Limits...
Drosophila roadblock and Chlamydomonas LC7: a conserved family of dynein-associated proteins involved in axonal transport, flagellar motility, and mitosis.

Bowman AB, Patel-King RS, Benashski SE, McCaffery JM, Goldstein LS, King SM - J. Cell Biol. (1999)

Bottom Line: The gene identified by robl encodes a 97-amino acid polypeptide that is 57% identical (70% similar) to the 105-amino acid Chlamydomonas outer arm dynein-associated protein LC7, also reported here.Furthermore, we demonstrate that members of this family of proteins are associated with both flagellar outer arm dynein and Drosophila and rat brain cytoplasmic dynein.We propose that roadblock/LC7 family members may modulate specific dynein functions.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Division of Cellular and Molecular Medicine, Department of Pharmacology, University of California San Diego, La Jolla, California 92093-0683, USA.

ABSTRACT
Eukaryotic organisms utilize microtubule-dependent motors of the kinesin and dynein superfamilies to generate intracellular movement. To identify new genes involved in the regulation of axonal transport in Drosophila melanogaster, we undertook a screen based upon the sluggish larval phenotype of known motor mutants. One of the mutants identified in this screen, roadblock (robl), exhibits diverse defects in intracellular transport including axonal transport and mitosis. These defects include intra-axonal accumulations of cargoes, severe axonal degeneration, and aberrant chromosome segregation. The gene identified by robl encodes a 97-amino acid polypeptide that is 57% identical (70% similar) to the 105-amino acid Chlamydomonas outer arm dynein-associated protein LC7, also reported here. Both robl and LC7 have homology to several other genes from fruit fly, nematode, and mammals, but not Saccharomyces cerevisiae. Furthermore, we demonstrate that members of this family of proteins are associated with both flagellar outer arm dynein and Drosophila and rat brain cytoplasmic dynein. We propose that roadblock/LC7 family members may modulate specific dynein functions.

Show MeSH

Related in: MedlinePlus

Coimmunostaining of larval segmental nerves for (left column) SYT and (right column) ChAT revealed distal axonal accumulations of synaptic cargo in robl mutants. (A and B) In wild type, there is only a low background staining observed. (C and D) Segmental nerves from roblz hemizygotes have accumulations of SYT and ChAT. (E and F) In roblz hemizygotes the segmental nerves at the anterior, coming off of the ventral ganglion (located at the bottom left quadrant in images E and F), show a decreased frequency of SYT accumulations but an increased frequency of ChAT accumulations when compared with more posterior regions. (G and H) The segmental nerves located at the posterior show an increased frequency of SYT accumulations with a corresponding decreased frequency of ChAT accumulations. (I and J) Segmental nerves from robl  homozygotes (roblk/roblk) show abundant accumulations of SYT and ChAT.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2199740&req=5

Figure 5: Coimmunostaining of larval segmental nerves for (left column) SYT and (right column) ChAT revealed distal axonal accumulations of synaptic cargo in robl mutants. (A and B) In wild type, there is only a low background staining observed. (C and D) Segmental nerves from roblz hemizygotes have accumulations of SYT and ChAT. (E and F) In roblz hemizygotes the segmental nerves at the anterior, coming off of the ventral ganglion (located at the bottom left quadrant in images E and F), show a decreased frequency of SYT accumulations but an increased frequency of ChAT accumulations when compared with more posterior regions. (G and H) The segmental nerves located at the posterior show an increased frequency of SYT accumulations with a corresponding decreased frequency of ChAT accumulations. (I and J) Segmental nerves from robl homozygotes (roblk/roblk) show abundant accumulations of SYT and ChAT.

Mentions: Mutations in robl cause phenotypes similar to other axonal transport mutants in Drosophila. Previous analysis of kinesin heavy chain (khc) and kinesin light chain mutants demonstrated massive accumulations of axonal cargo and motors distributed randomly along the entire length of the larval segmental nerves. These accumulations were shown to be massive local axonal swellings that fill with organelles and vesicles (Hurd and Saxton 1996; Gindhart et al. 1998). The accumulation phenotype correlates with the other common axonal transport phenotypes in Drosophila, tail flipping and posterior paralysis. It was proposed that these mutants disrupt the processive movement of their cargo within the axon, causing the axons to swell, filling with transported axonal material. Immunostaining of roblz/ hemizygous and robl homozygous larvae reveals frequent accumulations of synaptotagmin (SYT) (Fig. 5C and Fig. I) and choline acetyltransferase (ChAT) (Fig. 5D and Fig. J) in the larval segmental nerves. In contrast, SYT (Fig. 5 A) and ChAT (Fig. 5 B) show only a low background level staining in wild-type segmental nerves. Additionally, axonal transport motors (of the kinesin I and kinesin II family), cysteine string protein, and a marker for endocytic traffic are also observed to accumulate in the axons of robl mutants (data not shown). Thus, robl mutants have a gross phenotype similar to that previously described for axonal transport mutants in Drosophila; a progressive larval posterior paralysis, tail flipping, and segmental nerve axonal cargo accumulation.


