Evidence that talin alternative splice variants from Ciona intestinalis have different roles in cell adhesion
© Singiser and McCann; licensee BioMed Central Ltd. 2006
Received: 13 September 2006
Accepted: 06 December 2006
Published: 06 December 2006
Talins are large, modular cytoskeletal proteins found in animals and amoebozoans such as Dictyostelium discoideum. Since the identification of a second talin gene in vertebrates, it has become increasingly clear that vertebrate Talin1 and Talin2 have non-redundant roles as essential links between integrins and the actin cytoskeleton in distinct plasma membrane-associated adhesion complexes. The conserved C-terminal I/LWEQ module is important for talin function. This structural element mediates the interaction of talins with F-actin. The I/LWEQ module also targets mammalian Talin1 to focal adhesion complexes, which are dynamic multicomponent assemblies required for cell adhesion and cell motility. Although Talin1 is essential for focal adhesion function, Talin2 is not targeted to focal adhesions. The nonvertebrate chordate Ciona intestinalis has only one talin gene, but alternative splicing of the talin mRNA produces two proteins with different C-terminal I/LWEQ modules. Thus, C. intestinalis contains two talins, Talin-a and Talin-b, with potentially different activities, despite having only one talin gene.
We show here that, based on their distribution in cDNA libraries, Talin-a and Talin-b are differentially expressed during C. intestinalis development. The I/LWEQ modules of the two proteins also have different affinities for F-actin. Consistent with the hypothesis that Talin-a and Talin-b have different roles in cell adhesion, the distinct I/LWEQ modules of Talin-a and Talin-b possess different subcellular targeting determinants. The I/LWEQ module of Talin-a is targeted to focal adhesions, where it most likely serves as the link between integrin and the actin cytoskeleton. The Talin-b I/LWEQ module is not targeted to focal adhesions, but instead preferentially labels F-actin stress fibers. These different properties of C. intestinalis the Talin-a and Talin-b I/LWEQ modules mimic the differences between mammalian Talin1 and Talin2.
Vertebrates and D. discoideum contain two talin genes that encode proteins with different functions. The urochordate C. intestinalis has a single talin gene but produces two separate talins by alternative splicing that vary in a domain crucial for talin function. This suggests that multicellular organisms require multiple talins as components of adhesion complexes. In C. intestinalis, alternative splicing, rather than gene duplication followed by neo-functionalization, accounts for the presence of multiple talins with different properties. Given that C. intestinalis is an excellent model system for chordate biology, the study of Talin-a and Talin-b will lead to a deeper understanding of cell adhesion in the chordate lineage and how talin functions have been parceled out to multiple proteins during metazoan evolution.
In contrast to D. discoideum and vertebrates, invertebrates including Drosophila melanogaster and Caenorhabditis elegans and the nonvertebrate chordate Ciona intestinalis have only one talin gene . However, alternative splicing of the talin mRNA in D. melanogaster and C. intestinalis produces multiple talins that vary at their C-termini . Therefore, despite having only one talin gene, multiple talin proteins can account for the biological functions of talin in these organisms.
Little is currently known about the functions of these talin splice variants. In this report we have determined that the I/LWEQ modules of the alternative splice variants of C. intestinalis talin have different actin-binding affinities and different subcellular targeting signals. We show that the Talin-b I/LWEQ module has a higher affinity for F-actin than the Talin-a I/LWEQ module and is also targeted to actin stress fibers in cells. In contrast, the Talin-a I/LWEQ module does not label stress fibers, but is instead targeted to focal adhesions. These differences are similar to those seen with Talin1 and Talin2 in vertebrates, where Talin1 is the link between integrins and the actin cytoskeleton in focal adhesions [11, 13]. These results suggest that alternative splicing is an effective mechanism for producing distinct talin proteins with different functions in C. intestinalis. The recent completion of its genome sequence has increased the utility of C. intestinalis as a model chordate [14–17]. Thus, further studies of Talin-a and Talin-b in C. intestinalis will be useful in identifying fundamental roles for distinct talin proteins in cell adhesion in chordates.
