Computational modeling of the bHLH domain of the transcription factor TWIST1 and R118C, S144R and K145E mutants

BMC Bioinformatics

Eliana Abdelhay¹  Ernesto R Caffarena²  André L Mencalha¹  João HM da Silva²  Amanda M Maia¹ 

[1]Laboratório de Célula-tronco – CEMO/INCA, Praça da Cruz Vermelha 23 6 andar, Centro, Rio de Janeiro/RJ, Brasil

[2]Laboratório de Biofísica Computacional e Modelagem Molecular – PROCC/ FIOCRUZ, Av Brasil, 4365, Manguinhos, Rio de Janeiro/RJ, Brasil

关键词: Collective motions;    Molecular dynamics simulation;    Comparative modeling;    bHLH;    Transcription factor;    Twist1;   

Others  :  1088186
DOI  :  10.1186/1471-2105-13-184

 received in 2012-02-16, accepted in 2012-07-17,  发布年份 2012

【 摘 要 】

Background

Human TWIST1 is a highly conserved member of the regulatory basic helix-loop-helix (bHLH) transcription factors. TWIST1 forms homo- or heterodimers with E-box proteins, such as E2A (isoforms E12 and E47), MYOD and HAND2. Haploinsufficiency germ-line mutations of the twist1 gene in humans are the main cause of Saethre-Chotzen syndrome (SCS), which is characterized by limb abnormalities and premature fusion of cranial sutures. Because of the importance of TWIST1 in the regulation of embryonic development and its relationship with SCS, along with the lack of an experimentally solved 3D structure, we performed comparative modeling for the TWIST1 bHLH region arranged into wild-type homodimers and heterodimers with E47. In addition, three mutations that promote DNA binding failure (R118C, S144R and K145E) were studied on the TWIST1 monomer. We also explored the behavior of the mutant forms in aqueous solution using molecular dynamics (MD) simulations, focusing on the structural changes of the wild-type versus mutant dimers.

Results

The solvent-accessible surface area of the homodimers was smaller on wild-type dimers, which indicates that the cleft between the monomers remained more open on the mutant homodimers. RMSD and RMSF analyses indicated that mutated dimers presented values that were higher than those for the wild-type dimers. For a more careful investigation, the monomer was subdivided into four regions: basic, helix I, loop and helix II. The basic domain presented a higher flexibility in all of the parameters that were analyzed, and the mutant dimer basic domains presented values that were higher than the wild-type dimers. The essential dynamic analysis also indicated a higher collective motion for the basic domain.

Conclusions

Our results suggest the mutations studied turned the dimers into more unstable structures with a wider cleft, which may be a reason for the loss of DNA binding capacity observed for in vitro circumstances.

【 授权许可】

   
2012 Maia et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

Figure 7.

