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Gene organization, evolution and expression of the microtubule-associated protein ASAP (MAP9).

Venoux M, Delmouly K, Milhavet O, Vidal-Eychenié S, Giorgi D, Rouquier S - BMC Genomics (2008)

Bottom Line: The human gene is strongly expressed in brain and testis as a 2.6 Kb transcript encoding a approximately110 KDa protein.The protein contains MAP, MIT-like and THY domains in the C-terminal part indicative of microtubule interaction, while the N-terminal part is more divergent.It may have a role in spermatogenesis and also represents a potential new target for antitumoral drugs.

View Article: PubMed Central - HTML - PubMed

Affiliation: Groupe Microtubules et Cycle Cellulaire, Institut de Génétique Humaine, CNRS UPR 1142, rue de cardonille, 34396 Montpellier cédex 5, France. Magali.Venoux@igh.cnrs.fr

ABSTRACT

Background: ASAP is a newly characterized microtubule-associated protein (MAP) essential for proper cell-cycling. We have previously shown that expression deregulation of human ASAP results in profound defects in mitotic spindle formation and mitotic progression leading to aneuploidy, cytokinesis defects and/or cell death. In the present work we analyze the structure and evolution of the ASAP gene, as well as the domain composition of the encoded protein. Mouse and Xenopus cDNAs were cloned, the tissue expression characterized and the overexpression profile analyzed.

Results: Bona fide ASAP orthologs are found in vertebrates with more distantly related potential orthologs in invertebrates. This single-copy gene is conserved in mammals where it maps to syntenic chromosomal regions, but is also clearly identified in bird, fish and frog. The human gene is strongly expressed in brain and testis as a 2.6 Kb transcript encoding a approximately110 KDa protein. The protein contains MAP, MIT-like and THY domains in the C-terminal part indicative of microtubule interaction, while the N-terminal part is more divergent. ASAP is composed of approximately 42% alpha helical structures, and two main coiled-coil regions have been identified. Different sequence features may suggest a role in DNA damage response. As with human ASAP, the mouse and Xenopus proteins localize to the microtubule network in interphase and to the mitotic spindle during mitosis. Overexpression of the mouse protein induces mitotic defects similar to those observed in human. In situ hybridization in testis localized ASAP to the germ cells, whereas in culture neurons ASAP localized to the cell body and growing neurites.

Conclusion: The conservation of ASAP indicated in our results reflects an essential function in vertebrates. We have cloned the ASAP orthologs in mouse and Xenopus, two valuable models to study the function of ASAP. Tissue expression of ASAP revealed a high expression in brain and testis, two tissues rich in microtubules. ASAP associates to the mitotic spindle and cytoplasmic microtubules, and represents a key factor of mitosis with possible involvement in other cell cycle processes. It may have a role in spermatogenesis and also represents a potential new target for antitumoral drugs. Possible involvement in neuron dynamics also highlights ASAP as a candidate target in neurodegenerative diseases.

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Phylogenetic analysis and pairwise comparison of ASAP from different species. (A) Phylogenetic tree of the ASAP protein from fish, frog, mouse, dog and human. The predicted protein sequences were compared using the ClustalW software to generate a rooted phylogenetic tree using the bootstrap method. Branch length and bootstrap numbers are indicated. (B) Pairwise comparison (%) of ASAP proteins between the species depicted in A. The values above the diagonal show protein sequence identities, those under the diagonal show protein sequence similarities. These values were estimated using the Gap program of the GCG package. (C) Clustal alignment of the phylogenetic analysis performed in (A) plus a partial chicken sequence, visualized using Jalview and Clustal X colour codes [63]. On top of the aligment, colour bars indicate conserved domains; blue, MIT-like; green, MAP; red, NLS. Phosphoserines 132 (SQ motif) and 625 are squared. Canonical (+) and (-) residues of the MIT domain are squared.
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Figure 2: Phylogenetic analysis and pairwise comparison of ASAP from different species. (A) Phylogenetic tree of the ASAP protein from fish, frog, mouse, dog and human. The predicted protein sequences were compared using the ClustalW software to generate a rooted phylogenetic tree using the bootstrap method. Branch length and bootstrap numbers are indicated. (B) Pairwise comparison (%) of ASAP proteins between the species depicted in A. The values above the diagonal show protein sequence identities, those under the diagonal show protein sequence similarities. These values were estimated using the Gap program of the GCG package. (C) Clustal alignment of the phylogenetic analysis performed in (A) plus a partial chicken sequence, visualized using Jalview and Clustal X colour codes [63]. On top of the aligment, colour bars indicate conserved domains; blue, MIT-like; green, MAP; red, NLS. Phosphoserines 132 (SQ motif) and 625 are squared. Canonical (+) and (-) residues of the MIT domain are squared.