Drosophila roadblock and Chlamydomonas LC7: a conserved family of dynein-associated proteins involved in axonal transport, flagellar motility, and mitosis.

Bowman AB, Patel-King RS, Benashski SE, McCaffery JM, Goldstein LS, King SM - J. Cell Biol. (1999)

Coimmunostaining of larval segmental nerves for (left column) SYT and (right column) ChAT revealed distal axonal accumulations of synaptic cargo in robl mutants. (A and B) In wild type, there is only a low background staining observed. (C and D) Segmental nerves from roblz hemizygotes have accumulations of SYT and ChAT. (E and F) In roblz hemizygotes the segmental nerves at the anterior, coming off of the ventral ganglion (located at the bottom left quadrant in images E and F), show a decreased frequency of SYT accumulations but an increased frequency of ChAT accumulations when compared with more posterior regions. (G and H) The segmental nerves located at the posterior show an increased frequency of SYT accumulations with a corresponding decreased frequency of ChAT accumulations. (I and J) Segmental nerves from robl  homozygotes (roblk/roblk) show abundant accumulations of SYT and ChAT.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199740&req=5

Figure 5: Coimmunostaining of larval segmental nerves for (left column) SYT and (right column) ChAT revealed distal axonal accumulations of synaptic cargo in robl mutants. (A and B) In wild type, there is only a low background staining observed. (C and D) Segmental nerves from roblz hemizygotes have accumulations of SYT and ChAT. (E and F) In roblz hemizygotes the segmental nerves at the anterior, coming off of the ventral ganglion (located at the bottom left quadrant in images E and F), show a decreased frequency of SYT accumulations but an increased frequency of ChAT accumulations when compared with more posterior regions. (G and H) The segmental nerves located at the posterior show an increased frequency of SYT accumulations with a corresponding decreased frequency of ChAT accumulations. (I and J) Segmental nerves from robl homozygotes (roblk/roblk) show abundant accumulations of SYT and ChAT.
Mentions: Mutations in robl cause phenotypes similar to other axonal transport mutants in Drosophila. Previous analysis of kinesin heavy chain (khc) and kinesin light chain mutants demonstrated massive accumulations of axonal cargo and motors distributed randomly along the entire length of the larval segmental nerves. These accumulations were shown to be massive local axonal swellings that fill with organelles and vesicles (Hurd and Saxton 1996; Gindhart et al. 1998). The accumulation phenotype correlates with the other common axonal transport phenotypes in Drosophila, tail flipping and posterior paralysis. It was proposed that these mutants disrupt the processive movement of their cargo within the axon, causing the axons to swell, filling with transported axonal material. Immunostaining of roblz/ hemizygous and robl homozygous larvae reveals frequent accumulations of synaptotagmin (SYT) (Fig. 5C and Fig. I) and choline acetyltransferase (ChAT) (Fig. 5D and Fig. J) in the larval segmental nerves. In contrast, SYT (Fig. 5 A) and ChAT (Fig. 5 B) show only a low background level staining in wild-type segmental nerves. Additionally, axonal transport motors (of the kinesin I and kinesin II family), cysteine string protein, and a marker for endocytic traffic are also observed to accumulate in the axons of robl mutants (data not shown). Thus, robl mutants have a gross phenotype similar to that previously described for axonal transport mutants in Drosophila; a progressive larval posterior paralysis, tail flipping, and segmental nerve axonal cargo accumulation.

Bottom Line: The gene identified by robl encodes a 97-amino acid polypeptide that is 57% identical (70% similar) to the 105-amino acid Chlamydomonas outer arm dynein-associated protein LC7, also reported here.Furthermore, we demonstrate that members of this family of proteins are associated with both flagellar outer arm dynein and Drosophila and rat brain cytoplasmic dynein.We propose that roadblock/LC7 family members may modulate specific dynein functions.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Division of Cellular and Molecular Medicine, Department of Pharmacology, University of California San Diego, La Jolla, California 92093-0683, USA.

ABSTRACT
Eukaryotic organisms utilize microtubule-dependent motors of the kinesin and dynein superfamilies to generate intracellular movement. To identify new genes involved in the regulation of axonal transport in Drosophila melanogaster, we undertook a screen based upon the sluggish larval phenotype of known motor mutants. One of the mutants identified in this screen, roadblock (robl), exhibits diverse defects in intracellular transport including axonal transport and mitosis. These defects include intra-axonal accumulations of cargoes, severe axonal degeneration, and aberrant chromosome segregation. The gene identified by robl encodes a 97-amino acid polypeptide that is 57% identical (70% similar) to the 105-amino acid Chlamydomonas outer arm dynein-associated protein LC7, also reported here. Both robl and LC7 have homology to several other genes from fruit fly, nematode, and mammals, but not Saccharomyces cerevisiae. Furthermore, we demonstrate that members of this family of proteins are associated with both flagellar outer arm dynein and Drosophila and rat brain cytoplasmic dynein. We propose that roadblock/LC7 family members may modulate specific dynein functions.

Show MeSH
Related in: MedlinePlus