Emerging mechanisms and consequences of calcium regulation of alternative splicing in neurons and endocrine cells

Pan Q, Shai O, Lee LJ, Frey BJ, Blencowe BJ (2008) Deep surveying of alternative splicing complexity in the human transcriptome by high-throughput sequencing. Nat Genet 40:1413–1415 Wang ET, Sandberg R, Luo S, Khrebtukova I, Zhang L, Mayr C, Kingsmore SF, Schroth GP, Burge CB (2008) Alternative isoform regulation in human tissue transcriptome. Nature 456:470–476 Li Q, Lee JA, Black DL (2007) Neuronal regulation of alternative pre-mRNA splicing. Nat Rev Neurosci 8:819–831 Xie J (2008) Control of alternative pre-mRNA splicing by Ca(++) signals. Biochim Biophys Acta 1779:438–452 Norris AD, Calarco JA (2012) Emerging roles of alternative pre-mRNA splicing regulation in neuronal development and function. Front Neurosci 6:122 Sharma A, Lou H (2011) Depolarization-mediated regulation of alternative splicing. Front Neurosci 5:141 Teraoka SN, Telatar M, Becker-Catania S, Liang T, Onengut S, Tolun A, Chessa L, Sanal O, Bernatowska E, Gatti RA, Concannon P (1999) Splicing defects in the ataxia-telangiectasia gene, ATM: underlying mutations and consequences. Am J Hum Genet 64:1617–1631 Concannon P, Gatti RA (1997) Diversity of ATM gene mutations detected in patients with ataxia-telangiectasia. Hum Mutat 10:100–107 Ars E, Serra E, Garcia J, Kruyer H, Gaona A, Lazaro C, Estivill X (2000) Mutations affecting mRNA splicing are the most common molecular defects in patients with neurofibromatosis type 1. Hum Mol Genet 9:237–247 Will CL, Luhrmann R (2011) Spliceosome structure and function. Cold Spring Harb Perspect in Biol. 3:a003707 Chen M, Manley JL (2009) Mechanisms of alternative splicing regulation: insights from molecular and genomics approaches. Nat Rev Mol Cell Biol 10:741–754 Underwood JG, Boutz PL, Dougherty JD, Stoilov P, Black DL (2005) Homologues of the Caenorhabditis elegans Fox-1 protein are neuronal splicing regulators in mammals. Mol Cell Biol 25:10005–10016 Nakahata S, Kawamoto S (2005) Tissue-dependent isoforms of mammalian Fox-1 homologs are associated with tissue-specific splicing activities. Nucleic Acids Res 33:2078–2089 Gehman LT, Stoilov P, Maguire J, Damianov A, Lin CH, Shiue L, Ares M Jr, Mody I, Black DL (2011) The splicing regulator Rbfox1 (A2BP1) controls neuronal excitation in the mammalian brain. Nat Genet 43:706–711 Gehman LT, Meera P, Stoilov P, Shiue L, O’Brien JE, Meisler MH, Ares M Jr, Otis TS, Black DL (2012) The splicing regulator Rbfox2 is required for both cerebellar development and mature motor function. Genes Dev 26:445–460 Zhang C, Zhang Z, Castle J, Sun S, Johnson J, Krainer AR, Zhang MQ (2008) Defining the regulatory network of the tissue-specific splicing factors Fox-1 and Fox-2. Genes Dev 22:2550–2563 Boutz PL, Stoilov P, Li Q, Lin CH, Chawla G, Ostrow K, Shiue L, Ares M Jr, Black DL (2007) A post-transcriptional regulatory switch in polypyrimidine tract-binding proteins reprograms alternative splicing in developing neurons. Genes Dev 21:1636–1652 Zhu H, Hasman RA, Barron VA, Luo G, Lou H (2006) A nuclear function of Hu proteins as neuron-specific alternative RNA processing regulators. Mol Biol Cell 17:5105–5114 Szabo A, Dalmau J, Manley G, Rosenfeld M, Wong E, Henson J, Posner JB, Furneaux HM (1991) HuD, a paraneoplastic encephalomyelitis antigen, contains RNA-binding domains and is homologous to Elav and Sex-lethal. Cell 67:325–333 Buckanovich RJ, Posner JB, Darnell RB (1993) Nova, the paraneoplastic