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Pericentrosomal targeting of Rab6 secretory vesicles by Bicaudal-D-related protein 1 (BICDR-1) regulates neuritogenesis.

Schlager MA, Kapitein LC, Grigoriev I, Burzynski GM, Wulf PS, Keijzer N, de Graaff E, Fukuda M, Shepherd IT, Akhmanova A, Hoogenraad CC - EMBO J. (2010)

Bottom Line: BICDR-1 expression is high during early neuronal development and strongly declines during neurite outgrowth.Later during development, BICDR-1 expression is strongly reduced, which permits anterograde secretory transport required for neurite outgrowth.These results indicate an important role for BICDR-1 as temporal regulator of secretory trafficking during the early phase of neuronal differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Erasmus Medical Center, Rotterdam, The Netherlands.

ABSTRACT
Membrane and secretory trafficking are essential for proper neuronal development. However, the molecular mechanisms that organize secretory trafficking are poorly understood. Here, we identify Bicaudal-D-related protein 1 (BICDR-1) as an effector of the small GTPase Rab6 and key component of the molecular machinery that controls secretory vesicle transport in developing neurons. BICDR-1 interacts with kinesin motor Kif1C, the dynein/dynactin retrograde motor complex, regulates the pericentrosomal localization of Rab6-positive secretory vesicles and is required for neural development in zebrafish. BICDR-1 expression is high during early neuronal development and strongly declines during neurite outgrowth. In young neurons, BICDR-1 accumulates Rab6 secretory vesicles around the centrosome, restricts anterograde secretory transport and inhibits neuritogenesis. Later during development, BICDR-1 expression is strongly reduced, which permits anterograde secretory transport required for neurite outgrowth. These results indicate an important role for BICDR-1 as temporal regulator of secretory trafficking during the early phase of neuronal differentiation.

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BICDR-1 interacts with the dynein/dynactin motor complex. (A) Immunoprecipitations from extracts of Hela cells transfected with the indicated constructs and probed for DHC, p150glued or dynein intermediate chain 74 (IC74). (B) Ratio of pericentrosomal versus cytoplasmic p150glued or IC74 fluorescence intensity in cells with and without GFP-BICDR-1 overexpression (average ±s.e.m.; p150glued control, n=29; p150glued+BICDR-1, n=30; IC74 control, n=32; IC74+BICDR-1, n=19 cells). (C) Percentage of cells, transfected with indicated siRNAs, with endogenous pericentrosomal BICDR-1 (average ±s.e.m.; control, n=3040; DHC siRNA, n=533; p150glued, n=792 cells). (D) Representative image of a Vero cell stained for endogenous BICDR-1 (green) and for p150glued (red). (E) Representative image of Hela cells with and without GFP-BICDR-1 (green) overexpression stained for p150glued (red). (F) Vero cells transfected with p150glued-specific siRNA (right) and untransfected (left). Stained for BICDR-1 (green) and p150glued (red). (D–F) Solid lines indicate the cell edge and dashed lines indicate the nucleus. The insets show magnifications of boxed areas. Scale bars, 10 μm. ***P<0.001.
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f4: BICDR-1 interacts with the dynein/dynactin motor complex. (A) Immunoprecipitations from extracts of Hela cells transfected with the indicated constructs and probed for DHC, p150glued or dynein intermediate chain 74 (IC74). (B) Ratio of pericentrosomal versus cytoplasmic p150glued or IC74 fluorescence intensity in cells with and without GFP-BICDR-1 overexpression (average ±s.e.m.; p150glued control, n=29; p150glued+BICDR-1, n=30; IC74 control, n=32; IC74+BICDR-1, n=19 cells). (C) Percentage of cells, transfected with indicated siRNAs, with endogenous pericentrosomal BICDR-1 (average ±s.e.m.; control, n=3040; DHC siRNA, n=533; p150glued, n=792 cells). (D) Representative image of a Vero cell stained for endogenous BICDR-1 (green) and for p150glued (red). (E) Representative image of Hela cells with and without GFP-BICDR-1 (green) overexpression stained for p150glued (red). (F) Vero cells transfected with p150glued-specific siRNA (right) and untransfected (left). Stained for BICDR-1 (green) and p150glued (red). (D–F) Solid lines indicate the cell edge and dashed lines indicate the nucleus. The insets show magnifications of boxed areas. Scale bars, 10 μm. ***P<0.001.

Mentions: Earlier studies have shown that members of BICD family interact with the microtubule minus-end-directed dynein/dynactin motor complex. Therefore, we investigated the possible interaction between BICDR-1 and dynein/dynactin. Co-immunoprecipitation experiments from Hela cells transfected with GFP-BICDR-1 showed that BICDR-1 precipitates the major dynein/dynactin subunits, whereas no binding is seen with control GFP (Figure 4A). Consistently, both dynactin (Figure 4D) and dynein (Supplementary Figure S5B) co-localize with endogenous BICDR-1 around the centrosome in Vero cells. Overexpression of GFP-BICDR-1 caused an approximately two-fold increase in pericentrosomal dynein/dynactin fluorescent staining intensity (Figure 4B and E; Supplementary Figure S5C), indicating that BICDR-1 recruits the dynein/dynactin motor complex to the pericentrosomal region.