Identification of C. intestinalis Talin-a and Talin-b
Distribution of Talin-a and Talin-b cDNAs.
gastrula (C. savignyi)
larva (M. tectiformis)
Actin binding activity of the Talin-a and Talin-b I/LWEQ modules
Subcellular targeting of Talin-a and Talin-b
Analysis of EST data produced primarily by Nori Satoh and colleagues of Kyoto University  has shown that alternative splicing of talin pre-mRNA leads to the production of two proteins with different C-termini in C. intestinalis. The variable distributions of Talin-a and Talin-b in different cDNA libraries suggest that Talin-a and Talin-b are differentially expressed during C. intestinalis development. We have also shown that the variant I/LWEQ modules of Talin-a and Talin-b have different biological properties: Talin-b has the higher affinity for F-actin and preferentially labels actin stress fibers, while Talin-a is targeted to focal adhesions. Therefore, alternative processing is a mechanism for producing multiple talin proteins, with distinct activities associated with a critical structural element, in these nonvertebrate chordates, even though the C. intestinalis genome contains only one talin gene.
Alternative splicing is a widespread mechanism for expanding the proteome. In humans, 30–65% of all gene products are subject to alternative splicing, and similar levels are also seen in other organisms, including the mouse, fruitfly, and nematode . Alternative splicing frequently results in changes in protein structure that alter protein-protein interactions. For example, alternative splicing of the non-erythroid protein 4.1R produces variants that alter the efficiency of the interaction with the spectrin/actin complex by a factor of two . This is similar to differences we have identified in the C. intestinalis talin alternative splice variants, where the Talin-b I/LWEQ module interacts with F-actin with a four-fold higher affinity than that of Talin-a. The higher affinity for F-actin may explain the preferential localization of the Talin-b I/LWEQ module to actin stress fibers in cells.
Alternative splicing can also alter subcellular targeting determinants of the resultant proteins . HeLa cells represent a heterologous system for the study of tunicate talins in vivo, but we have recently used them to determine that the C-terminal I/LWEQ module of Talin1 contains a focal adhesion targeting determinant . We have shown here that the Talin-a I/LWEQ module, but not that of Talin-b, is targeted to focal adhesions in HeLa cells. Although focal adhesions have not yet been directly observed in C. intestinalis cells, the C. intestinalis genome contains genes for several characteristic focal adhesion components that were previously thought to be restricted to vertebrates. These include at least 14 α-integrins, 5 β-integrins, focal adhesion kinase, vinculin, and α-actinin . Vinculin and β-integrin are well-characterized Talin1 binding partners. Given the presence of these classical focal adhesion proteins in C. intestinalis, our results suggest that Talin-a is likely to function similarly to mammalian Talin1 as an adhesion complex component during C. intestinalis development. The lack of a focal adhesion-targeting determinant in the C-terminus of Talin-b may indicate that this protein is a component of other adhesion complexes in the organism, as we have seen with mammalian Talin2. Talin2 is not targeted to focal adhesions in undifferentiated cells but is preferentially targeted to costameres and intercalated disks, which are stable adhesions in differentiated striated muscle .
The presence of two related, but ancient and highly divergent talins (30% identical) with different physiological roles in D. discoideum [4–6] supports the hypothesis that gene duplication, followed by neo-functionalization, produced multiple talins that are required for cell adhesion during cell differentiation [3, 29]. A similar explanation may also account for the maintenance of Talin1 and Talin2 in vertebrates since duplication of the ancestral animal talin early in the chordate lineage, more than 450 million years ago . In support of this hypothesis regarding vertebrate talins, several recent studies have shown that these distinct proteins indeed have different roles. Talin1 and Talin2 interact with different proteins [3, 30, 31] and have different affinities for their common partner F-actin . Talin1 is required for mammalian embryogenesis, probably as an essential component of adhesion complexes required for cell motility during gastrulation . Talin2 is unable to complement this lethal phenotype. We have recently shown that the adult tissue distributions of mammalian Talin1 and Talin2 also vary, with Talin1 being the more widely expressed isoform . Talin2 is most abundant in brain and muscle. We have shown that Talin2 is induced during muscle differentiation along with other muscle-specific proteins such as archvillin, integrin-β1D, and metavinculin, which is an alternative splice variant of vinculin . As expected, Talin1 is a component of focal adhesions in undifferentiated myoblasts, but Talin2 is not found in focal adhesions of these cells or other mammalian cells. Talin2 is instead a component of stable adhesion assemblies such as costameres and intercalated disks in mature striated muscle . Taken together, these studies indicate that vertebrate Talin1 and Talin2 are differentially expressed during cell differentiation and organismal development and that they function as non-redundant components of distinct adhesion complexes in different cells and tissues. Thus, in both D. discoideum and vertebrates, multiple talins are involved in talin functions.