Ri antigen, is homologous to an RNA-binding protein and is specifically expressed in the developing motor system. Neuron 11:657–672 Yang YY, Yin GL, Darnell RB (1998) The neuronal RNA-binding protein Nova-2 is implicated as the autoantigen targeted in POMA patients with dementia. Proc Natl Acad Sci USA 95:13254–13259 Jensen KB, Dredge BK, Stefani G, Zhong R, Buckanovich RJ, Okano HJ, Yang YY, Darnell RB (2000) Nova-1 regulates neuron-specific alternative splicing and is essential for neuronal viability. Neuron 25:359–371 Stamm S (2008) Regulation of alternative splicing by reversible protein phosphorylation. J Biol Chem 283:1223–1227 Xiao SH, Manley JL (1997) Phosphorylation of the ASF/SF2 RS domain affects both protein–protein and protein–RNA interactions and is necessary for splicing. Genes Dev 11:334–344 Allemand E, Guil S, Myers M, Moscat J, Caceres JF, Krainer AR (2005) Regulation of heterogenous nuclear ribonucleoprotein A1 transport by phosphorylation in cells stressed by osmotic shock. Proc Natl Acad Sci USA 102:3605–3610 Liu G, Razanau A, Hai Y, Yu J, Sohail M, Lobo VG, Chu J, Kung SK, Xie J (2012) A conserved serine of heterogeneous nuclear ribonucleoprotein L (hnRNP L) mediates depolarization-regulated alternative splicing of potassium channels. J Biol Chem 287:22709–22716 Xie J, Lee JA, Kress TL, Mowry KL, Black DL (2003) Protein kinase A phosphorylation modulates transport of the polypyrimidine tract-binding protein. Proc Natl Acad Sci USA 100:8776–8781 Black DL (2003) Mechanisms of alternative pre-messenger RNA splicing. Annu Rev Biochem 72:291–336 Clapham DE (2007) Calcium signaling. Cell 131:1047–1058 Sudhof TC (2012) Calcium control of neurotransmitter release. Cold Spring Harb Perspect Biol 4(1):a011353 Zhang SJ, Zou M, Lu L, Lau D, Ditzel DA, Delucinge-Vivier C, Aso Y, Descombes P, Bading H (2009) Nuclear calcium signaling controls expression of a large gene pool: identification of a gene program for acquired neuroprotection induced by synaptic activity. PLoS Genet 5:e1000604 Alonso MT, Garcia-Sancho J (2011) Nuclear Ca(2+) signalling. Cell Calcium 49:280–289 Mellstrom B, Savignac M, Gomez-Villafuertes R, Naranjo JR (2008) Ca2+-operated transcriptional networks: molecular mechanisms and in vivo models. Physiol Rev 88:421–449 Dolmetsch RE, Pajvani U, Fife K, Spotts JM, Greenberg ME (2001) Signaling to the nucleus by an l-type calcium channel-calmodulin complex through the MAP kinase pathway. Science 294:333–339 Krebs J (2009) The influence of calcium signaling on the regulation of alternative splicing. Biochim Biophys Acta 1793:979–984 Wayman GA, Lee YS, Tokumitsu H, Silva AJ, Soderling TR (2008) Calmodulin-kinases: modulators of neuronal development and plasticity. Neuron 59:914–931 Saito N, Shirai Y (2002) Protein kinase C gamma (PKC gamma): function of neuron-specific isotype. J Biochem 132:683–687 Rosen LB, Ginty DD, Weber MJ, Greenberg ME (1994) Membrane depolarization and calcium influx stimulate MEK and MAP kinase via activation of Ras. Neuron 12:1207–1221 Cooper DM, Mons N, Karpen JW (1995) Adenylyl cyclases and the interaction between calcium and cAMP signalling. Nature 374:421–424 Revil T, Toutant J, Shkreta L, Garneau D, Cloutier P, Chabot B (2007) Protein kinase C-dependent control of Bcl-x alternative splicing. Mol Cell Biol 27:8431–8441 Shultz JC, Vu N, Shultz MD, Mba MU, Shapiro BA, Chalfant CE (2012) The Proto-oncogene PKCiota regulates the alternative splicing of Bcl-x pre-mRNA. Mol cancer Res 10:660–669 Zhao