Limits...
The Drosophila gene abnormal spindle encodes a novel microtubule-associated protein that associates with the polar regions of the mitotic spindle.

Saunders RD, Avides MC, Howard T, Gonzalez C, Glover DM - J. Cell Biol. (1997)

Bottom Line: Wild-type Asp protein copurifies with microtubules and is not released by salt concentrations known to dissociate most other microtubule-associated proteins.The central 579-amino acid segment of the molecule contains one short motif homologous to sequences in a number of actin bundling proteins and a second motif present at the calmodulin binding sites of several proteins.Immunofluorescence studies show that the wild-type Asp protein is localized to the polar regions of the spindle immediately surrounding the centrosome.

View Article: PubMed Central - PubMed

Affiliation: Cancer Research Campaign, Cell Cycle Genetics Group, Department of Anatomy and Physiology, University of Dundee, Dundee DD1 4HN, Scotland.

ABSTRACT
abnormal spindle, a gene required for normal spindle structure and function in Drosophila melanogaster, lies immediately adjacent the gene tolloid at 96A/B. It encodes a 220-kD polypeptide with a predicted pI of 10.8. The recessive mutant allele asp1 directs the synthesis of a COOH terminally truncated or internally deleted peptide of approximately 124 kD. Wild-type Asp protein copurifies with microtubules and is not released by salt concentrations known to dissociate most other microtubule-associated proteins. The bacterially expressed NH2-terminal 512-amino acid peptide, which has a number of potential phosphorylation sites for p34(cdc2) and MAP kinases, strongly binds to microtubules. The central 579-amino acid segment of the molecule contains one short motif homologous to sequences in a number of actin bundling proteins and a second motif present at the calmodulin binding sites of several proteins. Immunofluorescence studies show that the wild-type Asp protein is localized to the polar regions of the spindle immediately surrounding the centrosome. These findings are discussed in relation to the known spindle abnormalities in asp mutants.

Show MeSH

Related in: MedlinePlus

A cytogenetic and  molecular map of the asp region. A is a representation of  divisions 95 and 96 of the  polytene map of chromosome arm 3R. Subdivisions  A and B are indicated. The  rearrangements used by  Gonzalez et al. (1989) to define the asp region, In(3R)Ubx7LLatsR and Df(3R)A117PB197D are indicated  above the chromosomes. B  shows the proximal section  of the chromosome walk  linked to the cytological map.  B, R, and S represent BamHI,  EcoRI, and SalI restriction  sites, respectively. Horizontal lines below the restriction  map indicate phage lambda  and cosmid clones isolated  from genomic libraries. Boxed  fragments indicate fragments  mapped by in situ hybridization. One contains the distal  breakpoint of In(3R)Ubx7LLatsR (ats), while the other is  an EcoRI fragment recovered by microcloning. C is an  expansion of the proximal  section of the chromosome  walk. The distal breakpoints  of In(3R)Ubx7LLatsR and  Df(3R)Hdγ1 are indicated.  Uncertainties about the cytological limits of the deficiencies are indicated by the thinner lines. The limits of the  deletion associated with the  P element insertion tld allele  tld68–62 are indicated by the  bar. D illustrates the transcription units of tld and asp  as well as the two fragments  used in P element mediated rescue experiments. pMBO1367 contains an 18-kb SalI fragment containing both tld and asp. pMBO1366  contains a 14-kb SalI fragment containing tld but only the 5′ portion of asp.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2139842&req=5