Mentions: Most vertebrate orthologs are contained in chromosomal regions that are syntenic to human (not shown), i.e., on chromosome 3F1 in mouse (this region is syntenic with human chromosome 4q32.1 as described in , chromosome 3 in chimpanzee (at 174 Mb), chromosome 15 in dog (at 55.9 Mb) and chromosome 4 in chicken (at 21.2 Mb). The X. laevis cDNA (645 amino-acids) was assembled using clones AW764964 and AW764609 (Table 1). The zebrafish gene is located on chromosome 1 at 26.02 Mb and has a similar intron-exon structure to that of human (not shown). Using the ASAP protein sequences from selected species we performed a phylogenetic comparison using Clustalw with a bootstrap method. As shown in Fig. 2, the human and dog proteins are very similar (>78% PSI) whereas mouse ASAP is more divergent. Although frog and fish ASAP are more distant than those from the other species, they do not constitute a separate family, since they have a similar topology and share >32–38% PSI and >44–48% sequence similarity with the human protein. Pairwise sequence comparisons confirmed these results (Fig. 2B).


Gene organization, evolution and expression of the microtubule-associated protein ASAP (MAP9).

Venoux M, Delmouly K, Milhavet O, Vidal-Eychenié S, Giorgi D, Rouquier S - BMC Genomics (2008)

Phylogenetic analysis and pairwise comparison of ASAP from different species. (A) Phylogenetic tree of the ASAP protein from fish, frog, mouse, dog and human. The predicted protein sequences were compared using the ClustalW software to generate a rooted phylogenetic tree using the bootstrap method. Branch length and bootstrap numbers are indicated. (B) Pairwise comparison (%) of ASAP proteins between the species depicted in A. The values above the diagonal show protein sequence identities, those under the diagonal show protein sequence similarities. These values were estimated using the Gap program of the GCG package. (C) Clustal alignment of the phylogenetic analysis performed in (A) plus a partial chicken sequence, visualized using Jalview and Clustal X colour codes [63]. On top of the aligment, colour bars indicate conserved domains; blue, MIT-like; green, MAP; red, NLS. Phosphoserines 132 (SQ motif) and 625 are squared. Canonical (+) and (-) residues of the MIT domain are squared.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2551623&req=5