Y, Koebis M, Suo S, Ohno S, Ishiura S (2012) Regulation of the alternative splicing of sarcoplasmic reticulum Ca2+-ATPase1 (SERCA1) by phorbol 12-myristate 13-acetate (PMA) via a PKC pathway. Biochem Biophys Res Commun 423:212–217 Matter N, Herrlich P, Konig H (2002) Signal-dependent regulation of splicing via phosphorylation of Sam68. Nature 420:691–695 Cao W, Sohail M, Liu G, Koumbadinga GA, Lobo VG, Xie J (2011) Differential effects of PKA-controlled CaMKK2 variants on neuronal differentiation. RNA Biol 8:1061–1072 Cao W, Razanau A, Feng D, Lobo VG, Xie J (2012) Control of alternative splicing by forskolin through hnRNP K during neuronal differentiation. Nucleic Acids Res 40:8059–8071 Li H, Liu G, Yu J, Cao W, Lobo VG, Xie J (2009) In vivo selection of kinase-responsive RNA elements controlling alternative splicing. J Biol Chem 284:16191–16201 Selbert MA, Anderson KA, Huang QH, Goldstein EG, Means AR, Edelman AM (1995) Phosphorylation and activation of Ca(2+)-calmodulin-dependent protein kinase IV by Ca(2+)-calmodulin-dependent protein kinase Ia kinase: phosphorylation of threonine 196 is essential for activation. J Biol Chem 270:17616–17621 Chatila T, Anderson KA, Ho N, Means AR (1996) A unique phosphorylation-dependent mechanism for the activation of Ca2+/calmodulin-dependent protein kinase type IV/GR. J Biol Chem 271:21542–21548 Xie J, McCobb DP (1998) Control of alternative splicing of potassium channels by stress hormones. Science 280:443–446 Saito M, Nelson C, Salkoff L, Lingle CJ (1997) A cysteine-rich domain defined by a novel exon in a Slo variant in rat adrenal chromaffin cells and PC12 cells. J Biol Chem 272:11710–11717 Xie J, Black DL (2001) A CaMK IV responsive RNA element mediates depolarization-induced alternative splicing of ion channels. Nature 410:936–939 Xie J, Jan C, Stoilov P, Park J, Black DL (2005) A consensus CaMK IV-responsive RNA sequence mediates regulation of alternative exons in neurons. RNA 11:1825–1834 Yu J, Hai Y, Liu G, Fang T, Kung SK, Xie J (2009) The heterogeneous nuclear ribonucleoprotein L is an essential component in the Ca2+/calmodulin-dependent protein kinase IV-regulated alternative splicing through cytidine-adenosine repeats. J Biol Chem 284:1505–1513 An P, Grabowski PJ (2007) Exon silencing by UAGG motifs in response to neuronal excitation. PLoS Biol 5:e36 Lee JA, Xing Y, Nguyen D, Xie J, Lee CJ, Black DL (2007) Depolarization and CaM kinase IV modulate NMDA receptor splicing through two essential RNA elements. PLoS Biol 5:e40 Iijima T, Wu K, Witte H, Hanno-Iijima Y, Glatter T, Richard S, Scheiffele P (2011) SAM68 regulates neuronal activity-dependent alternative splicing of neurexin-1. Cell 147:1601–1614 Rozic G, Lupowitz Z, Piontkewitz Y, Zisapel N (2011) Dynamic changes in neurexins alternative splicing: role of Rho-associated protein kinases and relevance to memory formation. PLoS ONE 6:e18579 Rozic G, Lupowitz Z, Zisapel N (2013) Exonal elements and factors involved in the depolarization-induced alternative splicing of neurexin 2. J Mol Neurosci 50:221–233 Lee JA, Tang ZZ, Black DL (2009) An inducible change in Fox-1/A2BP1 splicing modulates the alternative splicing of downstream neuronal target exons. Genes Dev 23:2284–2293 Daoud R, Mies G, Smialowska A, Olah L, Hossmann KA, Stamm S (2002) Ischemia induces a translocation of the splicing factor Tra2-beta 1 and changes alternative splicing patterns in the brain. J Neurosci. 