The in Vivo Phosphorylation Sites of Rat Brain Dynamin I

Abstract

Dynamin I (dynI) is phosphorylated in synaptosomes at Ser774 and Ser778 by cyclin-dependent kinase 5 to regulate recruitment of syndapin I for synaptic vesicle endocytosis, and in PC12 cells on Ser857. Hierarchical phosphorylation of Ser774 precedes phosphorylation of Ser778. In contrast, Thr780 phosphorylation by cdk5 has been reported as the sole site (Tomizawa, K., Sunada, S., Lu, Y. F., Oda, Y., Kinuta, M., Ohshima, T., Saito, T., Wei, F. Y., Matsushita, M., Li, S. T., Tsutsui, K., Hisanaga, S. I., Mikoshiba, K., Takei, K., and Matsui, H. (2003) J. Cell Biol. 163, 813–824). To resolve the discrepancy and to better understand the biological roles of dynI phosphorylation, we undertook a systematic identification of all phosphorylation sites in rat brain nerve terminal dynI. Using phosphoamino acid analysis, exclusively phospho-serine residues were found. Thr780 phosphorylation was not detectable. Mutation of Ser774, Ser778, and Thr780 confirmed that Thr780 phosphorylation is restricted to in vitro conditions. Mass spectrometry of 32P-labeled phosphopeptides separated by two-dimensional mapping revealed seven in vivo phosphorylation sites: Ser774, Ser778, Ser822, Ser851, Ser857, Ser512, and Ser347. Quantification of 32P radiation in each phosphopeptide showed that Ser774 and Ser778 were the major sites (up to 69% of the total), followed by Ser851 and Ser857 (12%), and Ser853 (2%). Phosphorylation of Ser851 and Ser857 was restricted to the long tail splice variant dynIxa and was not hierarchical. Co-purified, 32P-labeled dynIII was phosphorylated at Ser759, Ser763, and Ser853. Ser853 is homologous to Ser851 in dynIxa. The results identify all major and several minor phosphorylation sites in dynI and provide the first measure of their relative abundance and relative responses to depolarization. The multiple phospho-sites suggest subtle regulation of synaptic vesicle endocytosis by new protein kinases and new protein-protein interactions. The homologous dynI and dynIII phosphorylation indicates a high mechanistic similarity. The results suggest a unique role for the long splice variants of dynI and dynIII in nerve terminals. Dynamin I (dynI) is phosphorylated in synaptosomes at Ser774 and Ser778 by cyclin-dependent kinase 5 to regulate recruitment of syndapin I for synaptic vesicle endocytosis, and in PC12 cells on Ser857. Hierarchical phosphorylation of Ser774 precedes phosphorylation of Ser778. In contrast, Thr780 phosphorylation by cdk5 has been reported as the sole site (Tomizawa, K., Sunada, S., Lu, Y. F., Oda, Y., Kinuta, M., Ohshima, T., Saito, T., Wei, F. Y., Matsushita, M., Li, S. T., Tsutsui, K., Hisanaga, S. I., Mikoshiba, K., Takei, K., and Matsui, H. (2003) J. Cell Biol. 163, 813–824). To resolve the discrepancy and to better understand the biological roles of dynI phosphorylation, we undertook a systematic identification of all phosphorylation sites in rat brain nerve terminal dynI. Using phosphoamino acid analysis, exclusively phospho-serine residues were found. Thr780 phosphorylation was not detectable. Mutation of Ser774, Ser778, and Thr780 confirmed that Thr780 phosphorylation is restricted to in vitro conditions. Mass spectrometry of 32P-labeled phosphopeptides separated by two-dimensional mapping revealed seven in vivo phosphorylation sites: Ser774, Ser778, Ser822, Ser851, Ser857, Ser512, and Ser347. Quantification of 32P radiation in each phosphopeptide showed that Ser774 and Ser778 were the major sites (up to 69% of the total), followed by Ser851 and Ser857 (12%), and Ser853 (2%). Phosphorylation of Ser851 and Ser857 was restricted to the long tail splice variant dynIxa and was not hierarchical. Co-purified, 32P-labeled dynIII was phosphorylated at Ser759, Ser763, and Ser853. Ser853 is homologous to Ser851 in dynIxa. The results identify all major and several minor phosphorylation sites in dynI and provide the first measure of their relative abundance and relative responses to depolarization. The multiple phospho-sites suggest subtle regulation of synaptic vesicle endocytosis by new protein kinases and new protein-protein interactions. The homologous dynI and dynIII phosphorylation indicates a high mechanistic similarity. The results suggest a unique role for the long splice variants of dynI and dynIII in nerve terminals. Synaptic vesicle endocytosis (SVE) 2The abbreviations used are: SVE, synaptic vesicle endocytosis; dyn, dynamin; cdk5, cyclin-dependent kinase 5; MALDI-TOF, matrix-assisted laser desorption ionization time-of-flight; MS, mass spectrometry; MS/MS, tandem mass spectrometry; QqTOF, quadrupole-TOF hybrid; PRD, proline-rich domain; SH3, src-3 homology; TOF, time of flight; sdpnI, syndapin I; amphI, amphiphysin I; GST, glutathione S-transferase; endoI, endophilin I; LC, liquid chromatography; HPLC, high-performance liquid chromatography. is triggered by a coordinated dephosphorylation of a group of at least eight proteins called the dephosphins (1Cousin M.A. Robinson P.J. Trends Neurosci. 