Development in C. intestinalis is a complex, multistage process. Following fertilization and subsequent embryogenesis, a free-swimming tadpole larva containing a notochord, nervous system, and musculature settles upon a solid substrate and metamorphoses into a sessile, filter-feeding organism. The adult contains organs common to other chordates, including a digestive system, heart, and nervous system . C. intestinalis has only one talin gene, but the existence of the splice variants Talin-a and Talin-b, which vary in the important C-terminal I/LWEQ module , further supports the emerging paradigm that talin function is due to the actions of distinct proteins, which are expressed at different times and in different places during chordate development [3, 10]. The results presented here suggest that alternative splicing is the means by which two talins with different roles in cell adhesion, and perhaps cell differentiation, are produced from one talin gene in C. intestinalis. Thus, this mechanism may bridge the one-talin bottleneck present in invertebrates between the multicellular amoebozoan D. discoideum  and vertebrates . Further studies at the cellular and molecular levels will identify when and where Talin-a and Talin-b are produced in C. intestinalis, and how they contribute to the assembly and function of adhesion complexes during development of this model organism.
Taken together with previous studies showing that talin function is due to the activities of distinct proteins, which are differentially expressed and targeted to different adhesion complexes in multicellular organisms, our results show that alternative splicing of the C. intestinalis talin mRNA produces two different proteins, Talin-a and Talin-b. These proteins vary in the C-terminal I/LWEQ module, which is critical for talin function. Moreover, these differences lead to different activities of the alternative splice variants. Our data suggest that C. intestinalis Talin-a is involved in cell adhesion and motility in assemblies that are similar to focal adhesions. Identification of the structural determinants responsible for the different properties of Talin-a and Talin-b will expand our knowledge of how different talins are involved in cell adhesion in chordates. Given that C. intestinalis has recently re-emerged as a model for the study of chordate development [33, 35], further research comparing the roles of Talin-a and Talin-b during C. intestinalis development will also increase our knowledge of how talin functions have evolved during animal evolution.
Identification of Talin-a and Talin-b
Talin-a and Talin-b cDNAs were identified in BLAST searches  of the GenBank non-human, non-mouse EST database using the C-terminal 250 amino acid sequence of Ciona intestinalis talin [GenBank: AABS01000025] as the query sequence . Unambiguous Talin-a and Talin-b cDNAs were identified by comparison of the C-terminal protein sequences with the original Talin-a and Talin-b that we identified previously . Full-length and partial C-terminal chordate talin sequences were aligned with CLUSTAL W  using the European Bioinformatics Institute server as previously described . Unrooted trees were then calculated from these alignments using TreeView . The sequence alignments in Fig. 2 were prepared for display using MacBoxShade as previously described .
PCR amplification of Talin-a/b from Ciona intestinalis and protein preparation
Whole Ciona intestinalis adults were obtained from the Marine Biological Laboratory (Woods Hole, MA). Organisms were frozen in liquid nitrogen, pulverized, and homogenized. RNA was extracted using RNAqueous-Midi (Ambion). First strand cDNA was synthesized using Superscript II reverse transcriptase (Invitrogen) with 1 μg total RNA and the oligo dT primer (5'-GACTCGAGTCGACATCGATTTTTTTTTTTTTTTTT-3'). Talin-a/b regions were then amplified by PCR using gene specific primers with the first-strand cDNA as a template: Talin-a/b sense, 5'-CCGGAATTCATTTTGGAAGCCGCAAAATCAATCGC-3' Talin-a antisense, 5'-CCTGGCGCGCCTTAATCGGATTCAGAATCATCCTTGT-3' Talin-b antisense, 5'-CCTGGCGCGCCCTATTTCTTTTGGTTTTGCTCGTATTTC-3' PCR amplicons were recovered using pCR2.1-TOPO (Invitrogen) and then subcloned into pET-41 (Novagen) for production of GST-fusion proteins. The proteins produced correspond to amino acids 2341–2531 for both Talin-a and Talin-b. All constructs were verified by DNA sequencing. Recombinant GST fusion proteins were prepared as previously described [2, 22].
