Search PubMed⌕ Search

Biomedical subjects

Kira E Poskanzer

Publications and source records attributed to Kira E Poskanzer.

5 recordsLinked to original sources
PubMed2006

Discrete residues in the c(2)b domain of synaptotagmin I independently specify endocytic rate and synaptic vesicle size.

It has been demonstrated that synapses lacking functional synaptotagmin I (Syt I) have a decreased rate of synaptic vesicle endocytosis. Beyond this, the function of Syt I during endocytosis remains undefined. Here, we demonstrate that a decreased rate of endocytosis in syt(null) mutants correlates with a stimulus-dependent perturbation of membrane internalization, assayed ultrastructurally. We then separate the mechanisms that control endocytic rate and vesicle size by mapping these processes to discrete residues in the Syt I C(2)B domain. Mutation of a poly-lysine motif alters vesicle size but not endocytic rate, whereas the mutation of calcium-coordinating aspartate residues (syt-D3,4N) alters endocytic rate but not vesicle size. Finally, slowed endocytic rate in the syt-D3,4N animals, but not syt(null) animals, can be rescued by elevating extracellular calcium concentration, supporting the conclusion that calcium coordination within the C(2)B domain contributes to the control of endocytic rate.

Adaptor Protein Complex alpha Subunits↗
PubMed2004

Dap160/intersectin scaffolds the periactive zone to achieve high-fidelity endocytosis and normal synaptic growth.

Dap160/Intersectin is a multidomain adaptor protein that colocalizes with endocytic machinery in the periactive zone at the Drosophila NMJ. We have generated severe loss-of-function mutations that eliminate Dap160 protein from the NMJ. dap160 mutant synapses have decreased levels of essential endocytic proteins, including dynamin, endophilin, synaptojanin, and AP180, while other markers of the active zone and periactive zone are generally unaltered. Functional analyses demonstrate that dap160 mutant synapses are unable to sustain high-frequency transmitter release, show impaired FM4-64 loading, and show a dramatic increase in presynaptic quantal size consistent with defects in synaptic vesicle recycling. The dap160 mutant synapse is grossly malformed with abundant, highly ramified, small synaptic boutons. We present a model in which Dap160 scaffolds both endocytic machinery and essential synaptic signaling systems to the periactive zone to coordinately control structural and functional synapse development.

Adaptor Proteins, Signal Transducing↗
PubMed2004

Mobilization and fusion of a non-recycling pool of synaptic vesicles under conditions of endocytic blockade.

At vertebrate central synapses, it has been demonstrated that a resting pool of synaptic vesicles (SVs) exists that normally does not participate in SV release and recycling. It remains unclear whether SVs within the resting pool are capable of mobilization and fusion. Here, we combine live imaging of SV exo- and endocytosis using pH-sensitive GFP (synapto-pHluorins) with pharmacological and genetic manipulations of the SV cycle at the Drosophila NMJ. We demonstrate that a resting pool of SVs exists at this synapse that encompasses 30-41% of the total SV pool. Under conditions of endocytic blockade, using a temperature-sensitive dynamin mutation, the resting pool of SVs can be mobilized and released. We present a model for the presence of a resting pool of SVs that does not require molecular specification of a subpopulation of SVs.

Animals↗
PubMed2003

Synaptotagmin I is necessary for compensatory synaptic vesicle endocytosis in vivo.

Neurotransmission requires a balance of synaptic vesicle exocytosis and endocytosis. Synaptotagmin I (Syt I) is widely regarded as the primary calcium sensor for synaptic vesicle exocytosis. Previous biochemical data suggest that Syt I may also function during synaptic vesicle endocytosis; however, ultrastructural analyses at synapses with impaired Syt I function have provided an indirect and conflicting view of the role of Syt I during synaptic vesicle endocytosis. Until now it has not been possible experimentally to separate the exocytic and endocytic functions of Syt I in vivo. Here, we test directly the role of Syt I during endocytosis in vivo. We use quantitative live imaging of a pH-sensitive green fluorescent protein fused to a synaptic vesicle protein (synapto-pHluorin) to measure the kinetics of endocytosis in sytI-null Drosophila. We then combine live imaging of the synapto-pHluorins with photoinactivation of Syt I, through fluorescein-assisted light inactivation, after normal Syt I-mediated vesicle exocytosis. By inactivating Syt I only during endocytosis, we demonstrate that Syt I is necessary for the endocytosis of synaptic vesicles that have undergone exocytosis using a functional Syt I protein.

Animals↗