Binding of Rab3A to Synaptic Vesicles

Chou, Judy H.; Jahn, Reinhard

doi:10.1074/jbc.275.13.9433

Cited by 32 publications

(24 citation statements)

References 47 publications

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“…Monoclonal antibodies specific for Rab3A (Cl42.2 [37]), syncollin (Cl87.1 [2]), synaptobrevin 2 (Cl69.1 [16]), GDI (Cl81.1 [14]), and rabphilin (clone Cl76.2 [5]) as well as polyclonal antibodies specific for Rab3B (O. M. Schluetter et al, unpublished data); Rab3C (20); endobrevin (19); and syntaxins 2, 3, 4, 7, and 13 were described previously (4,5,25) and are commercially available from Synaptic Systems (Göttingen, Germany). Cellubrevin (3) and syntaxin 8 (4) antisera were described previously.…”

Section: Methodsmentioning

confidence: 99%

Rab3D Is Not Required for Exocrine Exocytosis but for Maintenance of Normally Sized Secretory Granules

Riedel

Antonin

Fernández‐Chacón

et al. 2002

Molecular and Cellular Biology

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127

106

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Rab3D, a member of the Rab3 subfamily of the Rab/ypt GTPases, is expressed on zymogen granules in the pancreas as well as on secretory vesicles in mast cells and in the parotid gland. To shed light on the function of Rab3D, we have generated Rab3D-deficient mice. These mice are viable and have no obvious phenotypic changes. Secretion of mast cells is normal as revealed by capacitance patch clamping. Furthermore, enzyme content and overall morphology are unchanged in pancreatic and parotid acinar cells of knockout mice. Both the exocrine pancreas and the parotid gland show normal release kinetics in response to secretagogue stimulation, suggesting that Rab3D is not involved in exocytosis. However, the size of secretory granules in both the exocrine pancreas and the parotid gland is significantly increased, with the volume being doubled. We conclude that Rab3D exerts its function during granule maturation, possibly by preventing homotypic fusion of secretory granules.

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Section: Methodsmentioning

confidence: 99%

Rab3D Is Not Required for Exocrine Exocytosis but for Maintenance of Normally Sized Secretory Granules

Riedel

Antonin

Fernández‐Chacón

et al. 2002

Molecular and Cellular Biology

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127

106

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“…Monoclonal antibodies specific for syncollin (Cl 87.1 [2]), synaptobrevin 2 (Cl 69.1 [12]), and GDP dissociation inhibitor (Cl 81.1 [9]) as well as polyclonal antibodies specific for syntaxin 4 (18) were described previously and are commercially available from Synaptic Systems (Göttingen, Germany). A monoclonal antibody specific for rabphilin (clone Cl76.2; subclass immunoglobulin M) was raised against recombinant rabphilin using standard procedures (24).…”

Section: Methodsmentioning

confidence: 99%

Loss of the Zymogen Granule Protein Syncollin Affects Pancreatic Protein Synthesis and Transport but Not Secretion

Antonin¹,

Wagner

Riedel³

et al. 2002

Molecular and Cellular Biology

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Syncollin is a small protein that is abundantly expressed in pancreatic acinar cells and that is tightly associated with the lumenal side of the zymogen granule membrane. To shed light on the hitherto unknown function of syncollin, we have generated syncollin-deficient mice. The mice are viable and show a normal pancreatic morphology as well as normal release kinetics in response to secretagogue stimulation. Although syncollin is highly enriched in zymogen granules, no change was found in the overall protein content and in the levels of chymotrypsin, trypsin, and amylase. However, syncollin-deficient mice reacted to caerulein hyperstimulation with a more severe pancreatitis. Furthermore, the rates of both protein synthesis and intracellular transport of secretory proteins were reduced. We conclude that syncollin plays a role in maturation and/or concentration of zymogens in zymogen granules.The exocrine pancreas is specialized for the synthesis, storage, and release of digestive proenzymes. Since the pancreas exhibits the highest rate of protein synthesis among all mammalian tissues (8), it has served for many years as the model system for the study of intracellular protein transport and secretion (31). In pancreatic acinar cells, secretory proteins are synthesized at the rough endoplasmic reticulum and transported to the Golgi apparatus. In the trans-Golgi network, proteins are segregated into condensing vacuoles that undergo further maturation to zymogen granules (for a review, see reference 37). During maturation and condensation, granular content proteins are segregated from proteins destined for constitutive secretion (for a review, see references 3 and 43). Mature granules are stored in the apical pole of the acinar cell. Upon nervous or hormonal stimulation, a selective rise in the local Ca 2ϩ concentration is observed in the apical pole, which triggers exocytotic discharge of the granule content into the pancreatic duct (32).Formation, maturation, and exocytosis of zymogen granules are multistep processes that are only incompletely understood at the molecular level. For instance, it is largely unknown how zymogens are sorted in the trans-Golgi network and how they are packaged and concentrated in zymogen granules. Furthermore, the molecular steps involved in granule docking and fusion still need to be elucidated. As in many other intracellular fusion events, members of the Rab and SNARE (for soluble N-ethylmaleimide sensitive factor attachment protein receptor) protein families were shown to be involved in pancreatic secretion (16,18,29,30,44), but how they are regulated and which proteins are involved in transmitting the calcium signal to the exocytotic machinery are still unknown.