Actin was obtained from Cytoskeleton, Inc (Denver, CO). The binding of the I/LWEQ modules of Talin-a and Talin-b to F-actin was determined as previously described [2, 22]. Briefly, F-actin co-sedimentation assays were performed in buffer A (2 mM Tris, pH 8.0, 0.2 mM CaCl2, 0.2 mM ATP, and 0.5 mM dithiothreitol), keeping the concentration of GST-Talin-a/b.2341–2531 constant at 4.0 μM while increasing the actin concentration from 0 to 25 μM in a total volume of 50 μl. Actin polymerization was induced by addition of 2 mM MgCl2 and 50 mM KCl and allowed to proceed for 60 minutes at 22°C. The critical concentration for actin polymerization is 0.71 μM under these conditions. Following centrifugation at 160000 g for 20 minutes at 22°C, the supernatant and pellet fractions were separated for subsequent analysis with SDS-PAGE using 12% gels. Proteins were stained with Coomassie Blue G250. Digital quantification of the cosedimentation data was performed using NIH Image v1.62 and binding curves were calculated using GraphPad Prism 4.0 as previously described [2, 22]. The R2 values for the binding curves were 0.95 for Talin-a and 0.99 for Talin-b. Neither GST-Talin-a.2341–2531 nor GST-Talin-b.2341–2531 sedimented at 160,000 g in the absence of F-actin. We have determined previously that the presence of the GST purification tag has no effect on actin binding of I/LWEQ module proteins [2, 22]. The concentration of actin was determined using its molar extinction coefficient at 290 nm. The concentrations of GST-Talin-a and GST-Talin-b were determined using the BCA protein assay (Pierce) and by calculating the extinction coefficient of each protein .
C. intestinalis Talin-a/b I/LWEQ module constructs were subcloned from pET-41 to pDsRedC2-monomer (BD Biosciences) and verified by DNA sequencing. HeLa cells (American Type Culture Collection CCL-2) were maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% FBS, 100 μg/ml penicillin, 100 μg/ml streptomycin and 10 μg/ml gentamicin. Glass coverslips were ethanol-washed and coated with 10 μg/ml fibronectin. HeLa cells were plated on the coverslips in complete growth medium and allowed to adhere for 12 hours. Transfection of HeLa cells was performed with Lipofectamine 2000 (Invitrogen, Carlsbad, CA). Cells were transfected with 0.8 μg of DNA and 2 μl Lipofectamine 2000 in normal growth medium for 24 hours. Cells were then fixed with 4% paraformadehyde, permeabilized briefly on ice with 1% Triton X-100, and labeled with either anti-vinculin antibody at a dilution of 1:300 (V9131, Sigma) or FITC-phalloidin at 1:400 dilution as previously described in our analysis of the subcellular targeting of mammalian Talin1 . Fluorescence images were obtained with a Zeiss Axiovert 200 microscope and processed using OpenLab (Improvision). Expression of intact dsRed-Talin-a/b constructs in transfected cells was confirmed in immunoblots using an anti-dsRed polyclonal antibody. We have shown previously that the addition of a fluorescence tag to the N-terminus of both full-length talin and talin I/LWEQ module constructs has no effect on the talin subcellular targeting [10, 13]. Transfection of HeLa cells with the dsRed vector alone did not affect the morphology of HeLa cells and dsRed protein was diffusely distributed throughout the cytoplasm.