Figure 1: A cytogenetic and molecular map of the asp region. A is a representation of divisions 95 and 96 of the polytene map of chromosome arm 3R. Subdivisions A and B are indicated. The rearrangements used by Gonzalez et al. (1989) to define the asp region, In(3R)Ubx7LLatsR and Df(3R)A117PB197D are indicated above the chromosomes. B shows the proximal section of the chromosome walk linked to the cytological map. B, R, and S represent BamHI, EcoRI, and SalI restriction sites, respectively. Horizontal lines below the restriction map indicate phage lambda and cosmid clones isolated from genomic libraries. Boxed fragments indicate fragments mapped by in situ hybridization. One contains the distal breakpoint of In(3R)Ubx7LLatsR (ats), while the other is an EcoRI fragment recovered by microcloning. C is an expansion of the proximal section of the chromosome walk. The distal breakpoints of In(3R)Ubx7LLatsR and Df(3R)Hdγ1 are indicated. Uncertainties about the cytological limits of the deficiencies are indicated by the thinner lines. The limits of the deletion associated with the P element insertion tld allele tld68–62 are indicated by the bar. D illustrates the transcription units of tld and asp as well as the two fragments used in P element mediated rescue experiments. pMBO1367 contains an 18-kb SalI fragment containing both tld and asp. pMBO1366 contains a 14-kb SalI fragment containing tld but only the 5′ portion of asp.

Mentions: Previous genetic mapping studies (Gonzalez et al., 1989) mapped asp to the cytological interval 96A21-96B10 between the distal breakpoint of In(3R)Ubx7LLatsR and the breakpoint of T(Y;3)B197 (Fig. 1). At the onset of this work there were no obvious molecular entry points into this region, and so we chose the technique of chromosome microdissection and microcloning to achieve this end. Two chromosomal segments corresponding to the asp interval were microdissected and DNA fragments extracted for cloning in the bacteriophage λ insertion vector λ1149. The chromosomal origins of these clones were confirmed by in situ hybridization, before using them to screen phage λ and cosmid genomic libraries (see Materials and Methods). A resulting chromosome walk of 130 kb was carried out and clones correlated with the polytene chromosome map by in situ hybridization. The distal breakpoint of In(3R)Ubx7LLatsR, which does not uncover asp and defines the proximal boundary of the region known to contain asp, was mapped to a 5.8-kb BamHI fragment at the extreme proximal end of the walk. The distal end of the chromosome walk maps to the interband between 96A21-25 and 96B1-10. Subsequently, we found that a cytologically invisible deficiency Df(3R)Hdγ1 uncovers not only the nearby zygotic embryonic lethal tolloid (tld), as reported by Shimell et al. (1991), but also asp.


The Drosophila gene abnormal spindle encodes a novel microtubule-associated protein that associates with the polar regions of the mitotic spindle.

Saunders RD, Avides MC, Howard T, Gonzalez C, Glover DM - J. Cell Biol. (1997)

A cytogenetic and  molecular map of the asp region. A is a representation of  divisions 95 and 96 of the  polytene map of chromosome arm 3R. Subdivisions  A and B are indicated. The  rearrangements used by  Gonzalez et al. (1989) to define the asp region, In(3R)Ubx7LLatsR and Df(3R)A117PB197D are indicated  above the chromosomes. B  shows the proximal section  of the chromosome walk  linked to the cytological map.  B, R, and S represent BamHI,  EcoRI, and SalI restriction  sites, respectively. Horizontal lines below the restriction  map indicate phage lambda  and cosmid clones isolated  from genomic libraries. Boxed  fragments indicate fragments  mapped by in situ hybridization. One contains the distal  breakpoint of In(3R)Ubx7LLatsR (ats), while the other is  an EcoRI fragment recovered by microcloning. C is an  expansion of the proximal  section of the chromosome  walk. The distal breakpoints  of In(3R)Ubx7LLatsR and  Df(3R)Hdγ1 are indicated.  Uncertainties about the cytological limits of the deficiencies are indicated by the thinner lines. The limits of the  deletion associated with the  P element insertion tld allele  tld68–62 are indicated by the  bar. D illustrates the transcription units of tld and asp  as well as the two fragments  used in P element mediated rescue experiments. pMBO1367 contains an 18-kb SalI fragment containing both tld and asp. pMBO1366  contains a 14-kb SalI fragment containing tld but only the 5′ portion of asp.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139842&req=5