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“…The protein concentrations from both membrane and cytosol fractions were measured using the Micro BCA protein assay kit. The expression of p12 I , Ras-related GTP binding protein 1B (Rab1B), and Fas-associated death domain protein (FADD) in both membrane and cytosol fractions were tested by immunoblot analysis using mouse monoclonal anti-HA antibody, rabbit polyclonal anti-Rab1B antibody (Zymed), and mouse monoclonal anti-FADD antibody (Transduction Lab), respectively, as described previously (7,27,45).…”

Section: Methodsmentioning

confidence: 99%

Endoplasmic Reticulum andcis-Golgi Localization of Human T-Lymphotropic Virus Type 1 p12^I: Association with Calreticulin and Calnexin

Ding¹,

Albrecht²,

Luo³

et al. 2001

J Virol

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Human T-lymphotropic virus type 1 (HTLV-1) is a complex retrovirus encoding regulatory and accessory genes in four open reading frames (ORF I to IV) of the pX region. We have demonstrated an important role of pX ORF I expression, which encodes p12I , in establishment of HTLV-1 infection in a rabbit model and for optimal viral infectivity in quiescent primary lymphocytes. These data indicated that p12 I may enhance lymphocyte activation and thereby promote virus infection. To further define the role of p12 I in cell activation, we characterized the subcellular localization of p12 I in transfected 293T cells and HeLa-Tat cells by multiple methods, including immunofluorescence confocal microscopy, electron microscopy, and subcellular fractionation. Herein, we demonstrate that p12 I accumulates in the endoplasmic reticulum (ER) and cis-Golgi apparatus. The location of p12 I was unchanged following treatments with both cycloheximide (blocking de novo protein synthesis) and brefeldin A (disrupting ER-to-Golgi protein transport), indicating that the protein is retained in the ER and cis-Golgi. Moreover, using coimmunoprecipitation assays, we identify the direct binding of p12 I with both calreticulin and calnexin, resident ER proteins which regulate calcium storage. Our results indicate that p12 I directly binds key regulatory proteins involved in calcium-mediated cell signaling and suggest a role of p12 I in the establishment of HTLV-1 infection by activation of host cells.Human T-lymphotropic virus type 1 (HTLV-1) is the etiologic agent of adult T-cell leukemia and lymphoma and appears to initiate a variety of immune-mediated disorders, including the chronic neural degenerative disease HTLV-1-associated myelopathy/tropical spastic paraparesis (16,44). As a complex retrovirus, HTLV-1 contains the common retroviral genes gag, pol, and env, as well as several regulatory and accessory genes. These regulatory and accessory genes are present in four different open reading frames (ORFs) in the pX region between env and the 3Ј long terminal repeat (LTR) (10,15,39,40). ORFs IV and III encode the regulatory proteins Tax and Rex, respectively, which have been extensively characterized. Tax is a 40-kDa nuclear-localizing protein that increases viral transcription from the HTLV-1 LTR as well as a number of cellular genes involved in host cell proliferation (17,30,34). Rex is a 27-kDa nucleolar-localizing protein that acts at the posttranscriptional level by promoting the cytoplasmic accumulation of unspliced and singly spliced viral RNA (29).Recent studies have provided important new data that indicate a role of the highly conserved ORF I-encoded protein p12 I in HTLV-1 infection. ORF I mRNA has been detected in HTLV-1-infected cells derived from patients with both adult T-cell leukemia and lymphoma and HTLV-1-associated myelopathy/tropical spastic paraparesis and from asymptomatic carriers (3, 5, 9, 10). Moreover, recombinant p12 I is recognized by sera from naturally infected humans and experimentally infected rabbits (14). Pep...

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Binding of Rab3A to Synaptic Vesicles

Cited by 32 publications

References 47 publications

Rab3D Is Not Required for Exocrine Exocytosis but for Maintenance of Normally Sized Secretory Granules

Rab3D Is Not Required for Exocrine Exocytosis but for Maintenance of Normally Sized Secretory Granules

Loss of the Zymogen Granule Protein Syncollin Affects Pancreatic Protein Synthesis and Transport but Not Secretion

Endoplasmic Reticulum andcis-Golgi Localization of Human T-Lymphotropic Virus Type 1 p12^I: Association with Calreticulin and Calnexin

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Binding of Rab3A to Synaptic Vesicles

Cited by 32 publications

References 47 publications

Rab3D Is Not Required for Exocrine Exocytosis but for Maintenance of Normally Sized Secretory Granules

Rab3D Is Not Required for Exocrine Exocytosis but for Maintenance of Normally Sized Secretory Granules

Loss of the Zymogen Granule Protein Syncollin Affects Pancreatic Protein Synthesis and Transport but Not Secretion

Endoplasmic Reticulum andcis-Golgi Localization of Human T-Lymphotropic Virus Type 1 p12I: Association with Calreticulin and Calnexin

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Endoplasmic Reticulum andcis-Golgi Localization of Human T-Lymphotropic Virus Type 1 p12^I: Association with Calreticulin and Calnexin