2001; 24: 659-665Abstract Full Text Full Text PDF PubMed Scopus (275) Google Scholar). The dephosphins are constitutively phosphorylated in nerve terminals, and their collective rephosphorylation after SVE is necessary for maintaining synaptic vesicle recycling and thus synaptic transmission. One such dephosphin is dynamin I (dynI), a large GTPase enzyme that is crucial for the fission stage of SVE (2Sweitzer S.M. Hinshaw J.E. Cell. 1998; 93: 1021-1029Abstract Full Text Full Text PDF PubMed Scopus (551) Google Scholar). During SVE, dynI is dephosphorylated by the calcium-dependent phosphatase calcineurin (3Liu J.P. Sim A.T.R. Robinson P.J. Science. 1994; 265: 970-973Crossref PubMed Scopus (184) Google Scholar) and is subsequently rephosphorylated by cyclin-dependent kinase 5 (cdk5) on Ser774 and Ser778 (4Tan T.C. Valova V.A. Malladi C.S. Graham M.A. Robinson P.J. Cell Biol. PubMed Scopus Google Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). phosphorylation in the proline-rich are to regulate the the src-3 proteins in long of proteins has been to the dynI in we syndapin I as the dynI in vivo and that dynI is crucial to SVE Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google Scholar). phosphorylation of Ser774 and Ser778 by cdk5, been a of on dynI phosphorylation sites and their protein in vivo and in is phosphorylated on Ser857 by in and the phosphorylation of dynI to amphiphysin I and Y. PubMed Scopus Google Scholar). phosphorylation was to in PC12 cells and was to a has been reported that dynI is phosphorylated at Thr780 by cdk5, to S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar). is in in vivo phosphorylation site of not phosphorylation at site Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). that kinases of dynI in the in vivo of has not been H. S. 1994; PubMed Scopus Google P.J. J.P. T.C. PubMed Scopus Google S. J.P. PubMed Scopus Google Y. PubMed Scopus Google Scholar). and are for the kinase in cells on and S. J. Y. J. Biol. Full Text Full Text PDF PubMed Scopus Google J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). The sites are all dynamin the phosphorylation of dynI has been reported in cells is not to S. J. Y. J. Biol. Full Text Full Text PDF PubMed Scopus Google that of S. J. Y. J. Biol. Full Text Full Text PDF PubMed Scopus Google J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). that in vivo phosphorylation of phospho-sites in dynI In we to were in vivo dynI phosphorylation in to Ser774, Ser778, and Ser857, that to for a that of 32P-labeled dynI that of the phosphorylation sites were that dynI was exclusively phosphorylated on residues by acid new dynI phosphorylation sites at and and new dynI phosphorylation sites at and showed that phosphorylation of Thr780 by cdk5 was restricted to in vitro not in vivo and not regulate of and dynI was the dynamin and as Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google Scholar). were the and confirmed by was was of I was and were and of proteins were in and to the synaptosomes were rat brain and 32P as P.J. J.P. T.C. PubMed Scopus Google Scholar). synaptosomes were for at The synaptosomes were and as (4Tan T.C. Valova V.A. Malladi C.S. Graham M.A. Robinson P.J. Cell Biol. PubMed Scopus Google Scholar). was the to as (4Tan T.C. Valova V.A. Malladi C.S. Graham M.A. Robinson P.J. Cell Biol. PubMed Scopus Google Scholar). and were used as to dynI as Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google Scholar). were in and on and each of were and and 32P-labeled dynI were used in each two-dimensional dynI were and to the for two-dimensional dynI were used in the The were in of in a of The were in at The was to and the were after of was after of was acid after of The was to a of dynI were as Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). the was 5 and to of acid in acid acid in in the of The was a to a and and a of the of The was in and and 32P-labeled dynI was by phosphoamino acid as (4Tan T.C. Valova V.A. Malladi C.S. Graham M.A. Robinson P.J. Cell