The work was supported by research grants from the National Institutes of Health (P20RR20171) and the Ohio Valley Affiliate of the American Heart Association (0365218B) to R.O. McCann. R.H. Singiser is supported by a predoctoral fellowship from the Ohio Valley Affiliate of the American Heart Association (0615261B).
- Critchley DR: Talin. Guidebook to the Extracellular Matrix, Anchor, and Adhesion Proteins. Edited by: Kreis T, Vale R. 1999, Oxford , Oxford University Press, 82-85.
- McCann RO, Craig SW: The I/LWEQ module: a conserved sequence that signifies F-actin binding in functionally diverse proteins from yeast to mammals. Proc Natl Acad Sci U S A. 1997, 94 (11): 5679-5684. 10.1073/pnas.94.11.5679.PubMed CentralView ArticlePubMed
- Senetar MA, McCann RO: Gene duplication and functional divergence during evolution of the cytoskeletal linker protein talin. Gene. 2005, 362: 141-152. 10.1016/j.gene.2005.08.012.View ArticlePubMed
- Kreitmeier M, Gerisch G, Heizer C, Muller-Taubenberger A: A talin homologue of Dictyostelium rapidly assembles at the leading edge of cells in response to chemoattractant. J Cell Biol. 1995, 129 (1): 179-188. 10.1083/jcb.129.1.179.View ArticlePubMed
- Niewohner J, Weber I, Maniak M, Muller-Taubenberger A, Gerisch G: Talin-null cells of Dictyostelium are strongly defective in adhesion to particle and substrate surfaces and slightly impaired in cytokinesis. J Cell Biol. 1997, 138 (2): 349-361. 10.1083/jcb.138.2.349.PubMed CentralView ArticlePubMed
- Tsujioka M, Machesky LM, Cole SL, Yahata K, Inouye K: A unique talin homologue with a villin headpiece-like domain is required for multicellular morphogenesis in Dictyostelium. Curr Biol. 1999, 9 (7): 389-392. 10.1016/S0960-9822(99)80169-9.View ArticlePubMed
- Tsujioka M, Yoshida K, Inouye K: Talin B is required for force transmission in morphogenesis of Dictyostelium. Embo J. 2004, 23 (11): 2216-2225. 10.1038/sj.emboj.7600238.PubMed CentralView ArticlePubMed
- McCann RO, Craig SW: Identification of a novel isoform of the focal adhesion protein talin. Mol Biol Cell. 1998, 9 (Suppl S Nov 1998): 798-
- McCann RO, Craig SW: Functional genomic analysis reveals the utility of the I/LWEQ module as a predictor of protein:actin interaction. Biochem Biophys Res Commun. 1999, 266 (1): 135-140. 10.1006/bbrc.1999.1776.View ArticlePubMed
- Senetar MA, Moncman CL, McCann RO: Talin2 is Induced During Striated Muscle Differentiation and is Targeted to Stable Adhesion Complexes in Mature Muscle. Cell Motil Cytoskel. 2006, In press:
- Franco SJ, Rodgers MA, Perrin BJ, Han J, Bennin DA, Critchley DR, Huttenlocher A: Calpain-mediated proteolysis of talin regulates adhesion dynamics. Nat Cell Biol. 2004, 6: 977-983. 10.1038/ncb1175.View ArticlePubMed
- Baldauf SL: The deep roots of eukaryotes. Science. 2003, 300: 1703-1706. 10.1126/science.1085544.View ArticlePubMed
- Franco SJ, Senetar MA, Simonson W, Huttenlocher A, McCann RO: The conserved C-terminal I/LWEQ module targets Talin1 to focal adhesions. Cell Motility and the Cytoskeleton. 2006, 63: 563-581. 10.1002/cm.20145.View ArticlePubMed
