation of CSP by PKA had effect upon its to HSC70 is in the binding of CSP to G-protein and have been to the of proteins 2000; Full Text Full Text PDF PubMed Scopus Google Scholar), and the phosphorylation of these interactions was approach binding of G-protein to or was for these a of was that of and to CSP of phosphorylation not further the of the To the role of CSP phosphorylation in the Ser10 in for or was that the CSP non-phosphorylatable and as a the of the phosphorylated CSP. To this the effect of the on syntaxin which have as a interaction by wild type or mutant with the of syntaxin in cells. In or of syntaxin wild type a a in syntaxin Indeed, of syntaxin with wild type CSP and This vivo interaction between the proteins and that the Ser10 not in the mutant The syntaxin binding by to wild type may constitutive phosphorylation of wild type CSP in the cells. of syntaxin binding were with that this failed to the protein not We the non-phosphorylatable to the role of Ser10 phosphorylation in of CSP in chromaffin cells has effects on exocytosis as by Burgoyne R. J. Neurosci. 2000; PubMed Google as a inhibition of exocytosis as a in and a effect on the of the To Ser10 phosphorylation these effects of chromaffin cells were transfected with wild type or Both were with protein detect transfected and exocytosis from cells was by of and was by Burgoyne R. J. Neurosci. 2000; PubMed Google Scholar). The example in as of CSP reduced the number of We a in number for the of However, analysis of the defined in revealed between of the wild type CSP and the Ser10 mutant type CSP the of as an increase in the and increase in Burgoyne R. J. Neurosci. 2000; PubMed Google Scholar). In contrast, from cells and to control these that the inhibition of exocytosis to CSP is of Ser10 phosphorylation but that this is for the modulation of exocytosis by CSP. The mechanism by which protein kinases modulate the late of regulated exocytosis is the of exocytosis in most secretory by PKA or its is a but have upon proteins involved in exocytosis that are of PKA In this have that CSP is a PKA exocytotic PKA and α-SNAP is a in for PKA and H. Scheller R.H. Sci. A. 1996; PubMed Scopus Google have that α-SNAP is not phosphorylated in either chromaffin cells or immunoprecipitation of of protein from an of the is phosphorylated in by PKA Sudhof T.C. J. Neurosci. 1995; PubMed Google and in vivo in to or in the of the G. Sudhof T.C. J. Neurosci. PubMed Google Scholar). However, effects of phosphorylation example modulation of its interaction with have been and in the in or R.A. R. R.C. Sudhof T.C. J. Neurosci. 1999; 19: PubMed Google Scholar). In contrast, CSP in Science. PubMed Scopus Google Scholar, Full Text PDF PubMed Scopus Google Scholar), and not only is CSP phosphorylated in but have of its of this were P. J. Cell Biol. PubMed Scopus Google the synaptic vesicle protein to be a PKA in As have for is and phosphorylation its in in binding to and cellular assays of a mutant in chromaffin cells exocytosis The of PKA and on the synaptic vesicle may to the sophisticated of by However, as is Neurosci. 1999; PubMed Scopus Google Scholar), CSP a of PKA in exocytosis in other cell of the CSP acid by the and J. Biol. 1999; PubMed Scopus Google Scholar, A. 1999; PubMed Scopus Google phosphorylation for a of including Ca2+/calmodulin-dependent protein kinase II kinase and kinase II and kinase PKA and and However, are with PKA a CSP kinase and Ser10 as its of in vitro, CSP is phosphorylated by PKA only at an was not In can CSP phosphorylation in chromaffin cells, and phosphorylation of CSP PC12 cells in a Ser10. Furthermore, an kinase PKA to be the only kinase from cell that could CSP. on not Ser10 as a PKA This that the of the protein has a on kinase and the to protein phosphorylation An interaction between CSP and syntaxin in and in vivo is in the M.E. Burgoyne R.D. J. Cell Sci. PubMed Google R. P. J. Neurosci. 1999; 19: PubMed Google Scholar, 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). In CSP and syntaxin can be and the of mutant syntaxin can be by the of CSP R. P. J. Neurosci. 1999; 19: PubMed Google Scholar). We have that in with to that in studies R. P. J. Neurosci. 1999; 19: PubMed Google Scholar, 1999; Full Text Full Text PDF PubMed Scopus Google using the We have CSP and the of syntaxin from a a cellular interaction of the is known that the of CSP is for binding HSC70 Burgoyne R.D. Biochem. J. PubMed Scopus Google Scholar, Sudhof T.C. J. Cell Biol. 1999; PubMed Scopus Google Scholar, Scheller R.H. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar), the CSP that with syntaxin is Ser10 the CSP that is known to with HSC70 is not that have phosphorylation not its of HSC70 In effect of phosphorylation on the binding of CSP to or 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). of binding to syntaxin by phosphorylation of CSP on Ser10 the of CSP has a role in syntaxin This is with the that and CSP syntaxin the of each protein little Burgoyne R.D. J. Neurochem. 