Biol. PubMed Scopus Google P.J. J.P. T.C. PubMed Scopus Google Scholar). phosphopeptide mapping was on by at J. J.E. in Science. Scholar) followed by in and The were by the and the as J. J.E. in Science. that the was in acid The phosphopeptides were and by matrix-assisted laser desorption spectrometry as Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). two-dimensional phosphopeptide was by the by the at and by the mass of the phosphopeptide as by J. J.E. in Science. Scholar). The in the is on relative was the Y. J. PubMed Scopus Google Scholar) as The was by for each group to for the of phosphorylated and at of dynI phosphopeptides was the a of in 5 was a of the at The was acid in to and in to in to in and to in The was a a quadrupole-TOF a on the the of phosphopeptides of a mass the was at the of the phosphopeptide in a was by a the were for The for the was the of a the of a was In the was first to the phosphopeptides reported in were in at least In on was phosphorylated by in a of and at Phosphorylation for and were by of and were by and were by each of in vitro phosphorylated and to were as and for phosphopeptides as Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). The phosphopeptides were by by the at a as rat brain was as Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google Scholar). proteins were the of at for were in the of were in and by and I was The and to and in dynI were reported (4Tan T.C. Valova V.A. Malladi C.S. Graham M.A. Robinson P.J. Cell Biol. PubMed Scopus Google Scholar). rat brain synaptosomes were as P.J. J.P. T.C. PubMed Scopus Google Scholar). were separated by on and to were by the of by all in vivo phosphorylation sites in dynI rat brain we used a of and all dynI as by and reported that the of dynI of phosphorylation Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google Scholar). has that the of dynI is S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar). we the of proteins of amphI, endoI, and to dynI by and a large of dynI protein and 32P-labeled dynI. of the was to dynI in the first was dynI protein and in the first and In contrast, was dynI and in the first and is in a that is to dynI phosphorylation in vitro Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google Scholar) and that dynI protein after SH3, SH3, and the of and dynI the and of was to all of dynI was in the thus used of in all to all of in vivo phosphorylation on phosphorylation on Thr780 has been J. Cell Biol. PubMed Scopus Google Scholar). was not in a in vivo Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar) was to after in vitro phosphorylation cdk5 S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar). to a protein is phosphorylated on residues is by phosphoamino acid after 32P phosphorylation sites that are not in dynI sites are to to SVE, not 32P in of The first of phosphoamino acid on dynI showed that was phosphorylated P.J. PubMed Scopus Google Scholar). The of dynI in that was the of a by protein The was reported dynI was by a the Phosphorylation at was (4Tan T.C. Valova V.A. Malladi C.S. Graham M.A. Robinson P.J. Cell Biol. PubMed Scopus Google Scholar). In we phosphoamino acid dynI by 32P-labeled synaptosomes and was phosphorylated that dynI is exclusively phosphorylated on residues 32P on and of in of dynI 32P-labeled synaptosomes was to two-dimensional phosphopeptide mapping a of dynI synaptosomes for was in The 32P-labeled phosphopeptides were by on the two-dimensional The of the radiation was in been to phosphopeptides and the dynI Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google Scholar). and were to exclusively and and were to phosphorylated that are phosphorylated at Ser774 and Ser778 Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). of Phosphorylation in for all the 32P that is the dynI phosphorylation the phosphopeptides each on the two-dimensional were the and by and tandem the dynI not that we Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google Scholar). relative radiation and were by MS, phosphopeptides were phosphopeptides were in and of radiation and and the tail of the long splice variant were in and The in was to Ser851 Ser857 is phosphorylated not phosphorylation site was in of phosphopeptides of The in I was to Ser851 and Ser857 are phosphorylated was and and as a and was revealed as the for the phosphorylation site in the the and In to the sites in the dynI Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google to and regulation and protein that was to phosphatase was by in was used to