Difference between revisions of "Kinase Family SCYL"

Revision as of 00:30, 20 February 2012

Kinase Classification: Group Other: Family SCYL

SCYL is a family of inactive kinases involved in Golgi trafficking and nuclear tRNA export. This family was also previously known as the SCY1 family.

Classification and Evolution

SCYL kinases are found in almost all eukaryotes examined (lost in kinetoplastids and severely obligate parasites). The family is named after the SCY1 gene of yeast (SCYL = SCY1-Like). There are three subfamilies, of which SCYL2 is found throughout eukaryotes, SCYL1 in plants and unikonts (animals, most fungi, Dictyostelium) and SCYL3 is found in most eumetazoans.

Domain Structure

All SCYL have an N-terminal kinase domain and a longer C-terminal region which constitutes a Pfam-B domain (Pfam-B_17727) that frequently also appears as an array of HEAT repeats, which are known to be involved in cytoskeletal interactions.

Functions

Protein Trafficking

SCYL1 binds COP-I vesicles that mediate retrograde Golgi-to ER transport, through an SCYL1-specific RKLD motif at the extreme C terminus [1]. Knockdown os SCYL1 disrupts Golgi morphology and blocks retrograte COPI-mediated transport from Golgi to ER [2]. The Gorab protein (aka NTKL-BP1, SCYL-BP1) was found as a interactor of mouse Scyl1 by Y2H and coIP [3]. Gorab is a member of the golgin family, localized to the Golgi.

=SCYL1 mutations

The mdf mouse is a model for neuromuscular atrophy. This defect was mapped to a mutation in Scyl1 [4], correlating with the high expression of Scyl1 in neurons, neuromuscular junctions and synapses.

Human SCYL2 (aka CVAK104) is a coated vesicle associated (CVA) protein which binds clathrin and the plasma membrane adaptor complex, AP2 [5].

Scyl1 and Scyl2 both bound AP2 adaptor complex:

One protein that we find in α- and β-appendage pull-downs from HeLa, liver and brain extracts is Scy1-like1. This is a distant homologue of Scy1-like2 which is found only in α-appendage pull-downs (Figure 1C). There is 16% amino acid identity between Scy1-like1 and 2. Scy1-like2 was previously found in CCVs and was thus renamed as clathrin-coated vesicle associated kinase of 104kDa (CVAK104)[16]. Scy1-like1 contains multiple DxF motifs of the type that bind to the top site of the α-appendage and thus is likely to function in CCV formation. To retain nomenclature we call it CVAK90 (as the longest human splice form is predicted to be 90kDa). We verified a direct interaction of CVAK104 and CVAK90 with both the α- and β-appendages using yeast-2-hybrid (unpublished data).

SCY1 in yeast is poorly studied, but has been implicated in sterol transport from the cell surface to the ER [6].

yata/CG1973, the Drosophila SCYL1 gene, interacts genetically with APPL, the A-beta amyloid precursor protein [7]. yata mutants showed reduced lifespan, small brains and eye vacuolization. Overexpression of APPL could partially rescue these phenotypes, and double mutants had stronger phenotypes. APPL was mislocalized in yata mutants. RNAi screens found yata to be involved in cell size regulation [8].

Centrosomes and Telomeres

One splice isoform of human SCYL1 (aka NTKL) is found at the centrosomes during mitosis [9]. SCYL1 is also named TEIF (Telomerase transcriptional Elements Interacting Factor) due to its ability to bind DNA and transactivate the hTERT telomerase and DNA polymerase beta genes [10, 11]. SCYL1 levels correlated with centrosomal amplification in cancers, and manipulation of SCYL1 caused centrosome abnormalities [12].

tRNA Export

Human SCYL1 functions in nuclear export of tRNAs [13]. It binds tRNAs and interacts with the nuclear pore through Nup98, and copurifies with a complex of exportin-t (XPOT) and exportin-5, RanGTP, and eEF-1A which transports aminoacyl-tRNAs to the ribosomes. Arabidopsis SCYL1 (At2g40730, CTEXP) was also shown to be involved in tRNA export. It binds tRNAs, RanGTP, the exportin-t (PAUSED), and associates with the nuclear pore (Johnstone et al, http://www.nrcresearchpress.com/doi/abs/10.1139/B10-090, doi:10.1139/B10-090). In both human and Arabidopsis, the Ran association is GTP-dependent.