Figure 2: Phylogenetic analysis and pairwise comparison of ASAP from different species. (A) Phylogenetic tree of the ASAP protein from fish, frog, mouse, dog and human. The predicted protein sequences were compared using the ClustalW software to generate a rooted phylogenetic tree using the bootstrap method. Branch length and bootstrap numbers are indicated. (B) Pairwise comparison (%) of ASAP proteins between the species depicted in A. The values above the diagonal show protein sequence identities, those under the diagonal show protein sequence similarities. These values were estimated using the Gap program of the GCG package. (C) Clustal alignment of the phylogenetic analysis performed in (A) plus a partial chicken sequence, visualized using Jalview and Clustal X colour codes [63]. On top of the aligment, colour bars indicate conserved domains; blue, MIT-like; green, MAP; red, NLS. Phosphoserines 132 (SQ motif) and 625 are squared. Canonical (+) and (-) residues of the MIT domain are squared.
Mentions: Most vertebrate orthologs are contained in chromosomal regions that are syntenic to human (not shown), i.e., on chromosome 3F1 in mouse (this region is syntenic with human chromosome 4q32.1 as described in , chromosome 3 in chimpanzee (at 174 Mb), chromosome 15 in dog (at 55.9 Mb) and chromosome 4 in chicken (at 21.2 Mb). The X. laevis cDNA (645 amino-acids) was assembled using clones AW764964 and AW764609 (Table 1). The zebrafish gene is located on chromosome 1 at 26.02 Mb and has a similar intron-exon structure to that of human (not shown). Using the ASAP protein sequences from selected species we performed a phylogenetic comparison using Clustalw with a bootstrap method. As shown in Fig. 2, the human and dog proteins are very similar (>78% PSI) whereas mouse ASAP is more divergent. Although frog and fish ASAP are more distant than those from the other species, they do not constitute a separate family, since they have a similar topology and share >32–38% PSI and >44–48% sequence similarity with the human protein. Pairwise sequence comparisons confirmed these results (Fig. 2B).

Bottom Line: The human gene is strongly expressed in brain and testis as a 2.6 Kb transcript encoding a approximately110 KDa protein.The protein contains MAP, MIT-like and THY domains in the C-terminal part indicative of microtubule interaction, while the N-terminal part is more divergent.It may have a role in spermatogenesis and also represents a potential new target for antitumoral drugs.

View Article: PubMed Central - HTML - PubMed

Affiliation: Groupe Microtubules et Cycle Cellulaire, Institut de Génétique Humaine, CNRS UPR 1142, rue de cardonille, 34396 Montpellier cédex 5, France. Magali.Venoux@igh.cnrs.fr

ABSTRACT

Background: ASAP is a newly characterized microtubule-associated protein (MAP) essential for proper cell-cycling. We have previously shown that expression deregulation of human ASAP results in profound defects in mitotic spindle formation and mitotic progression leading to aneuploidy, cytokinesis defects and/or cell death. In the present work we analyze the structure and evolution of the ASAP gene, as well as the domain composition of the encoded protein. Mouse and Xenopus cDNAs were cloned, the tissue expression characterized and the overexpression profile analyzed.

Results: Bona fide ASAP orthologs are found in vertebrates with more distantly related potential orthologs in invertebrates. This single-copy gene is conserved in mammals where it maps to syntenic chromosomal regions, but is also clearly identified in bird, fish and frog. The human gene is strongly expressed in brain and testis as a 2.6 Kb transcript encoding a approximately110 KDa protein. The protein contains MAP, MIT-like and THY domains in the C-terminal part indicative of microtubule interaction, while the N-terminal part is more divergent. ASAP is composed of approximately 42% alpha helical structures, and two main coiled-coil regions have been identified. Different sequence features may suggest a role in DNA damage response. As with human ASAP, the mouse and Xenopus proteins localize to the microtubule network in interphase and to the mitotic spindle during mitosis. Overexpression of the mouse protein induces mitotic defects similar to those observed in human. In situ hybridization in testis localized ASAP to the germ cells, whereas in culture neurons ASAP localized to the cell body and growing neurites.

Conclusion: The conservation of ASAP indicated in our results reflects an essential function in vertebrates. We have cloned the ASAP orthologs in mouse and Xenopus, two valuable models to study the function of ASAP. Tissue expression of ASAP revealed a high expression in brain and testis, two tissues rich in microtubules. ASAP associates to the mitotic spindle and cytoplasmic microtubules, and represents a key factor of mitosis with possible involvement in other cell cycle processes. It may have a role in spermatogenesis and also represents a potential new target for antitumoral drugs. Possible involvement in neuron dynamics also highlights ASAP as a candidate target in neurodegenerative diseases.

Show MeSH
Related in: MedlinePlus