by the at The was to Ser853 is phosphorylated was to dynI synaptosomes Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). and and large were on the of the the of the phosphorylation site was by crucial such as the and the phosphorylation site to the of Ser853 In to the homologous phosphorylation of the dynI and dynIII Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google we revealed that has a in vivo phosphorylation site in a that is homologous to the dynIxa for site is in the splice variant of was by MS, and a phosphopeptide was as confirmed by dephosphorylation phosphatase not was to The was dynI is phosphorylated and the were used to the phosphorylation site and the of the to and to a at were not and were the phosphorylation site to is the in the dynIxa that was to phosphorylated in we used to of minor phosphorylation of phosphopeptides after two-dimensional mapping not to to to the of phosphopeptides on 32P we used to for all phosphopeptides a dynI and new dynI phosphorylation sites were found. of the at a unique phosphopeptide the of dynI is The was by The phosphorylation site was as the and In the we and that was phosphorylated in phosphopeptide not of the at unique phosphopeptide dynI is the of phosphorylation the of The phosphorylation site was to at the residues phosphorylated to and after the residues is the first time that in vivo dynI sites been of the is that sites were 32P-labeled were not by the and in the two-dimensional in were the the for each of the dynI and dynIII to are The the of the phosphopeptides and In the the was a of for the large in and on mass the has and and The phosphopeptides and are to in the the at phosphorylated for at least of the in the of are and and for by to phosphorylated at Ser851 Ser857. The of to a and to the of and for is for and 32P-labeled in depolarization. the of dynI and minor phosphopeptides in the two-dimensional of the 32P the of in each a of the in vivo of each site in of that that to 32P was not in The 32P-labeled were by and the of each was and as a of the radiation in all Using the of phosphopeptide for each was to that and were 32P-labeled in a of to Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google Scholar). to all phosphorylation The phosphopeptides in each are each of the 32P and Ser774 Ser774 Ser778. reported sites 69% of the dynI. that a of radiation is dynIII Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google was in the that dynIII phosphopeptides are and the homologous dynI phosphopeptides (4Tan T.C. Valova V.A. Malladi C.S. Graham M.A. Robinson P.J. Cell Biol. PubMed Scopus Google Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). was dynIII Ser853 and of the 32P and I for of the The of the 32P was in was a of Ser851 Ser857 The of was in was phosphorylated on the phosphorylation is of the radiation and a of of the 32P to Ser512, the of sites phosphopeptides proteins of the dynI and of the sites were is to in to the has of the dynI all the sites to in of and of 32P radiation on all dynI and dynIII phosphorylation sites two-dimensional phosphopeptide mapping and of 32P 32P in a new To the biological of each phosphorylation site we sites were to a measure of the of each site to The by each phosphopeptide in was was and to the phosphorylation site and The of 32P-labeled was by and was by the of radiation dynIII and in was the phosphopeptide the sites Ser851 and Ser857 were not the phosphorylation was by of the phosphorylation sites in dynIxa are dephosphorylated depolarization. The phosphopeptide was dephosphorylated by and in the phosphorylation of was 32P was of as we was was in the phosphopeptide to Ser347. Phosphorylation of Thr780 on to in was reported that dynI is phosphorylated by cdk5 at Thr780 in vitro and that all phosphorylation is by of Thr780 to S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar). suggest Thr780 is the sole phosphorylation site in the dynI and for Thr780 phosphorylation in vivo Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). Mass spectrometry is not a for discrepancy used to the of a phosphorylation we used a to a on Thr780 phosphorylation of dynI on Ser774 Ser778 by cdk5 in The results of S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar). a of of Ser774, Ser778, Thr780 to in for the of the phosphorylation of the by of Ser774 and Ser778 on the of phosphorylation by cdk5, as by and by was a on Thr780 and all sites in the were to phosphorylation was of and that of of sites results in a in phosphorylation at of the is all sites are that