Curr. Issues Mol. Biol. 8: 27–50. Online journal at www.cimb.org The Role of Tyrosine Phosphorylation in Regulation of Signal Transduction Pathways in Unicellular Eukaryotes Irina V. Schemarova transducer and activator of transcription) proteins located in the cytoplasm. This transmission is also provided by Sechenov Institute of Evolutionary Physiology and the SH2 domain contacts responsible for coupling of Biochemistry, Russian Academy of Sciences, St. phosphotyrosine-containing proteins (Darnell, 1997). Petersburg 194223, Russia The structural-functional organization of Ras-MAPK and STAT signaling pathways in vertebrate cells has Abstract been intensively studied for more than 20 years. By The review summarizes for the frst time the current the present time an extensive experimental material concepts of the role of tyrosine phosphorylation in has been accumulated, which allows judging about the regulation of signal transduction pathways in unicellular events in cells of the higher eukaryotes at each stage of eukaryotes. Evolutionary concepts are developed about transmission of proliferative signal. Much less is known the origin of protein tyrosine kinases (PTK)-signaling. about mechanisms of transmission of growth signals in cells of primitive eukaryotes. There are practically no data Introduction in the literature about the existence of their STAT and Ras- At the end of the last century, in mammalian cells a multi- MAPK pathways of the signal transmission; meanwhile, it cascade pathway was discovered to be responsible for is obvious that RTKs exist not only in mammals, but in transmission of proliferative signals into genome. The multicellular invertebrates, including the most primitive key role in the signal transmission through this pathway — sponges and coelenterates (Schartl, Barnekow, 1982; is played by processes of phosphorylation of protein Schartl et al., 1989; Kruse et al., 1997). tyrosine residues (Hunter, 1995; Marshall, 1995; Feig et Our analysis of experimental and literature data al., 1996; Mustelin et al., 2002). has shown that tyrosine phosphorylation also takes Ushiro and Cohen (1980) were the frst to establish place in cells of unicellular eukaryotes. The presence the important role of phosphorylation of tyrosine as a in eukaryotic microorganisms of tyrosine kinases and regulator of intracellular processes and to reveal changes proteins phosphorylated at tyrosine in processes of of tyrosine kinase activity of proteins in mammalian cells growth, cellular differentiation, and adaptive response in response to action of growth factors. Subsequently, the indicates a more ancient origin of mechanisms of PTK- change of the protein tyrosine kinase activity was shown signaling than this was believed earlier. to underlie the Ras-MAPK signaling pathway regulated This review focuses on the role of mammalian growth by Mitogen-Activated Protein (MAP) kinases (Hunter and factors, tyrosine kinases, and phosphotyrosine-containing Cooper, 1985). proteins in signaling in unicellular eukaryotes. Phosphorylation of tyrosine residues in proteins is catalyzed by retrovirus protein tyrosine kinases (PTKs), Effects of mammalian growth factors on unicellular their cellular homologues (Bishop, 1983) or by tyrosine organisms kinases associated with growth factor receptors (Yarden Due to the accepted concept that PTK-signaling takes and Ullrich, 1988; Broome and Hunter, 1996). These place only in cells of the higher eukaryotes, many aspects proteins were studied in detail and classifed in vertebrate of the problem of evolutionary origin of this signaling are animals (Hanks and Hunter, 1995). kept in the shade. Thus, for instance, nothing, in fact, is The classical scheme of transmission of the prolifera- known about evolution of mechanisms of activation of the tive signals through the pathway mediated by growth signal pathways dependent on tyrosine phosphorylation. factors (Ras-MAPK pathway) includes: (1) association of In this connection, it seems important to growth factor with receptor, (2) dimerization of receptor and consider pathways of signal transmission by tyrosine autophosphorylation of receptor tyrosine kinase (RTK), phosphorylation in unicellular eukaryotes, whose cells (3) module coupling of RTK with adaptor SH2 domain have no typical growth receptors and RTKs. proteins; activation of Ras, (4) phosphorylation and It would have been logical to suggest that unicellular activation of MAP kinases, (5) transmission of signal into eukaryotes deprived of typical receptors of mammalian genome. growth factors could not and should not respond to signals At the end of the last century, another pathway of initiated by mammalian growth factors. However, in reality, transmission of proliferative signals into genome, with