22:5889–5899 Janowicz A, Michalak M, Krebs J (2011) Stress-induced subcellular distribution of ALG-2, RBM22, and hSlu7. Biochim Biophys Acta 1813:1045–1049 Schor IE, Rascovan N, Pelisch F, Allo M, Kornblihtt AR (2009) Neuronal cell depolarization induces intragenic chromatin modifications affecting NCAM alternative splicing. Proc Natl Acad Sci USA 106:4325–4330 Brown SJ, Stoilov P, Xing Y (2012) Chromatin and epigenetic regulation of pre-mRNA processing. Hum Mol Genet 21:R90–R96 Lee KM, Tarn WY (2013) Coupling pre-mRNA processing to transcription on the RNA factory assembly line. RNA Biol 10:380–390 Montaville P, Dai Y, Cheung CY, Giller K, Becker S, Michalak M, Webb SE, Miller AL, Krebs J (2006) Nuclear translocation of the calcium-binding protein ALG-2 induced by the RNA-binding protein RBM22. Biochim Biophys Acta 1763:1335–1343 Park JW, Parisky K, Celotto AM, Reenan RA, Graveley BR (2004) Identification of alternative splicing regulators by RNA interference in Drosophila. Proc Natl Acad Sci USA 101:15974–15979 Krebs J, Groenendyk J, Michalak M (2011) Ca2+-signaling, alternative splicing and endoplasmic reticulum stress responses. Neurochem Res 36:1198–1211 Heyd F, Lynch KW (2011) Degrade, move, and regroup: signaling control of splicing proteins. Trends Biochem Sci 36:397–404 Wang Z, Burge CB (2008) Splicing regulation: from a parts list of regulatory elements to an integrated splicing code. RNA 14:802–813 McKee AE, Neretti N, Carvalho LE, Meyer CA, Fox EA, Brodsky AS, Silver PA (2007) Exon expression profiling reveals stimulus-mediated exon use in neural cells. Genome Biol 8:R159 Guerini D, Garcia-Martin E, Gerber A, Volbracht C, Leist M, Merino CG, Carafoli E (1999) The expression of plasma membrane Ca2+ pump isoforms in cerebellar granule neurons is modulated by Ca2+. J Biol Chem. 274:1667–1676 Zacharias DA, Strehler EE (1996) Change in plasma membrane Ca2(+)-ATPase splice-variant expression in response to a rise in intracellular Ca2+. Curr Biol 6:1642–1652 Strehler EE, Strehler-Page MA, Vogel G, Carafoli E (1989) mRNAs for plasma membrane calcium pump isoforms differing in their regulatory domain are generated by alternative splicing that involves two internal donor sites in a single exon. Proc Natl Acad Sci USA 86:6908–6912 Dean C, Scholl FG, Choih J, DeMaria S, Berger J, Isacoff E, Scheiffele P (2003) Neurexin mediates the assembly of presynaptic terminals. Nat Neurosci 6:708–716 Chih B, Gollan L, Scheiffele P (2006) Alternative splicing controls selective trans-synaptic interactions of the neuroligin-neurexin complex. Neuron 51:171–178 Penn AC, Balik A, Wozny C, Cais O, Greger IH (2012) Activity-mediated AMPA receptor remodeling, driven by alternative splicing in the ligand-binding domain. Neuron 76:503–510 Feng Y, Chen M, Manley JL (2008) Phosphorylation switches the general splicing repressor SRp38 to a sequence-specific activator. Nat Struct Mol Biol 15:1040–1048 Balik A, Penn AC, Nemoda Z, Greger IH (2013) Activity-regulated RNA editing in select neuronal subfields in hippocampus. Nucleic Acids Res 41:1124–1134 Zundorf G, Reiser G (2011) Calcium dysregulation and homeostasis of neural calcium in the molecular mechanisms of neurodegenerative diseases provide multiple targets for neuroprotection. Antioxid Redox Signal 14:1275–1288 Delorenzo RJ, Sun DA, Deshpande LS (2005) Cellular mechanisms underlying acquired epilepsy: the calcium hypothesis of the induction and maintenance of epilepsy. Pharmacol Ther 105:229–266 Bhalla K, Phillips HA, Crawford J, McKenzie OL, Mulley JC, Eyre H, Gardner AE, Kremmidiotis