title: Synaptotagmin III is a critical factor for the formation of the perinuclear endocytic recycling compartment and determination of secretory granules size.
abstract: Early endosomes and a perinuclear, Rab-11-positive compartment have been implicated in the recycling of internalized receptors. In this study, we show that synaptotagmin III (Syt III), a member of the Syt family of proteins, is required for the formation and delivery of cargo to the perinuclear endocytic recycling compartment (ERC). We demonstrate that rat basophilic leukemia (RBL-2H3) mast cells endogenously express Syt III, and >70% of this protein colocalizes with early endosomal markers, such as EEA1, annexin II and syntaxin 7, and the remaining protein colocalizes with secretory granule (SG) markers such as beta-hexosaminidase, histamine and serotonin. To study the functional role of Syt III, we stably transfected RBL cells with Syt III antisense cDNA and monitored the route of transferrin (Tfn) internalization in cells that displayed substantially reduced (<90%) levels of Syt III (RBL-Syt III(-)). In these cells, Tfn binding and internalization into early endosomes were unaltered. However, whereas in the mock-transfected cells Tfn was subsequently delivered to the ERC, in the RBL-Syt III(-) cells, Tfn remained associated with dispersed peripheral vesicles and Rab 11 remained cytosolic. Nevertheless, the rates of Tfn internalization and recycling were not affected. RBL-Syt III(-) cells also displayed enlarged SGs, reminiscent of the SGs present in Chediak-Higashi (beige) mice. However, morphometric analyses suggested that granule formation was unaltered and that the calculated unit granule volume is the same in both cell lines. Therefore, our results implicate Syt III as a critical factor for the generation and delivery of internalized cargo to the perinuclear endocytic recycling compartment and suggest a possible link between ERC and recycling from immature SGs during the process of SG maturation.
authors.affiliation: Department of Cell Biology and Histology, Sackler School of Medicine, Tel Aviv University, Tel Aviv, 69978, Israel.
authors.forename: Elena
authors.initials: E
authors.lastname: Grimberg
authors.forename: Ze
authors.initials: Z
authors.lastname: Peng
authors.forename: Ilan
authors.initials: I
authors.lastname: Hammel
authors.forename: Ronit
authors.initials: R
authors.lastname: Sagi-Eisenberg
mesh_terms: D000818:Animals; D015415:Biomarkers; D002135:Calcium-Binding Proteins; D002451:Cell Compartmentation / Q000235:genetics; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure; D048429:Cell Size / Q000235:genetics; D003593:Cytoplasm / Q000378:metabolism* / Q000648:ultrastructure; D004705:Endocytosis / Q000235:genetics*; D008407:Mast Cells / Q000378:metabolism* / Q000648:ultrastructure; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000172:deficiency* / Q000235:genetics; D000090204:Mesothelin; D008854:Microscopy, Electron; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000172:deficiency* / Q000235:genetics; D020319:Oligodeoxyribonucleotides, Antisense; D011485:Protein Binding / Q000235:genetics; D021381:Protein Transport / Q000235:genetics; D051381:Rats; D011956:Receptors, Cell Surface / Q000378:metabolism; D022142:Secretory Vesicles / Q000378:metabolism* / Q000648:ultrastructure; D050857:Synaptotagmins; D014168:Transferrin / Q000378:metabolism; D022161:Transport Vesicles / Q000378:metabolism* / Q000648:ultrastructure; D014407:Tumor Cells, Cultured; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000099193:rab11 GTP-Binding Proteins
chemical_list: D015415:Biomarkers; D002135:Calcium-Binding Proteins; D008562:Membrane Glycoproteins; D000090204:Mesothelin; D009419:Nerve Tissue Proteins; D020319:Oligodeoxyribonucleotides, Antisense; D011956:Receptors, Cell Surface; D050857:Synaptotagmins; D014168:Transferrin; D020691:rab GTP-Binding Proteins; D000099193:rab11 GTP-Binding Proteins; C000715529:Msln protein, rat; C494291:Syt3 protein, rat