Tyrosine Phosphorylation in Dictyostelium

Phosphotyrosine signaling in Dictyostelium

Dictyostelium has extensive pTyr-based signaling, despite the lack of TK-group tyrosine kinases. Conservation of some components and pathways with those of humans suggests that pTyr-based signaling preceded animal multicellularity and the invention of TKs, and was originally mediated by dual-specificity TKL kinases that were later superseded by TKs (See also: Tyrosine Kinase Evolution).

TKL kinases and pTyr

Dictyostelium has 66 TKL kinases, most of which do not fall into known metazoan families. These include 12 receptor kinases and six TKLs are known to phosphorylate tyrosine (ZakA and Dypk2-4 phosphorylate tyrosine, SplA and Shk1 are dual specificity [1]). Four other TKLs (Shk2-4) are fused to SH2 domains, and are paralogs of the Shk1 dual-specificity kinase, suggesting that these may also phosphorylate tyrosine.

A more recent report indicates that rk3 (VSK3) is a tyrosine specific receptor kinase, based on in vitro peptide activity, though it is expressed on internal vesicles rather than the plasma membrane [2]. This suggests that its paralogs, rk1-2, may also have TK activity. All three have a W in place of the Y in the YmAPE motif. Y is highly conserved across ser/thr kinases, but is almost always a W in TK-group kinases. The catalytic loop in this family is HRDLKSHN, more typical of Ser/Thr kinases. Shk1 and Shk4 also show this Y to W change, as do several other Dictyostelium TKL kinases. None of these kinases has an activation loop sequence that would suggest autophosphorylation on tyrosine.

SH2 proteins in Dictyostelium

In addition to these 5 TKLs, another 7 Dictyostelium proteins have SH2 domains, including 4 STAT proteins, a c-Cbl ortholog (both are orthologs of the human genes that are regulated by tyrosine phosphorylation) and two other proteins (LrrB is an LRR protein and FbxB is an F-box and ankyrin repeat containing protein; neither is homologous to any other published SH2-containing protein). Two of the STATs are known to be tyrosine phosphorylated, by DPYK family TKL kinases splB (Pyk2) and Pyk3 [3, 4] and one conserves the tyrosine residue known in human to be a phosphosite. Curiously, Pyk2 also contains a regulatory pseudokinase domain, just like the metazoan Jaks kinases that also phosphorylate STAT.

The SH2 domain of Shk1 kinase mediates its membrane localization, suggesting the presence of membrane-associated phosphotyrosine and receptor tyrosine kinases, and maybe reminiscent of Src and other receptor-associated kinases in metazoans.

pTyr conservation between Dictyostelium and mammals

GSKA in Dictyostelium is regulated by ZakA through tyrosine phosphorylation on the activation loop Tyr214 and probably Tyr220 (the equivalent of the autophosphorylation site Tyr216 in human GSK3b; this phosphorylation happens during translation of the GSK3 protein, and so is not regulated). This phosphorylation is driven by the presence of extracellular cAMP hormone. GSK3 in turn phosphorylates STATa, driving its nuclear export [5]. Another STAT, STATc, is activated by tyrosine phosphorylation during development, and dephosphorylated by the PTP, PTP3. That dephosphorylation is in turn blocked by CblA, the c-Cbl homolog [6] , thus tying several Ptyr-associated proteins together in developmental signaling cascades.

The tyrosine phosphorylation of GSK3 and STAT, the presence of SH2 domains and their association with putative tyrosine- or dual-specificity kinases and the involvement of Cbl in pTyr pathways are all conserved from Dictyostelium to human, suggesting that eukaryotic tyrosine phosphorylation pathways emerged using TKLs, and that TKLs were replaced by TKs in metazoans.

Dictyostelium has 4 PTP phosphatases and members of several other phosphatase families. It does not have any clear pTyr-binding PTB domains, though it has two Talin genes that contain FERM domains which in turn have a sub-domain that is similar to PTB.

References

1. Goldberg JM, Manning G, Liu A, Fey P, Pilcher KE, Xu Y, and Smith JL. The dictyostelium kinome--analysis of the protein kinases from a simple model organism. PLoS Genet. 2006 Mar;2(3):e38. DOI:10.1371/journal.pgen.0020038 |
2. Fang J, Brzostowski JA, Ou S, Isik N, Nair V, and Jin T. A vesicle surface tyrosine kinase regulates phagosome maturation. J Cell Biol. 2007 Jul 30;178(3):411-23. DOI:10.1083/jcb.200701023 |
3. Araki T, Kawata T, and Williams JG. Identification of the kinase that activates a nonmetazoan STAT gives insights into the evolution of phosphotyrosine-SH2 domain signaling. Proc Natl Acad Sci U S A. 2012 Jul 10;109(28):E1931-7. DOI:10.1073/pnas.1202715109 |
4. Araki T, Vu LH, Sasaki N, Kawata T, Eichinger L, and Williams JG. Two Dictyostelium tyrosine kinase-like kinases function in parallel, stress-induced STAT activation pathways. Mol Biol Cell. 2014 Oct 15;25(20):3222-33. DOI:10.1091/mbc.E14-07-1182 |
5. Ginger RS, Dalton EC, Ryves WJ, Fukuzawa M, Williams JG, and Harwood AJ. Glycogen synthase kinase-3 enhances nuclear export of a Dictyostelium STAT protein. EMBO J. 2000 Oct 16;19(20):5483-91. DOI:10.1093/emboj/19.20.5483 |
6. Langenick J, Araki T, Yamada Y, and Williams JG. A Dictyostelium homologue of the metazoan Cbl proteins regulates STAT signalling. J Cell Sci. 2008 Nov 1;121(Pt 21):3524-30. DOI:10.1242/jcs.036798 |