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A unique insertion in STARD9's motor domain regulates its stability.

Senese S, Cheung K, Lo YC, Gholkar AA, Xia X, Wohlschlegel JA, Torres JZ - Mol. Biol. Cell (2014)

Bottom Line: These phosphorylation events are important for targeting a pool of STARD9-MD for ubiquitination by the SCFβ-TrCP ubiquitin ligase and proteasome-dependent degradation.Of interest, overexpression of nonphosphorylatable/nondegradable STARD9-MD mutants leads to spindle assembly defects.Our results with STARD9-MD imply that in vivo the protein levels of full-length STARD9 could be regulated by Plk1 and SCFβ-TrCP to promote proper mitotic spindle assembly.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, CA 90095.

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The STARD9 motor domain loop 12 harbors a unique insertion with Plk1 and β-TrCP binding sites. (A) Schematic of the STARD9 modular domain composition. (B) Molecular modeling of the STARD9 motor domain using the KIF1A motor domain crystal structure (PDB ID 1IA0). Note that loops 7, 11, and 12 are extended. (C) Compared with KIF1A, STARD9 contains a 26–amino acid insertion in loop 12 with putative Plk1 polo-box domain–binding motifs (SS) and a β-TrCP DSGXXS–binding motif. (D) The 26–amino acid insertion and DSGXXS motif are highly conserved among STARD9 mammalian orthologues. (E) Alignment of STARD9, IκBα, claspin, and Emi1 β-TrCP DSGXXS–binding motifs and consensus.
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Figure 1: The STARD9 motor domain loop 12 harbors a unique insertion with Plk1 and β-TrCP binding sites. (A) Schematic of the STARD9 modular domain composition. (B) Molecular modeling of the STARD9 motor domain using the KIF1A motor domain crystal structure (PDB ID 1IA0). Note that loops 7, 11, and 12 are extended. (C) Compared with KIF1A, STARD9 contains a 26–amino acid insertion in loop 12 with putative Plk1 polo-box domain–binding motifs (SS) and a β-TrCP DSGXXS–binding motif. (D) The 26–amino acid insertion and DSGXXS motif are highly conserved among STARD9 mammalian orthologues. (E) Alignment of STARD9, IκBα, claspin, and Emi1 β-TrCP DSGXXS–binding motifs and consensus.

Mentions: The large size of STARD9 (∼517 kDa) greatly complicates its expression and molecular manipulation (Figure 1A). To better understand the function of STARD9 in cell division, we sought to characterize its kinesin motor domain, which plays a critical role in STARD9's function (Torres et al., 2011). To do this, we first modeled the structure of the STARD9-MD (amino acids 1–391) based on the KIF1A motor domain (with which it shares 48% sequence identity) crystal structure (Protein Data Bank [PDB] ID 1IA0) using the homology modeling program Modeller (Sali and Blundell, 1993; Figure 1B and Supplemental Figure S1). The STARD9-MD structure was evaluated with the structural analysis verification server and by Ramachandran plot, which showed that 92.1% of the residues were in the most favored region and 0% of the residues were in disallowed regions (Ramachandran et al., 1963; Luthy et al., 1992; Colovos and Yeates, 1993; Laskowski et al., 1993; Shen and Sali, 2006; Figure 1B and Supplemental Figures S1 and S2). Structure alignment of STARD9-MD with KIF1A-MD showed strong structural conservation, with the exception of a few extended loops (L7, L11, and L12) in STARD9 (Pettersen et al., 2004; Figure 1B and Supplemental Figure S3). Based on modeling, the STARD9 L12 was predicted to be unstructured and could potential adopt multiple conformations (one conformation is depicted in Figure 1B).


A unique insertion in STARD9's motor domain regulates its stability.

Senese S, Cheung K, Lo YC, Gholkar AA, Xia X, Wohlschlegel JA, Torres JZ - Mol. Biol. Cell (2014)

The STARD9 motor domain loop 12 harbors a unique insertion with Plk1 and β-TrCP binding sites. (A) Schematic of the STARD9 modular domain composition. (B) Molecular modeling of the STARD9 motor domain using the KIF1A motor domain crystal structure (PDB ID 1IA0). Note that loops 7, 11, and 12 are extended. (C) Compared with KIF1A, STARD9 contains a 26–amino acid insertion in loop 12 with putative Plk1 polo-box domain–binding motifs (SS) and a β-TrCP DSGXXS–binding motif. (D) The 26–amino acid insertion and DSGXXS motif are highly conserved among STARD9 mammalian orthologues. (E) Alignment of STARD9, IκBα, claspin, and Emi1 β-TrCP DSGXXS–binding motifs and consensus.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4310736&req=5

Figure 1: The STARD9 motor domain loop 12 harbors a unique insertion with Plk1 and β-TrCP binding sites. (A) Schematic of the STARD9 modular domain composition. (B) Molecular modeling of the STARD9 motor domain using the KIF1A motor domain crystal structure (PDB ID 1IA0). Note that loops 7, 11, and 12 are extended. (C) Compared with KIF1A, STARD9 contains a 26–amino acid insertion in loop 12 with putative Plk1 polo-box domain–binding motifs (SS) and a β-TrCP DSGXXS–binding motif. (D) The 26–amino acid insertion and DSGXXS motif are highly conserved among STARD9 mammalian orthologues. (E) Alignment of STARD9, IκBα, claspin, and Emi1 β-TrCP DSGXXS–binding motifs and consensus.
Mentions: The large size of STARD9 (∼517 kDa) greatly complicates its expression and molecular manipulation (Figure 1A). To better understand the function of STARD9 in cell division, we sought to characterize its kinesin motor domain, which plays a critical role in STARD9's function (Torres et al., 2011). To do this, we first modeled the structure of the STARD9-MD (amino acids 1–391) based on the KIF1A motor domain (with which it shares 48% sequence identity) crystal structure (Protein Data Bank [PDB] ID 1IA0) using the homology modeling program Modeller (Sali and Blundell, 1993; Figure 1B and Supplemental Figure S1). The STARD9-MD structure was evaluated with the structural analysis verification server and by Ramachandran plot, which showed that 92.1% of the residues were in the most favored region and 0% of the residues were in disallowed regions (Ramachandran et al., 1963; Luthy et al., 1992; Colovos and Yeates, 1993; Laskowski et al., 1993; Shen and Sali, 2006; Figure 1B and Supplemental Figures S1 and S2). Structure alignment of STARD9-MD with KIF1A-MD showed strong structural conservation, with the exception of a few extended loops (L7, L11, and L12) in STARD9 (Pettersen et al., 2004; Figure 1B and Supplemental Figure S3). Based on modeling, the STARD9 L12 was predicted to be unstructured and could potential adopt multiple conformations (one conformation is depicted in Figure 1B).

Bottom Line: These phosphorylation events are important for targeting a pool of STARD9-MD for ubiquitination by the SCFβ-TrCP ubiquitin ligase and proteasome-dependent degradation.Of interest, overexpression of nonphosphorylatable/nondegradable STARD9-MD mutants leads to spindle assembly defects.Our results with STARD9-MD imply that in vivo the protein levels of full-length STARD9 could be regulated by Plk1 and SCFβ-TrCP to promote proper mitotic spindle assembly.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, CA 90095.

Show MeSH
Related in: MedlinePlus