【 参考文献 】

• [1]Anderson DJ, et al.: Cell lineage determination and the control of neuronal identity in the neural crest. Cold Spring Harb Symp Quant Biol 1997, 62:493-504.
• [2]Lister JA, Baron MH: Induction of basic helix-loop-helix protein-containing complexes during erythroid differentiation. Gene Expr 1998, 7(1):25-38.
• [3]Murray SS, et al.: Expression of helix-loop-helix regulatory genes during differentiation of mouse osteoblastic cells. J Bone Miner Res 1992, 7(10):1131-8.
• [4]Murre C, McCaw PS, Baltimore D: A new DNA binding and dimerization motif in immunoglobulin enhancer binding, daughterless, MyoD, and myc proteins. Cell 1989, 56(5):777-83.
• [5]Firulli BA, et al.: Altered Twist1 and Hand2 dimerization is associated with Saethre-Chotzen syndrome and limb abnormalities. Nat Genet 2005, 37(4):373-81.
• [6]Castanon I, et al.: Dimerization partners determine the activity of the Twist bHLH protein during Drosophila mesoderm development. Development 2001, 128(16):3145-59.
• [7]Kophengnavong T, Michnowicz JE, Blackwell TK: Establishment of distinct MyoD, E2A, and twist DNA binding specificities by different basic region-DNA conformations. Mol Cell Biol 2000, 20(1):261-72.
• [8]Hayashi M, et al.: Comparative roles of Twist-1 and Id1 in transcriptional regulation by BMP signaling. J Cell Sci 2007, 120(Pt 8):1350-7.
• [9]Funato N, et al.: Common regulation of growth arrest and differentiation of osteoblasts by helix-loop-helix factors. Mol Cell Biol 2001, 21(21):7416-28.
• [10]Sun XH, Baltimore D: An inhibitory domain of E12 transcription factor prevents DNA binding in E12 homodimers but not in E12 heterodimers. Cell 1991, 64(2):459-70.
• [11]Aronheim A, et al.: Distribution and characterization of helix-loop-helix enhancer-binding proteins from pancreatic beta cells and lymphocytes. Nucleic Acids Res 1991, 19(14):3893-9.
• [12]Lassar AB, et al.: Functional activity of myogenic HLH proteins requires hetero-oligomerization with E12/E47-like proteins in vivo. Cell 1991, 66(2):305-15.
• [13]Shen CP, Kadesch T: B-cell-specific DNA binding by an E47 homodimer. Mol Cell Biol 1995, 15(8):4518-24.
• [14]Chen ZF, Behringer RR: twist is required in head mesenchyme for cranial neural tube morphogenesis. Genes Dev 1995, 9(6):686-99.
• [15]Wilkie AO, Morriss-Kay GM: Genetics of craniofacial development and malformation. Nat Rev Genet 2001, 2(6):458-68.
• [16]Gripp KW, Zackai EH, Stolle CA: Mutations in the human TWIST gene. Hum Mutat 2000, 15(2):150-5.
• [17]El Ghouzzi V, et al.: Mutations within or upstream of the basic helix-loop-helix domain of the TWIST gene are specific to Saethre-Chotzen syndrome. Eur J Hum Genet 1999, 7(1):27-33.
• [18]El Ghouzzi V, et al.: Saethre-Chotzen mutations cause TWIST protein degradation or impaired nuclear location. Hum Mol Genet 2000, 9(5):813-9.
• [19]Puntervoll P, et al.: ELM server: a new resource for investigating short functional sites in modular eukaryotic proteins. Nucleic Acids Research 2003, 31:3625-3630.
• [20]Linding R, et al.: GlobPlot: Exploring protein sequences for globularity and disorder. Nucleic Acids Res 2003, 31(13):3701-8.
• [21]Altschul SF, et al.: Basic local alignment search tool. J Mol Biol 1990, 215(3):403-10.
• [22]Berman HM, et al.: The Protein Data Bank. Nucleic Acids Res 2000, 28(1):235-42.
• [23]Higgins D, et al.: CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Research 1994, 22:4673-4680.
• [24]Larkin MA, et al.: Clustal W and Clustal X version 2.0. Bioinformatics 2007, 23(21):2947-8.
• [25]Sali A, Blundell TL: Comparative protein modelling by satisfaction of spatial restraints. J Mol Biol 1993, 234(3):779-815.
• [26]Braun W, Go N: Calculation of protein conformations by proton-proton distance constraints. A new efficient algorithm. J Mol Biol 1985, 186(3):611-26.
• [27]Guex N, Peitsch MC: SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modeling. Electrophoresis 1997, 18(15):2714-23.
• [28]Benkert P, Tosatto SC, Schomburg D: QMEAN: A comprehensive scoring function for model quality assessment. Proteins 2008, 71(1):261-77.
• [29]Laskowski RA, et al.: PROCHECK: A program to check the stereochemical quality of protein structures. Journal of Applied Crystallography 1993, 26:283-291.
• [30]Zhou H, Zhou Y: Distance-scaled, finite ideal-gas reference state improves structure-derived potentials of mean force for structure selection and stability prediction. Protein Sci 2002, 11(11):2714-26.
• [31]Benkert P, Biasini M, Schwede T: Toward the estimation of the absolute quality of individual protein structure models. Bioinformatics 2011, 27(3):343-50.
• [32]Van Der Spoel D, et al.: GROMACS: fast, flexible, and free. J Comput Chem 2005, 26(16):1701-18.
• [33]Jorgensen WL, et al.: Comparison of Simple Potential Functions for Simulating Liquid Water. Journal of Chemical Physics 1983, 79(2):926-935.
• [34]Jorgensen WL, Maxwell DS, TiradoRives J: Development and testing of the OPLS all-atom force field on conformational energetics and properties of organic liquids. Journal of the American Chemical Society 1996, 118(45):11225-11236.
• [35]Barrett CP, Hall BA, Noble ME: Dynamite: a simple way to gain insight into protein motions. Acta Crystallogr D Biol Crystallogr 2004, 60(Pt 12 Pt 1):2280-7.
• [36]Humphrey W, Dalke A, Schulten K: VMD: visual molecular dynamics. J Mol Graph 1996, 14(1):33-8. 27–8
• [37]Longo A, Guanga GP, Rose RB: Crystal structure of E47-NeuroD1/beta2 bHLH domain-DNA complex: heterodimer selectivity and DNA recognition. Biochemistry 2008, 47(1):218-29.
• [38]Amadei A, Linssen AB, Berendsen HJ: Essential dynamics of proteins. Proteins 1993, 17(4):412-25.
• [39]Minezaki Y, et al.: Human transcription factors contain a high fraction of intrinsically disordered regions essential for transcriptional regulation. J Mol Biol 2006, 359(4):1137-49.
• [40]Hamamori Y, et al.: Regulation of histone acetyltransferases p300 and PCAF by the bHLH protein twist and adenoviral oncoprotein E1A. Cell 1999, 96(3):405-13.
• [41]Singh S, Gramolini AO: Characterization of sequences in human TWIST required for nuclear localization. BMC Cell Biol 2009, 10:47. BioMed Central Full Text
• [42]Bialek P, et al.: A twist code determines the onset of osteoblast differentiation. Dev Cell 2004, 6(3):423-35.
• [43]Atchley WR, Terhalle W, Dress A: Positional dependence, cliques, and predictive motifs in the bHLH protein domain. J Mol Evol 1999, 48(5):501-16.
• [44]Connerney J, et al.: Twist1 homodimers enhance FGF responsiveness of the cranial sutures and promote suture closure. Dev Biol 2008, 318(2):323-34.
• [45]Teachenor R, et al.: Biochemical and phosphoproteomic analysis of the helix-loop-helix protein e47. Mol Cell Biol 2012, 32(9):1671-82.
• [46]Etzioni S, et al.: Homodimeric MyoD preferentially binds tetraplex structures of regulatory sequences of muscle-specific genes. J Biol Chem 2005, 280(29):26805-12.
• [47]Wendt H, Thomas RM, Ellenberger T: DNA-mediated folding and assembly of MyoD-E47 heterodimers. J Biol Chem 1998, 273(10):5735-43.
• [48]El Ghouzzi V, et al.: Mutations in the basic domain and the loop-helix II junction of TWIST abolish DNA binding in Saethre-Chotzen syndrome. FEBS Lett 2001, 492(1–2):112-8.