- Dehal P, Satou Y, Campbell RK, Chapman J, Degnan B, De Tomaso A, Davidson B, Di Gregorio A, Gelpke M, Goodstein DM, Harafuji N, Hastings KE, Ho I, Hotta K, Huang W, Kawashima T, Lemaire P, Martinez D, Meinertzhagen IA, Necula S, Nonaka M, Putnam N, Rash S, Saiga H, Satake M, Terry A, Yamada L, Wang HG, Awazu S, Azumi K, Boore J, Branno M, Chin-Bow S, DeSantis R, Doyle S, Francino P, Keys DN, Haga S, Hayashi H, Hino K, Imai KS, Inaba K, Kano S, Kobayashi K, Kobayashi M, Lee BI, Makabe KW, Manohar C, Matassi G, Medina M, Mochizuki Y, Mount S, Morishita T, Miura S, Nakayama A, Nishizaka S, Nomoto H, Ohta F, Oishi K, Rigoutsos I, Sano M, Sasaki A, Sasakura Y, Shoguchi E, Shin-i T, Spagnuolo A, Stainier D, Suzuki MM, Tassy O, Takatori N, Tokuoka M, Yagi K, Yoshizaki F, Wada S, Zhang C, Hyatt PD, Larimer F, Detter C, Doggett N, Glavina T, Hawkins T, Richardson P, Lucas S, Kohara Y, Levine M, Satoh N, Rokhsar DS: The draft genome of Ciona intestinalis: insights into chordate and vertebrate origins. Science. 2002, 298 (5601): 2157-2167. 10.1126/science.1080049.View ArticlePubMed
- Sasakura Y, E. S, Takatori N, Wada S, Meinertzhagen IA, Satou Y, Satoh N: A genomewide survey of developmentally relevant genes in Ciona intestinalis X. Genes for cell junctions and extracellular matrix. Dev Genes and Evolution. 2003, 213: 303-313. 10.1007/s00427-003-0320-1.View Article
- Holland LZ, Gibson-Brown JJ: The Ciona intestinalis genome: when the constraints are off. Bioessays. 2003, 25 (6): 529-532. 10.1002/bies.10302.View ArticlePubMed
- Canestro C, Bassham S, Postlethwait JH: Seeing chordate evolution through the Ciona genome sequence. Genome Biol. 2003, 4 (3): 208-10.1186/gb-2003-4-3-208.PubMed CentralView ArticlePubMed
- Bateman A, Coin L, Durbin R, Finn RD, Hollich V, Griffiths-Jones S, Khanna A, Marshall MS, Moxon S, Sonnhammer EL, Studholme DJ, Yeats C, Eddy SR: The Pfam protein families database. Nucleic Acids Res. 2004, 32: D138-D141. 10.1093/nar/gkh121.PubMed CentralView ArticlePubMed
- Thompson JD, Higgins DG, Gibson TJ: CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 1994, 22 (22): 4673-4680.PubMed CentralView ArticlePubMed
- Page RD: TreeView: an application to display phylogenetic trees on personal computers. Comput Appl Biosci. 1996, 12 (4): 357-358.PubMed
- Engqvist-Goldstein AE, Kessels MM, Chopra VS, Hayden MR, Drubin DG: An actin-binding protein of the Sla2/Huntingtin interacting protein-1 family is a novel component of clathrin-coated pits and vesicles. J Cell Biol. 1999, 147 (7): 1503-1518. 10.1083/jcb.147.7.1503.PubMed CentralView ArticlePubMed
- Senetar MA, Foster SJ, McCann RO: Intrasteric Inhibition Mediates the Interaction of the I/LWEQ module proteins Talin1, Talin2, Hip1, and Hip12 with Actin. Biochemistry. 2004, 43: 15418-15428. 10.1021/bi0487239.View ArticlePubMed
- Critchley DR: Focal adhesions - the cytoskeletal connection. Curr Opin Cell Biol. 2000, 12 (1): 133-139. 10.1016/S0955-0674(99)00067-8.View ArticlePubMed
- Craig SW, Johnson RP: Assembly of focal adhesions: progress, paradigms, and portents. Curr Opin Cell Biol. 1996, 8 (1): 74-85. 10.1016/S0955-0674(96)80051-2.View ArticlePubMed
- Albiges-Rizo C, Frachet P, Block MR: Down regulation of talin alters cell adhesion and the processing of the alpha 5 beta 1 integrin. J Cell Sci. 1995, 108 (Pt 10): 3317-3329.PubMed