2000; PubMed Scopus Google Scholar), the the phosphorylation have high the of a CSP from to may an mechanism. We have that the effects of wild type CSP in chromaffin cells on Burgoyne R. J. Neurosci. 2000; PubMed Google Ser10. of wild type CSP in an increase in the and of the of However, of Ser10 to CSP in with control for and This effect is not to of the mutant protein this a in number to wild type CSP and CSP proteins were to upon in cells not the effects of wild type CSP on are to the only between the mutant and wild type CSP is that the mutant be phosphorylated at the As the is to the of from fusion to membrane fusion J. Burgoyne R.D. Science. PubMed Scopus Google Scholar), this a role for CSP phosphorylation at a late stage of in chromaffin cells has that of or other that increase cellular have the effects upon as of and A. R. Google Scholar). Furthermore, these effects were by with the a role for PKA in the of A. R. Google Scholar). PKA or of CSP the late of exocytosis and PKA inhibition or of non-phosphorylatable CSP these the modulation by PKA of the late of exocytosis by A. R. Google could be by its phosphorylation of CSP on Ser10. The may be effects of CSP on regulated exocytosis as a in the number of exocytotic and a of in the In the in number is to be to protein-protein interactions of CSP. HSC70 is an as its interaction with CSP is by Furthermore, HSC70 is for synaptic vesicle exocytosis in and interaction with CSP is for this P. 30: Full Text Full Text PDF PubMed Scopus Google Scholar). of G-protein and by CSP is the interaction of CSP with and has been in the of channel 2000; Full Text Full Text PDF PubMed Scopus Google Scholar), direct effects of on the exocytotic have been in various secretory cells, including neurons Science. PubMed Scopus (209) Google Scholar). However, be that the in exocytosis is by CSP binding to is a for this in of CSP can the effects of syntaxin upon R. P. J. Neurosci. 1999; 19: PubMed Google Scholar). the molecular basis of the inhibition of exocytosis by CSP is is not to chromaffin cells, as CSP in cells exocytosis H. Burgoyne R.D. J. J. Cell Sci. 1999; PubMed Google Scholar, H. R. G. G. Scheller R.H. J. PubMed Scopus Google Scholar). We propose that the second effect of CSP on exocytosis, the of is upon phosphorylation of CSP is not in cells non-phosphorylatable CSP. In the effects upon are in chromaffin cells of PKA A. R. Google Scholar). that the effect of CSP is to be to of syntaxin by phosphorylated CSP has an for Furthermore, analysis from cells syntaxin has been by effects upon number have with wild type and mutant CSP but to M.E. Burgoyne R.D. 2000; PubMed Scopus Google Scholar). could phosphorylated CSP the effects on is that phosphorylation of CSP syntaxin to with other binding that to the late of This with the physiological role of CSP as a syntaxin M.E. Burgoyne R.D. J. Cell Sci. PubMed Google Scholar, R. P. J. Neurosci. 1999; 19: PubMed Google Scholar, 1999; Full Text Full Text PDF PubMed Scopus Google Burgoyne R.D. J. Neurochem. 2000; PubMed Scopus Google Scholar). a of proteins could syntaxin CSP is the most by of the high of this interaction J. H. Scheller R.H. Full Text PDF PubMed Scopus Google Scholar). of an mutant with reduced for syntaxin the late of exocytosis, as a in and J. Burgoyne R.D. Science. PubMed Scopus Google Scholar), the of CSP In of the of endogenous wild type to the of membrane is to that the effect of CSP phosphorylation on exocytosis is to of An of the is that an involved in the late of exocytosis phosphorylated CSP studies have implicated CSP in the of channels modulation of a direct fusion of exocytosis Burgoyne R. J. Neurosci. 2000; PubMed Google R. P. J. Neurosci. 1999; 19: PubMed Google Scholar, 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, P. J. Neurosci. 2000; PubMed Google Scholar, Full Text PDF PubMed Scopus Google Scholar, R. P. J. Neurosci. PubMed Google Scholar, J. Neurosci. PubMed Google Scholar, H. Burgoyne R.D. J. PubMed Scopus Google Scholar). In this propose a of the physiological of CSP its phosphorylation by This could modulate exocytosis by the of syntaxin into protein involved in vesicle and fusion or by interactions with In phosphorylation of CSP could via reduced binding to which is known as a of ion channel Scheller R.H. Nature. 1995; PubMed Scopus Google Scholar, H. J. Neurosci. PubMed Google Scholar). CSP phosphorylation could in synaptic vesicle exocytosis, sophisticated control of Furthermore, as CSP has a and in exocytosis from a of cell types from cells to CSP phosphorylation by PKA could be a mechanism for the of We and for proteins and for the