title: Distinct and opposing roles for Rab27a/Mlph/MyoVa and Rab27b/Munc13-4 in mast cell secretion.
abstract: Mediator release from mast cells is a critical step in allergic and inflammatory disease. However, the processes regulating the latter stages of granule release are yet to be fully understood. Rab27 small GTPases regulate release of secretory lysosomes in a variety of cells, including mast cell granules. In the present study, using murine bone marrow-derived mast cells (BMMC) from Rab27-deficient mutant mice, we found that, in contrast to Rab27b, Rab27a primarily plays an inhibitory role in regulating degranulation. Immunofluorescence analysis revealed that resting Rab27a-deficient (ashen) BMMCs display abnormal cortical F-actin distribution. Actin disassembly prior to IgE cross-linking increased wild-type BMMC secretion to ashen levels, suggesting that changes in the integrity of cortical F-actin underlie the ashen phenotype. Comparison of the secretory impairment of Rab27b knockout and Rab27a/b double knockout BMMCs highlighted a secondary positive role for Rab27a in enhancing degranulation. Rab27 is known to interact with actin via its effectors melanophilin (Mlph) and myosin Va (MyoVa) in other cell types. To better understand the differing roles of Rab27 proteins, we analysed the secretory phenotype of BMMCs derived from mice lacking Rab27 effector proteins. These experiments revealed that the phenotype of BMMCs deficient in Mlph (leaden) and BMMCs deficient in MyoVa (dilute) resembles the hyper-secretion of ashen BMMCs, while Munc13-4-deficient (jinx) BMMCs phenocopy the Rab27b knockout and double Rab27a/b knockout secretory impairment. We conclude that Rab27a and Rab27b regulate distinct steps in the BMMC degranulation pathway, with Rab27a/Mlph/MyoVa regulating cortical actin stability upstream of Rab27a/b/Munc13-4-dependent granule exocytosis.
authors.affiliation: Molecular Medicine, National Heart and Lung Institute, Imperial College London, London, UK.
authors.forename: Rajesh K
authors.initials: RK
authors.lastname: Singh
authors.forename: Koichi
authors.initials: K
authors.lastname: Mizuno
authors.forename: Christina
authors.initials: C
authors.lastname: Wasmeier
authors.forename: Silene T
authors.initials: ST
authors.lastname: Wavre-Shapton
authors.forename: Chiara
authors.initials: C
authors.lastname: Recchi
authors.forename: Sergio D
authors.initials: SD
authors.lastname: Catz
authors.forename: Clare
authors.initials: C
authors.lastname: Futter
authors.forename: Tanya
authors.initials: T
authors.lastname: Tolmachova
authors.forename: Alistair N
authors.initials: AN
authors.lastname: Hume
authors.forename: Miguel C
authors.initials: MC
authors.lastname: Seabra
mesh_terms: D000199:Actins / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001854:Bone Marrow Cells / Q000378:metabolism / Q000502:physiology; D015550:Cell Degranulation / Q000235:genetics / Q000502:physiology; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism; D005089:Exocytosis / Q000235:genetics / Q000502:physiology; D005260:Female; D015151:Immunoblotting; D008297:Male; D008407:Mast Cells / Q000378:metabolism* / Q000502:physiology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D018995:Myosin Heavy Chains / Q000235:genetics / Q000378:metabolism*; D024701:Myosin Type V / Q000235:genetics / Q000378:metabolism*; D017455:Receptors, IgE / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000075745:rab27 GTP-Binding Proteins
chemical_list: D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D008565:Membrane Proteins; C435724:Mlph protein, mouse; C513082:Myo5a protein, mouse; D017455:Receptors, IgE; C520225:Unc13d protein, mouse; D000075745:rab27 GTP-Binding Proteins; D024701:Myosin Type V; C512958:Rab27b protein, mouse; C512962:Rab27a protein, mouse; D018995:Myosin Heavy Chains; D020691:rab GTP-Binding Proteins