this is not so. Studies performed for the last few years participation of growth factors and tyrosine kinases, was have shown that the mammalian growth factors (TGFA, discovered (Shuai et al., 1994). This is the monocascade EGF, FGF, HGF, etc.) can initiate proliferative effect not STAT pathway activated by receptors of growth factors only in mammalian cells, but also in unicellular animals. and cytokines. The essence of this transmission consists The stimulating effect of mammalian growth factors on in direct activation by tyrosine kinases of the STAT (signal proliferative growth of eukaryotic microorganisms was revealed for the frst time in the parasitic protozoa Giardia lamblia (Lujan et al, 1994). The mitogenic effect on For correspondence: [email protected] © Horizon Scientifc Press. Offprints from www.cimb.org 28 Schemarova growth of G. lamblia was produced by insulin-like factors and membrane-associated proteins that, in the authors’ IGF-I and IGF-II (Lujan et al, 1994). They were shown opinion (Gugssa et al., 2000), became involved in control to accelerate growth and L-cysteine incorporation in G. of growth and reproduction of T. musculi. lamblia trophozoites cultivated axenically in a serum- Platelet growth factor (PDGF), whose receptors free medium. The insulin-like factors act through the in mammals also have their own tyrosine kinase parasite membrane glycoprotein that has immunological activity, stimulated macromolecular synthesis in the characteristics similar to those of the mammalian IGF-I. ciliate Tetrahymena pyriformis (Andersen et al., 1984). In reaction of immunoprecipitation of lysate of G. lamblia Measurements of changes in the labeled uridine specifc trophozoites incubated with human recombinant IGF-II, activity in the UTP pool after PDGF treatment showed an with monoclonal antibody AIR3, proteins of a molecular initial increase lasting for about 40 min. Thereafter the mass of 70 kDa were found; one of them, as shown by RNA synthesis rate gradually decreased and became further studies using polyclonal antibody to the IGF- the same as in intact cells after 2 hr of incubation. PDGF I receptor, corresponded to the IGF-I receptor A-chain. also stimulated the labeled uridine incorporation in T. Protein of a molecular mass of 95 kDa that corresponded pyriformis. The increase begins shortly after addition of to the receptor IGF-I B-chain was discovered using this growth factor and levels off 2–3 h later. The data polyclonal antibody to phosphotyrosine. Based on the obtained by these authors (Andersen et al., 1984) rule obtained data, the authors suggested the existence in out a possibility of induction of changes in the nucleotide the membrane glycoproteins of G. lamblia trophozoites pool of precursors both of RNA and of DNA by PDGF in T. of internal tyrosine kinase activity and the capability for pyriformis. The authors suggested T. pyriformis to contain autophosphorylation. receptors on its surface for the growth factor. The promoter effect of IGF-I on cell growth was also Colony-stimulating growth factor (CSF) from revealed in cells of the parasitic protozoa Leishmania granulocytes stimulated proliferative response and mexicana (Gomes et al., 1998; 2001). The authors inhibited the heat-shock-induced apoptosis in Leishmania explain the stimulating IGF-I effect on leishmanian cells mexicana amazonensis (Charlab et al., 1990; Barcinski by a similarity of structure in receptors of leishmanian and et al., 1992; Welburn et al., 1996). Some mammalian mammalian cells. The receptor for IGF-I on L. mexicana cytokines (IL-3, IL-6, TNF, and IFN-G) are reported to promastigotes is a monomeric glycoprotein of a molecular induce in protozoan parasites a cellular reaction aimed at mass of 65 kDa and is antigenically related to the A-chain maintaining their viability (Olsson et al, 1992; Barcinski, of the human type 1 IGF-I receptor. Upon IGF-I stimulation Moreira, 1994; Csaba et al., 1995, Salotra et al., 1995; the receptor undergoes autophosphorylation at tyrosine Bakhiet et al., 1996). residues with activation of its signaling pathway. Activation Several works have demonstrated stimulation of of the IGF-I receptor also leads to phosphorylation of a proliferation of protozoa by EGF (Hide et al., 1989; 185-kDa molecule that is homologous to the substrate Sternberg, McGuigan, 1994; Selivanova et al., 2002; of the insulin receptor present in human cells, the insulin Shemarova et al., 2004). It was established that EGF receptor substrate 1 (IRS-1) (Gomes et al., 2001). produced a marked proliferative response in vivo in By the example of free-living ciliates Tetrahymena trypomastigotes of the Trypanosoma brucei strain Tc221 pyriformis and Tetrahymena thermophila, the vertebrate (Sternberg and McGuigan,