Pericentrosomal targeting of Rab6 secretory vesicles by Bicaudal-D-related protein 1 (BICDR-1) regulates neuritogenesis.

Schlager MA, Kapitein LC, Grigoriev I, Burzynski GM, Wulf PS, Keijzer N, de Graaff E, Fukuda M, Shepherd IT, Akhmanova A, Hoogenraad CC - EMBO J. (2010)

BICDR-1 interacts with the dynein/dynactin motor complex. (A) Immunoprecipitations from extracts of Hela cells transfected with the indicated constructs and probed for DHC, p150glued or dynein intermediate chain 74 (IC74). (B) Ratio of pericentrosomal versus cytoplasmic p150glued or IC74 fluorescence intensity in cells with and without GFP-BICDR-1 overexpression (average ±s.e.m.; p150glued control, n=29; p150glued+BICDR-1, n=30; IC74 control, n=32; IC74+BICDR-1, n=19 cells). (C) Percentage of cells, transfected with indicated siRNAs, with endogenous pericentrosomal BICDR-1 (average ±s.e.m.; control, n=3040; DHC siRNA, n=533; p150glued, n=792 cells). (D) Representative image of a Vero cell stained for endogenous BICDR-1 (green) and for p150glued (red). (E) Representative image of Hela cells with and without GFP-BICDR-1 (green) overexpression stained for p150glued (red). (F) Vero cells transfected with p150glued-specific siRNA (right) and untransfected (left). Stained for BICDR-1 (green) and p150glued (red). (D–F) Solid lines indicate the cell edge and dashed lines indicate the nucleus. The insets show magnifications of boxed areas. Scale bars, 10 μm. ***P<0.001.
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f4: BICDR-1 interacts with the dynein/dynactin motor complex. (A) Immunoprecipitations from extracts of Hela cells transfected with the indicated constructs and probed for DHC, p150glued or dynein intermediate chain 74 (IC74). (B) Ratio of pericentrosomal versus cytoplasmic p150glued or IC74 fluorescence intensity in cells with and without GFP-BICDR-1 overexpression (average ±s.e.m.; p150glued control, n=29; p150glued+BICDR-1, n=30; IC74 control, n=32; IC74+BICDR-1, n=19 cells). (C) Percentage of cells, transfected with indicated siRNAs, with endogenous pericentrosomal BICDR-1 (average ±s.e.m.; control, n=3040; DHC siRNA, n=533; p150glued, n=792 cells). (D) Representative image of a Vero cell stained for endogenous BICDR-1 (green) and for p150glued (red). (E) Representative image of Hela cells with and without GFP-BICDR-1 (green) overexpression stained for p150glued (red). (F) Vero cells transfected with p150glued-specific siRNA (right) and untransfected (left). Stained for BICDR-1 (green) and p150glued (red). (D–F) Solid lines indicate the cell edge and dashed lines indicate the nucleus. The insets show magnifications of boxed areas. Scale bars, 10 μm. ***P<0.001.
Mentions: Earlier studies have shown that members of BICD family interact with the microtubule minus-end-directed dynein/dynactin motor complex. Therefore, we investigated the possible interaction between BICDR-1 and dynein/dynactin. Co-immunoprecipitation experiments from Hela cells transfected with GFP-BICDR-1 showed that BICDR-1 precipitates the major dynein/dynactin subunits, whereas no binding is seen with control GFP (Figure 4A). Consistently, both dynactin (Figure 4D) and dynein (Supplementary Figure S5B) co-localize with endogenous BICDR-1 around the centrosome in Vero cells. Overexpression of GFP-BICDR-1 caused an approximately two-fold increase in pericentrosomal dynein/dynactin fluorescent staining intensity (Figure 4B and E; Supplementary Figure S5C), indicating that BICDR-1 recruits the dynein/dynactin motor complex to the pericentrosomal region.

Bottom Line: BICDR-1 expression is high during early neuronal development and strongly declines during neurite outgrowth.Later during development, BICDR-1 expression is strongly reduced, which permits anterograde secretory transport required for neurite outgrowth.These results indicate an important role for BICDR-1 as temporal regulator of secretory trafficking during the early phase of neuronal differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Erasmus Medical Center, Rotterdam, The Netherlands.

ABSTRACT
Membrane and secretory trafficking are essential for proper neuronal development. However, the molecular mechanisms that organize secretory trafficking are poorly understood. Here, we identify Bicaudal-D-related protein 1 (BICDR-1) as an effector of the small GTPase Rab6 and key component of the molecular machinery that controls secretory vesicle transport in developing neurons. BICDR-1 interacts with kinesin motor Kif1C, the dynein/dynactin retrograde motor complex, regulates the pericentrosomal localization of Rab6-positive secretory vesicles and is required for neural development in zebrafish. BICDR-1 expression is high during early neuronal development and strongly declines during neurite outgrowth. In young neurons, BICDR-1 accumulates Rab6 secretory vesicles around the centrosome, restricts anterograde secretory transport and inhibits neuritogenesis. Later during development, BICDR-1 expression is strongly reduced, which permits anterograde secretory transport required for neurite outgrowth. These results indicate an important role for BICDR-1 as temporal regulator of secretory trafficking during the early phase of neuronal differentiation.

Show MeSH
Related in: MedlinePlus