G, Callen DF (2004) The de novo chromosome 16 translocations of two patients with abnormal phenotypes (mental retardation and epilepsy) disrupt the A2BP1 gene. J Hum Genet 49:308–311 Barnby G, Abbott A, Sykes N, Morris A, Weeks DE, Mott R, Lamb J, Bailey AJ, Monaco AP (2005) Candidate-gene screening and association analysis at the autism-susceptibility locus on chromosome 16p: evidence of association at GRIN2A and ABAT. Am J Hum Genet 76:950–966 Martin CL, Duvall JA, Ilkin Y, Simon JS, Arreaza MG, Wilkes K, Alvarez-Retuerto A, Whichello A, Powell CM, Rao K, Cook E, Geschwind DH (2007) Cytogenetic and molecular characterization of A2BP1/FOX1 as a candidate gene for autism. Am J Med Genet B Neuropsychiatr Genet. 144B:869–876 Sebat J, Lakshmi B, Malhotra D, Troge J, Lese-Martin C, Walsh T, Yamrom B, Yoon S, Krasnitz A, Kendall J, Leotta A, Pai D, Zhang R, Lee YH, Hicks J, Spence SJ, Lee AT, Puura K, Lehtimaki T, Ledbetter D, Gregersen PK, Bregman J, Sutcliffe JS, Jobanputra V, Chung W, Warburton D, King MC, Skuse D, Geschwind DH, Gilliam TC, Ye K, Wigler M (2007) Strong association of de novo copy number mutations with autism. Science 316:445–449 Voineagu I, Wang X, Johnston P, Lowe JK, Tian Y, Horvath S, Mill J, Cantor RM, Blencowe BJ, Geschwind DH (2011) Transcriptomic analysis of autistic brain reveals convergent molecular pathology. Nature 474:380–384 Zhang Y, Vilaythong AP, Yoshor D, Noebels JL (2004) Elevated thalamic low-voltage-activated currents precede the onset of absence epilepsy in the SNAP25-deficient mouse mutant coloboma. J Neurosci 24:5239–5248 Papale LA, Beyer B, Jones JM, Sharkey LM, Tufik S, Epstein M, Letts VA, Meisler MH, Frankel WN, Escayg A (2009) Heterozygous mutations of the voltage-gated sodium channel SCN8A are associated with spike-wave discharges and absence epilepsy in mice. Hum Mol Genet 18:1633–1641 Martin MS, Tang B, Papale LA, Yu FH, Catterall WA, Escayg A (2007) The voltage-gated sodium channel Scn8a is a genetic modifier of severe myoclonic epilepsy of infancy. Hum Mol Genet 16:2892–2899 Pal S, Limbrick DD Jr, Rafiq A, DeLorenzo RJ (2000) Induction of spontaneous recurrent epileptiform discharges causes long-term changes in intracellular calcium homeostatic mechanisms. Cell Calcium 28:181–193 Pal S, Sun D, Limbrick D, Rafiq A, DeLorenzo RJ (2001) Epileptogenesis induces long-term alterations in intracellular calcium release and sequestration mechanisms in the hippocampal neuronal culture model of epilepsy. Cell Calcium 30:285–296 Sun DA, Sombati S, Blair RE, DeLorenzo RJ (2004) Long-lasting alterations in neuronal calcium homeostasis in an in vitro model of stroke-induced epilepsy. Cell Calcium 35:155–163 Sun DA, Sombati S, Blair RE, DeLorenzo RJ (2002) Calcium-dependent epileptogenesis in an in vitro model of stroke-induced “epilepsy”. Epilepsia. 43:1296–1305 DeLorenzo RJ, Pal S, Sombati S (1998) Prolonged activation of the N-methyl-d-aspartate receptor-Ca2+ transduction pathway causes spontaneous recurrent epileptiform discharges in hippocampal neurons in culture. Proc Natl Acad Sci USA 95:14482–14487 Pacheco Otalora LF, Hernandez EF, Arshadmansab MF, Francisco S, Willis M, Ermolinsky B, Zarei M, Knaus HG, Garrido-Sanabria ER (2008) Down-regulation of BK channel expression in the pilocarpine model of temporal lobe epilepsy. Brain Res 1200:116–131 Ermolinsky BS, Skinner F, Garcia I, Arshadmansab MF, Otalora LF, Zarei MM, Garrido-Sanabria ER (2011) Upregulation of STREX splice variant of the large conductance