Figure 1: A cytogenetic and molecular map of the asp region. A is a representation of divisions 95 and 96 of the polytene map of chromosome arm 3R. Subdivisions A and B are indicated. The rearrangements used by Gonzalez et al. (1989) to define the asp region, In(3R)Ubx7LLatsR and Df(3R)A117PB197D are indicated above the chromosomes. B shows the proximal section of the chromosome walk linked to the cytological map. B, R, and S represent BamHI, EcoRI, and SalI restriction sites, respectively. Horizontal lines below the restriction map indicate phage lambda and cosmid clones isolated from genomic libraries. Boxed fragments indicate fragments mapped by in situ hybridization. One contains the distal breakpoint of In(3R)Ubx7LLatsR (ats), while the other is an EcoRI fragment recovered by microcloning. C is an expansion of the proximal section of the chromosome walk. The distal breakpoints of In(3R)Ubx7LLatsR and Df(3R)Hdγ1 are indicated. Uncertainties about the cytological limits of the deficiencies are indicated by the thinner lines. The limits of the deletion associated with the P element insertion tld allele tld68–62 are indicated by the bar. D illustrates the transcription units of tld and asp as well as the two fragments used in P element mediated rescue experiments. pMBO1367 contains an 18-kb SalI fragment containing both tld and asp. pMBO1366 contains a 14-kb SalI fragment containing tld but only the 5′ portion of asp.
Mentions: Previous genetic mapping studies (Gonzalez et al., 1989) mapped asp to the cytological interval 96A21-96B10 between the distal breakpoint of In(3R)Ubx7LLatsR and the breakpoint of T(Y;3)B197 (Fig. 1). At the onset of this work there were no obvious molecular entry points into this region, and so we chose the technique of chromosome microdissection and microcloning to achieve this end. Two chromosomal segments corresponding to the asp interval were microdissected and DNA fragments extracted for cloning in the bacteriophage λ insertion vector λ1149. The chromosomal origins of these clones were confirmed by in situ hybridization, before using them to screen phage λ and cosmid genomic libraries (see Materials and Methods). A resulting chromosome walk of 130 kb was carried out and clones correlated with the polytene chromosome map by in situ hybridization. The distal breakpoint of In(3R)Ubx7LLatsR, which does not uncover asp and defines the proximal boundary of the region known to contain asp, was mapped to a 5.8-kb BamHI fragment at the extreme proximal end of the walk. The distal end of the chromosome walk maps to the interband between 96A21-25 and 96B1-10. Subsequently, we found that a cytologically invisible deficiency Df(3R)Hdγ1 uncovers not only the nearby zygotic embryonic lethal tolloid (tld), as reported by Shimell et al. (1991), but also asp.

Bottom Line: Wild-type Asp protein copurifies with microtubules and is not released by salt concentrations known to dissociate most other microtubule-associated proteins.The central 579-amino acid segment of the molecule contains one short motif homologous to sequences in a number of actin bundling proteins and a second motif present at the calmodulin binding sites of several proteins.Immunofluorescence studies show that the wild-type Asp protein is localized to the polar regions of the spindle immediately surrounding the centrosome.

View Article: PubMed Central - PubMed

Affiliation: Cancer Research Campaign, Cell Cycle Genetics Group, Department of Anatomy and Physiology, University of Dundee, Dundee DD1 4HN, Scotland.

ABSTRACT
abnormal spindle, a gene required for normal spindle structure and function in Drosophila melanogaster, lies immediately adjacent the gene tolloid at 96A/B. It encodes a 220-kD polypeptide with a predicted pI of 10.8. The recessive mutant allele asp1 directs the synthesis of a COOH terminally truncated or internally deleted peptide of approximately 124 kD. Wild-type Asp protein copurifies with microtubules and is not released by salt concentrations known to dissociate most other microtubule-associated proteins. The bacterially expressed NH2-terminal 512-amino acid peptide, which has a number of potential phosphorylation sites for p34(cdc2) and MAP kinases, strongly binds to microtubules. The central 579-amino acid segment of the molecule contains one short motif homologous to sequences in a number of actin bundling proteins and a second motif present at the calmodulin binding sites of several proteins. Immunofluorescence studies show that the wild-type Asp protein is localized to the polar regions of the spindle immediately surrounding the centrosome. These findings are discussed in relation to the known spindle abnormalities in asp mutants.

Show MeSH
Related in: MedlinePlus