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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 is mainly expressed in kidney and neural tissues during development. (A) Schematic structure of BICD2 and BICDR proteins (CC, coiled-coil region). Antibodies were raised against GST-tagged BICDR-1 fusion proteins containing amino acids 1–146 (antibody #12) and 161–387 (antibody #13). (B) Expression of BICDR-1 mRNA in various rat tissues. (C) BICDR-1 antibodies #12 and #13 specifically recognize GFP-tagged BICDR-1 not the homologous proteins BICDR-2, BICD1 or BICD2. (D) Western blot analysis of BICDR-1, Rab6A and Rab6B in various mouse tissues. (E) Developmental expression patterns of BICDR-1, Rab6A, Rab6B, p150glued and BICD2 in E10.5 (whole embryo), E13.5, E16, E18 and P1 (head only) mouse. (F–H) Lateral view of E10 mouse embryo hybridized with a BICDR-1-specific ribo-probe. Indicated structures: dorsal root ganglion (DRG); mesencephalon (Mes); rhombencephalon (Rhom); telencephalon (Tel). Scale bar in (F) 1 mm in (G–H) 100 μm. (I–L) Transverse cryosections (10 μm) of an E13.5 mouse kidney, hybridized with a BICDR-1 sense (I, K) or anti-sense (J, L) ribo-probe. Indicated structures: cortical region of metanephros (Ctx); medullary region of metanephros (Med); and metanephric tubule (Tub). Scale bars, 100 μm. (M–R) Transverse cryosections (10 μm) of an E13.5 mouse head, hybridized with a BICDR-1 sense (M, O, Q) or anti-sense (N, P, R) ribo-probe. Indicated structures: fourth ventricle (4v); third ventricle (3v); choroid plexus (Chor); myencephalon (My); metencephalon (Met); diencephalon (Di); trigeminal (V) ganglion (Trig); olfactory epithelium (Olf ep); ventricular zone (VZ); intermediate zone (IZ); neural layer of retina (NLR). Scale bars, 100 μm.
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f1: BICDR-1 is mainly expressed in kidney and neural tissues during development. (A) Schematic structure of BICD2 and BICDR proteins (CC, coiled-coil region). Antibodies were raised against GST-tagged BICDR-1 fusion proteins containing amino acids 1–146 (antibody #12) and 161–387 (antibody #13). (B) Expression of BICDR-1 mRNA in various rat tissues. (C) BICDR-1 antibodies #12 and #13 specifically recognize GFP-tagged BICDR-1 not the homologous proteins BICDR-2, BICD1 or BICD2. (D) Western blot analysis of BICDR-1, Rab6A and Rab6B in various mouse tissues. (E) Developmental expression patterns of BICDR-1, Rab6A, Rab6B, p150glued and BICD2 in E10.5 (whole embryo), E13.5, E16, E18 and P1 (head only) mouse. (F–H) Lateral view of E10 mouse embryo hybridized with a BICDR-1-specific ribo-probe. Indicated structures: dorsal root ganglion (DRG); mesencephalon (Mes); rhombencephalon (Rhom); telencephalon (Tel). Scale bar in (F) 1 mm in (G–H) 100 μm. (I–L) Transverse cryosections (10 μm) of an E13.5 mouse kidney, hybridized with a BICDR-1 sense (I, K) or anti-sense (J, L) ribo-probe. Indicated structures: cortical region of metanephros (Ctx); medullary region of metanephros (Med); and metanephric tubule (Tub). Scale bars, 100 μm. (M–R) Transverse cryosections (10 μm) of an E13.5 mouse head, hybridized with a BICDR-1 sense (M, O, Q) or anti-sense (N, P, R) ribo-probe. Indicated structures: fourth ventricle (4v); third ventricle (3v); choroid plexus (Chor); myencephalon (My); metencephalon (Met); diencephalon (Di); trigeminal (V) ganglion (Trig); olfactory epithelium (Olf ep); ventricular zone (VZ); intermediate zone (IZ); neural layer of retina (NLR). Scale bars, 100 μm.

Mentions: In a sequence homology search for new mammalian BICD-like proteins, we found two previously uncharacterized BICD homologues: hypothetical coiled-coil domain containing protein 64 (Ccdc64) and 64B (Ccdc64B). We renamed these proteins BICDR-1 and BICD-related protein 2 (BICDR-2), respectively. BICDR-1 and BICDR-2 genes are conserved in vertebrates, whereas BICDR-1 is also present in flies (Supplementary Figure S1A and B). In contrast to the three coiled-coil segments in BICD, BICDR proteins are shorter and only contain two predicted coiled-coil regions (Figure 1A). The highest degree of similarity to BICD is found in the cargo-binding domain at the C-terminus of BICDR (64% similarity; Supplementary Figure S1C). Northern blot analysis showed that BICDR-1 is predominantly expressed in kidney and brain, whereas some expression can be seen in testis and heart (Figure 1B). In contrast, BICDR-2 could not be detected on northern blots or tissue dot blots (data not shown). Indeed, expressed sequence tag profiles show that the BICDR-2 gene is expressed at low levels and only detected in a few tissues such as stomach, oesophagus and ovary. In this study, we decided to focus on BICDR-1 and generated rabbit polyclonal antibodies against both the N-terminal (#12) and central (#13) region of the protein (Figure 1A). Both antibodies reacted only to BICDR-1, and not to control proteins BICD1, BICD2 or BICDR-2 (Figure 1C). In agreement with mRNA expression data, western blot analysis of various adult mouse tissues showed that BICDR-1 expression is high in kidney and moderate in testis and brain (Figure 1D).