a in phosphorylation was in the that of the phosphorylation in the a major sites in the The dynI used in was on the long splice variant are at least sites for phosphorylation by Ser822, Ser851, and Ser857. all of for the phosphorylation in Using tandem mass we confirmed that the of the in vitro phosphorylated was phosphorylated at Ser774, Ser778, and Thr780 in vitro phosphorylation at Thr780 was by the dynI The phosphorylation at Thr780 was by the and The for the phosphorylated to the of F. PubMed Scopus Google was a that the phosphorylation site on was to of was a in the of the and phosphorylation to the phosphorylated was the of a in vitro by S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar). The results that a in a protein that is phosphorylated is to in vitro Thr780 is as in vitro phosphorylation not that Thr780 is phosphorylated in systematic the results and of S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar). of are several that to dynI is by the in vitro phosphorylation of dynI S. Y. H. J. Cell Biol. PubMed Scopus Google Science. 1998; PubMed Scopus (275) Google Scholar). To in vitro we a of at Ser774, Ser778, and Thr780 in and not the of as by The results were confirmed by of multiple a to dynI in the of the to the of dynI protein to the of phosphorylated dynI was at high that to dynI is not by dynI phosphorylation The that has is not to at the dynI and dynI. we used to the of dynI of synaptosomes for was to dynI dephosphorylation P.J. Neurosci. PubMed Scopus Google Scholar) by J. PubMed Scopus Google Scholar). synaptosomes and to was each of was dephosphorylated as by the the and dynI was not and we that is not a for in vivo in Phosphorylation was used to identify and all the phosphorylation sites in dynI. Mass spectrometry of 32P-labeled dynI phosphopeptides separated by two-dimensional mapping revealed seven in vivo phosphorylation sites at Ser774, Ser778, Ser822, Ser851, Ser857, Ser512, and Thr780 phosphorylation was not was that a Ser857 was phosphorylated in vitro by Y. PubMed Scopus Google a Ser851 was In we confirmed the of Ser857 as in vivo phosphorylation site Y. PubMed Scopus Google Scholar) and Ser851 as a new in vivo phosphorylation site the sites are restricted to the tail of the splice dynIxa that Ser851 and Ser857 were phosphorylated to a and was to their has been that Ser857 phosphorylation dynI to Y. PubMed Scopus Google Scholar). that dynI phosphorylation not of to the of phosphorylation at Ser857 on is of the radiation and we on cdk5 the The phosphorylation site at is the of the is not the on site is we were to a the of the were for on is in SVE, is in dynI The phosphorylation at is to the GTPase and on of the is to in dynI and J. PubMed Scopus Google Scholar). The of a role for the site on site is to of 32P in a of and is to role for in and are not in of a is in a on the of the phosphorylation not phosphorylated by the protein and acid followed by on the we that is at least protein cdk5 Y. PubMed Scopus Google that dynI in is in of a and phosphorylated by of the protein kinases that Ser774, Ser778, Ser851, Ser857. is on by The to are in and are called and J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). is that by the of Ser774 and Ser778 was revealed the of dephosphorylation of each site was showed that and are dephosphorylated to a the of phosphorylated dynI Ser774 and Ser778 was in to the phosphorylated a relative on the of that dynI phosphorylation at Ser774 has a role to phosphorylation at Ser778 in Phosphorylation at Thr780 in J. Cell Biol. PubMed Scopus Google was reported that cdk5 dynI at Ser774 and Ser778 in (4Tan T.C. Valova V.A. Malladi C.S. Graham M.A. Robinson P.J. Cell Biol. PubMed Scopus Google Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar) and at Thr780 in S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar). we Thr780 is phosphorylated in vitro by cdk5 and phosphorylation to a in vivo S. Y. H. J. Cell Biol. PubMed Scopus Google is that dynI is phosphorylated at Thr780 in vivo in synaptosomes the of The is that phosphorylation at Thr780 is restricted to in vitro is of cdk5 phosphorylation in that Thr780 is not phosphorylated by cdk5 in phosphoamino acid of dynI 32P-labeled synaptosomes revealed phosphorylation of dynI on the of Thr780 phosphorylation to a phosphorylation site not after in synaptosomes to not in synaptosomes at we were to Thr780 phosphorylation by two-dimensional phosphopeptide mapping of 32P-labeled dynI and the of the of a major of dynI phosphorylated on such of the phosphorylation