- Satou Y, Yamada L, Mochizuki Y, Takatori N, Kawashima T, Sasaki A, Hamaguchi M, Awazu S, Yagi K, Sasakura Y, Nakayama A, Ishikawa H, Inaba K, Satoh N: A cDNA resource from the basal chordate Ciona intestinalis. Genesis. 2002, 33 (4): 153-154. 10.1002/gene.10119.View ArticlePubMed
- Stamm S, Ben-Ari S, Rafalska I, Tang Y, Zhang Z, Toiber D, Thanaraj TA, Soreq H: Function of alternative splicing. Gene. 2005, 344: 1-20. 10.1016/j.gene.2004.10.022.View ArticlePubMed
- Kontrogianni-Konstantopoulos A, Frye CS, Benz EJ, Huang SC: The prototypical 4.1R-10-kDa domain and the 4.1g-10-kDa paralog mediate fodrin-actin complex formation. J Biol Chem. 2001, 276 (23): 20679-20687. 10.1074/jbc.M010581200.View ArticlePubMed
- He X, Zhang J: Rapid subfunctionalization accompanied by prolonged and substantial neofunctionalization in duplicate gene evolution. Genetics. 2005, 169: 1157-1164. 10.1534/genetics.104.037051.PubMed CentralView ArticlePubMed
- Di Paolo G, Pellegrini L, Letinic K, Cestra G, Zoncu R, Voronov S, Chang S, Guo J, Wenk MR, De Camilli P: Recruitment and regulation of phosphatidylinositol phosphate kinase type 1g by the FERM domain of talin. Nature. 2002, 420: 85-89. 10.1038/nature01147.View ArticlePubMed
- Ling K, Doughman RL, Firestone AJ, Bunce MW, Anderson RA: Type I gamma phosphatidylinositol phosphate kinase targets and regulates focal adhesions. Nature. 2002, 420 (6911): 89-93. 10.1038/nature01082.View ArticlePubMed
- Monkley SJ, Zhou XH, Kinston SJ, Giblett SM, Hemmings L, Brown JE, Pritchard CA, Critchley DR, Fassler R: Disruption of the talin gene arrests mouse development at the gastrulation stage. Dev Dyn. 2000, 219 (4): 560-574. 10.1002/1097-0177(2000)9999:9999<::AID-DVDY1079>3.0.CO;2-Y.View ArticlePubMed
- Kume M, Dan K: Invertebrate Embryology. 1988, New York , Garland
- Kessin RH: Dictyostelium: Evolution, Cell Biology, and the Development of Multicellularity. 2001, Cambridge , Cambridge University PressView Article
- Passamaneck YJ, Di Gregorio A: Ciona intestinalis: chordate development made simple. Dev Dyn. 2005, 233 (1): 1-19. 10.1002/dvdy.20300.View ArticlePubMed
- Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ: Basic local alignment search tool. J Mol Biol. 1990, 215: 403-410. 10.1006/jmbi.1990.9999.View ArticlePubMed
- Gill SC, von Hippel PH: Calculation of protein extinction coefficients from amino acid sequence data [published erratum appears in Anal Biochem 1990 Sep;189(2):283]. Anal Biochem. 1989, 182 (2): 319-326. 10.1016/0003-2697(89)90602-7.View ArticlePubMed
- Chishti AH, Kim AC, Marfatia SM, Lutchman M, Hanspal M, Jindal H, Liu SC, Low PS, Rouleau GA, Mohandas N, Chasis JA, Conboy JG, Gascard P, Takakuwa Y, Huang SC, Benz EJ, Bretscher A, Fehon RG, Gusella JF, Ramesh V, Solomon F, Marchesi VT, Tsukita S, Tsukita S, Arpin M, Louvard D, Tonks NK, Anderson JM, Fanning AS, Bryant PJ, Woods DF, Hoover KB: The FERM domain: a unique module involved in the linkage of cytoplasmic proteins to the membrane. Trends Biochem Sci. 1998, 23 (8): 281-282. 10.1016/S0968-0004(98)01237-7.View ArticlePubMed
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.