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 is mainly expressed in kidney and neural tissues during development. (A) Schematic structure of BICD2 and BICDR proteins (CC, coiled-coil region). Antibodies were raised against GST-tagged BICDR-1 fusion proteins containing amino acids 1–146 (antibody #12) and 161–387 (antibody #13). (B) Expression of BICDR-1 mRNA in various rat tissues. (C) BICDR-1 antibodies #12 and #13 specifically recognize GFP-tagged BICDR-1 not the homologous proteins BICDR-2, BICD1 or BICD2. (D) Western blot analysis of BICDR-1, Rab6A and Rab6B in various mouse tissues. (E) Developmental expression patterns of BICDR-1, Rab6A, Rab6B, p150glued and BICD2 in E10.5 (whole embryo), E13.5, E16, E18 and P1 (head only) mouse. (F–H) Lateral view of E10 mouse embryo hybridized with a BICDR-1-specific ribo-probe. Indicated structures: dorsal root ganglion (DRG); mesencephalon (Mes); rhombencephalon (Rhom); telencephalon (Tel). Scale bar in (F) 1 mm in (G–H) 100 μm. (I–L) Transverse cryosections (10 μm) of an E13.5 mouse kidney, hybridized with a BICDR-1 sense (I, K) or anti-sense (J, L) ribo-probe. Indicated structures: cortical region of metanephros (Ctx); medullary region of metanephros (Med); and metanephric tubule (Tub). Scale bars, 100 μm. (M–R) Transverse cryosections (10 μm) of an E13.5 mouse head, hybridized with a BICDR-1 sense (M, O, Q) or anti-sense (N, P, R) ribo-probe. Indicated structures: fourth ventricle (4v); third ventricle (3v); choroid plexus (Chor); myencephalon (My); metencephalon (Met); diencephalon (Di); trigeminal (V) ganglion (Trig); olfactory epithelium (Olf ep); ventricular zone (VZ); intermediate zone (IZ); neural layer of retina (NLR). Scale bars, 100 μm.
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Related In: Results  -  Collection

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Show All Figures
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f1: BICDR-1 is mainly expressed in kidney and neural tissues during development. (A) Schematic structure of BICD2 and BICDR proteins (CC, coiled-coil region). Antibodies were raised against GST-tagged BICDR-1 fusion proteins containing amino acids 1–146 (antibody #12) and 161–387 (antibody #13). (B) Expression of BICDR-1 mRNA in various rat tissues. (C) BICDR-1 antibodies #12 and #13 specifically recognize GFP-tagged BICDR-1 not the homologous proteins BICDR-2, BICD1 or BICD2. (D) Western blot analysis of BICDR-1, Rab6A and Rab6B in various mouse tissues. (E) Developmental expression patterns of BICDR-1, Rab6A, Rab6B, p150glued and BICD2 in E10.5 (whole embryo), E13.5, E16, E18 and P1 (head only) mouse. (F–H) Lateral view of E10 mouse embryo hybridized with a BICDR-1-specific ribo-probe. Indicated structures: dorsal root ganglion (DRG); mesencephalon (Mes); rhombencephalon (Rhom); telencephalon (Tel). Scale bar in (F) 1 mm in (G–H) 100 μm. (I–L) Transverse cryosections (10 μm) of an E13.5 mouse kidney, hybridized with a BICDR-1 sense (I, K) or anti-sense (J, L) ribo-probe. Indicated structures: cortical region of metanephros (Ctx); medullary region of metanephros (Med); and metanephric tubule (Tub). Scale bars, 100 μm. (M–R) Transverse cryosections (10 μm) of an E13.5 mouse head, hybridized with a BICDR-1 sense (M, O, Q) or anti-sense (N, P, R) ribo-probe. Indicated structures: fourth ventricle (4v); third ventricle (3v); choroid plexus (Chor); myencephalon (My); metencephalon (Met); diencephalon (Di); trigeminal (V) ganglion (Trig); olfactory epithelium (Olf ep); ventricular zone (VZ); intermediate zone (IZ); neural layer of retina (NLR). Scale bars, 100 μm.
Mentions: In a sequence homology search for new mammalian BICD-like proteins, we found two previously uncharacterized BICD homologues: hypothetical coiled-coil domain containing protein 64 (Ccdc64) and 64B (Ccdc64B). We renamed these proteins BICDR-1 and BICD-related protein 2 (BICDR-2), respectively. BICDR-1 and BICDR-2 genes are conserved in vertebrates, whereas BICDR-1 is also present in flies (Supplementary Figure S1A and B). In contrast to the three coiled-coil segments in BICD, BICDR proteins are shorter and only contain two predicted coiled-coil regions (Figure 1A). The highest degree of similarity to BICD is found in the cargo-binding domain at the C-terminus of BICDR (64% similarity; Supplementary Figure S1C). Northern blot analysis showed that BICDR-1 is predominantly expressed in kidney and brain, whereas some expression can be seen in testis and heart (Figure 1B). In contrast, BICDR-2 could not be detected on northern blots or tissue dot blots (data not shown). Indeed, expressed sequence tag profiles show that the BICDR-2 gene is expressed at low levels and only detected in a few tissues such as stomach, oesophagus and ovary. In this study, we decided to focus on BICDR-1 and generated rabbit polyclonal antibodies against both the N-terminal (#12) and central (#13) region of the protein (Figure 1A). Both antibodies reacted only to BICDR-1, and not to control proteins BICD1, BICD2 or BICDR-2 (Figure 1C). In agreement with mRNA expression data, western blot analysis of various adult mouse tissues showed that BICDR-1 expression is high in kidney and moderate in testis and brain (Figure 1D).

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