in dynI. a of Thr780 to not cdk5 phosphorylation in the in to reported by S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar). a on Ser774 and Ser778, on dynI phosphorylation by cdk5 in was all sites at Ser774, Ser778, and Thr780 were to that phosphorylation by cdk5 was that is a phosphorylation of sites the site is In we in phosphoamino acid that dynI was phosphorylated by cdk5 in vitro was dynI was 32P-labeled synaptosomes (4Tan T.C. Valova V.A. Malladi C.S. Graham M.A. Robinson P.J. Cell Biol. PubMed Scopus Google Scholar). the splice variants are The long was used in are in the the used by S. Y. H. J. Cell Biol. PubMed Scopus Google Scholar) was not reported on in vitro phosphorylation in vivo phosphorylation site not to The discrepancy and that of to the of their to was that the phosphorylation of Thr780 in synaptosomes and brain and was of S. Y. H. J. Cell Biol. PubMed Scopus Google the at is for was not that was a that all Ser774, Ser778, and and was phosphorylated in vitro to of is not to phosphorylation at Ser774 Ser778 and we showed that all are phosphorylated in vitro in the The site of the has not been and is to the results to Thr780 for phosphorylation at the Thr780 is not phosphorylated in Phosphorylation that a of dynIII the dynamin synaptosomes and were to identify the homologous phosphorylation sites to Ser774 and Ser778 for dynIII Graham Robinson P.J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). The acid sites are dynI and In we a new dynIII phosphorylation site at Ser853 that was has been reported to in the J. H. M.A. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). is that the dynIII that 32P we that the 32P-labeled dynIII was Ser853 is homologous to Ser851 of dynI. site is in the long splice that Ser857 dynI is not in dynIII homologous phosphorylation that dynIII to dynI in the nerve phosphorylation on the sites in synaptosomes that the protein kinases and that is a role for Ser853 and Ser851 The acid of the tail of the long splice variants of dynI and dynIII are not for that the phosphorylation site to all the phosphorylation sites in dynIII been the results suggest that the dynIII sites been found. the seven dynI and are in the of of been and phosphorylation at in dynIII is in sites were that were not homologous to dynI. the acid of dynIII are as to that protein are by The results suggest dynIII has a role in nerve a we for phosphorylation of in was in the as by and mass spectrometry not The and not we not that the of is is to In by on The of phosphorylation sites in synaptosomes is not that are is sites in dynIII were at of the seven dynI phospho-sites are in the of Ser774 is in Ser778 and are and are in the acid are The tail of is not to the as dynI and is in to the long tail of of are Ser857 The is that Ser851 Ser853 in has of the sites in of the and not to phosphorylated by a protein kinase in The of of phosphorylation of at and S. J. Y. J. Biol. Full Text Full Text PDF PubMed Scopus Google J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar) is not the synaptosomes were not sites phosphorylated in after the of dynI in to all the in vivo phosphorylation sites in dynI. is to that all sites in protein been reported sites were not Thr780 S. Y. H. J. Cell Biol. PubMed Scopus Google and in dynI S. J. Y. J. Biol. Full Text Full Text PDF PubMed Scopus Google S. S. Y. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar) and J. Biol. Full Text Full Text PDF PubMed Scopus Google Valova V.A. Malladi C.S. Robinson P.J. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). that phosphorylation of Thr780 and is restricted to in vitro for phosphorylation on several Phosphorylation of and was first in dynI thus phosphorylation in in vivo we for phosphorylation of dynI on and for a of sites in dynI phosphorylated in at of is that phosphopeptides not and been of phosphopeptides are not mass spectrometry not the of such phosphopeptides to such we that new sites in the of the phosphorylation, the of the 32P on dynI has been The on are the that a phosphopeptide been in of the the of high phosphorylation sites that not 32P after The of the sites in the of and of 32P and mass spectrometry has been to in vivo phosphorylation The of new in vivo dynI phosphorylation sites at Ser512, Ser822, Ser851, and in to the sites Ser778, and a for of the of dynI and to new phosphorylation sites in the regulate of syndapin I Graham Valova V.A. M.A. Robinson P.J. Neurosci. PubMed Scopus Google a role in the of endocytosis a major in a of the large of for