Ca2+-activated potassium (BK) channel in a rat model of mesial temporal lobe epilepsy. Neurosci Res 69:73–80 Jaekel B, Muhlberg K, Garcia de Arriba S, Reichenbach A, Verdaguer E, Pallas M, Camins A, Norenberg W, Allgaier C (2006) Neuroprotection associated with alternative splicing of NMDA receptors in rat cortical neurons. Br J Pharmacol 147:622–633 Ying Z, Babb TL, Comair YG, Bingaman W, Bushey M, Touhalisky K (1998) Induced expression of NMDAR2 proteins and differential expression of NMDAR1 splice variants in dysplastic neurons of human epileptic neocortex. J Neuropathol Exp Neurol 57:47–62 Vezzani A, Speciale C, Della Vedova F, Tamburin M, Benatti L (1995) Alternative splicing at the C-terminal but not at the N-terminal domain of the NMDA receptor NR1 is altered in the kindled hippocampus. Eur J Neurosci 7:2513–2517 Kraus JE, Nadler JV, McNamara JO (1996) Regulation of alternative splicing of NMDAR1 in the kindling model. Brain Res Mol Brain Res 41:97–104 Rujescu D, Soyka M, Dahmen N, Preuss U, Hartmann AM, Giegling I, Koller G, Bondy B, Moller HJ, Szegedi A (2005) GRIN1 locus may modify the susceptibility to seizures during alcohol withdrawal. Am J Med Genet B Neuropsychiatr Genet 133B:85–87 Witte OW, Bidmon HJ, Schiene K, Redecker C, Hagemann G (2000) Functional differentiation of multiple perilesional zones after focal cerebral ischemia. J Cereb Blood Flow Metab 20:1149–1165 Ruggiu M, Herbst R, Kim N, Jevsek M, Fak JJ, Mann MA, Fischbach G, Burden SJ, Darnell RB (2009) Rescuing Z+ agrin splicing in Nova null mice restores synapse formation and unmasks a physiologic defect in motor neuron firing. Proc Natl Acad Sci USA 106:3513–3518 Lotti F, Imlach WL, Saieva L, Beck ES, le Hao T, Li DK, Jiao W, Mentis GZ, Beattie CE, McCabe BD, Pellizzoni L (2012) An SMN-dependent U12 splicing event essential for motor circuit function. Cell 151:440–454 He H, Liyanarachchi S, Akagi K, Nagy R, Li J, Dietrich RC, Li W, Sebastian N, Wen B, Xin B, Singh J, Yan P, Alder H, Haan E, Wieczorek D, Albrecht B, Puffenberger E, Wang H, Westman JA, Padgett RA, Symer DE, de la Chapelle A (2011) Mutations in U4atac snRNA, a component of the minor spliceosome, in the developmental disorder MOPD I. Science 332:238–240 Edery P, Marcaillou C, Sahbatou M, Labalme A, Chastang J, Touraine R, Tubacher E, Senni F, Bober MB, Nampoothiri S, Jouk PS, Steichen E, Berland S, Toutain A, Wise CA, Sanlaville D, Rousseau F, Clerget-Darpoux F, Leutenegger AL (2011) Association of TALS developmental disorder with defect in minor splicing component U4atac snRNA. Science 332:240–243 Hutton M, Lendon CL, Rizzu P, Baker M, Froelich S, Houlden H, Pickering-Brown S, Chakraverty S, Isaacs A, Grover A, Hackett J, Adamson J, Lincoln S, Dickson D, Davies P, Petersen RC, Stevens M, de Graaff E, Wauters E, van Baren J, Hillebrand M, Joosse M, Kwon JM, Nowotny P, Che LK, Norton J, Morris JC, Reed LA, Trojanowski J, Basun H, Lannfelt L, Neystat M, Fahn S, Dark F, Tannenberg T, Dodd PR, Hayward N, Kwok JB, Schofield PR, Andreadis A, Snowden J, Craufurd D, Neary D, Owen F, Oostra BA, Hardy J, Goate A, van Swieten J, Mann D, Lynch T, Heutink P (1998) Association of missense and 5′-splice-site mutations in tau with the inherited dementia FTDP-17. Nature 393:702–705 Twyffels L, Gueydan C, Kruys V (2011) Shuttling SR proteins: more than splicing factors. FEBS J. 278:3246–3255 Dreyfuss G, Kim VN, Kataoka N (2002) Messenger-RNA-binding proteins and the messages they carry. Nat Rev Mol Cell Biol 3:195–205