SYCL3 functions

SCYL3 (PACE-1) has a conserved N-terminal myristoylation motif. Human SCYL3 is found in two subcellular locations: on the cytoplasmic face of the Golgi apparatus, dependent on the myristoylation motif, and in lamellipodia, where it may associate with ezrin, a cytoskeletal linker protein. The ezrin association was found by Y2H screening, and maps to the C-terminal regions of both proteins [14]. Drosophila SCYL3 (CG1344) was found in an RNAi screen to have spindle abnormalities [15]

BioGrid SCYL1 - GoRAB (Dynactin1) [only one interactor back] SCYL2 - NOP56 (nucleolar protein 5A) (83 interactors, incluing large chunks of ribosome (maybe all!), nuclear transport, topoisomerase (has pat1 domain,like some scyl), tubulins SCYL3 - ezrin CG1344 = CG17003 (tubulin n-acetyl transferase) W07G4.3 - GST-30, C35A5.8 (XPO7 exportin gene-Hs) (Exportin 7 defines a novel general nuclear export pathway.) SCY1 - 19 interactors including nuclear and RNA binding proteins, CCT chaperone, ER-golgi-trnasport, ubiquitin, sterol, pdi - lots of suspicious terms

Regulation and Activity

All SCYL proteins appear to be pseudokinases, as they have lost all three main catalytic residues, K72, D168 and D184, though the changes are conserved (K72F, D168N and D184G for all three human proteins) [16]. Human SCYL2 was shown to bind ATP and auto- and trans-phosphorylate in vitro in one study [5]. However, tagged SCYL3 constructs also showed in vitro kinase activity [14], but this was associated with the non-catalytic C-terminus, required an N-terminal myristoylation motif and could be eliminated with stringent purification methods, suggesting that SCYL3 (and maybe other SCYL) can bind an active kinase rather than having intrinsic kinase activity. All three human vertebrate SCYL have several phosphorylation sites, and SYCL3 has a T-153 site that is within the putative activation loop, but no upstream kinases are known. Yeast SCY1 also has two phosphorylation sites in the C-terminus (http://www.phosphogrid.org/sites/33167).

References

1. Burman JL, Hamlin JN, and McPherson PS. Scyl1 regulates Golgi morphology. PLoS One. 2010 Mar 4;5(3):e9537. DOI:10.1371/journal.pone.0009537 | PubMed ID:20209057 | HubMed [Burman]
2. Burman JL, Bourbonniere L, Philie J, Stroh T, Dejgaard SY, Presley JF, and McPherson PS. Scyl1, mutated in a recessive form of spinocerebellar neurodegeneration, regulates COPI-mediated retrograde traffic. J Biol Chem. 2008 Aug 15;283(33):22774-86. DOI:10.1074/jbc.M801869200 | PubMed ID:18556652 | HubMed [Burman2]
3. Di Y, Li J, Fang J, Xu Z, He X, Zhang F, Ling J, Li X, Xu D, Li L, Li YY, and Huo K. Cloning and characterization of a novel gene which encodes a protein interacting with the mitosis-associated kinase-like protein NTKL. J Hum Genet. 2003;48(6):315-321. DOI:10.1007/s10038-003-0031-5 | PubMed ID:12783284 | HubMed [Di]
4. Burman JL, Bourbonniere L, Philie J, Stroh T, Dejgaard SY, Presley JF, and McPherson PS. Scyl1, mutated in a recessive form of spinocerebellar neurodegeneration, regulates COPI-mediated retrograde traffic. J Biol Chem. 2008 Aug 15;283(33):22774-86. DOI:10.1074/jbc.M801869200 | PubMed ID:18556652 | HubMed [Schmidt]
5. Conner SD and Schmid SL. CVAK104 is a novel poly-L-lysine-stimulated kinase that targets the beta2-subunit of AP2. J Biol Chem. 2005 Jun 3;280(22):21539-44. DOI:10.1074/jbc.M502462200 | PubMed ID:15809293 | HubMed [Conner]
6. Sullivan DP, Georgiev A, and Menon AK. Tritium suicide selection identifies proteins involved in the uptake and intracellular transport of sterols in Saccharomyces cerevisiae. Eukaryot Cell. 2009 Feb;8(2):161-9. DOI:10.1128/EC.00135-08 | PubMed ID:19060182 | HubMed [Sullivan2]
7. Sone M, Uchida A, Komatsu A, Suzuki E, Ibuki I, Asada M, Shiwaku H, Tamura T, Hoshino M, Okazawa H, and Nabeshima Y. Loss of yata, a novel gene regulating the subcellular localization of APPL, induces deterioration of neural tissues and lifespan shortening. PLoS One. 2009;4(2):e4466. DOI:10.1371/journal.pone.0004466 | PubMed ID:19209226 | HubMed [Sone]
9. Kato M, Yano K, Morotomi-Yano K, Saito H, and Miki Y. Identification and characterization of the human protein kinase-like gene NTKL: mitosis-specific centrosomal localization of an alternatively spliced isoform. Genomics. 2002 Jun;79(6):760-7. DOI:10.1006/geno.2002.6774 | PubMed ID:12036289 | HubMed [Kato]
10. Gong Y, Sun Y, McNutt MA, Sun Q, Hou L, Liu H, Shen Q, Ling Y, Chi Y, and Zhang B. Localization of TEIF in the centrosome and its functional association with centrosome amplification in DNA damage, telomere dysfunction and human cancers. Oncogene. 2009 Mar 26;28(12):1549-60. DOI:10.1038/onc.2008.503 | PubMed ID:19198626 | HubMed [Tang]
11. Zhao Y, Zheng J, Ling Y, Hou L, and Zhang B. Transcriptional upregulation of DNA polymerase beta by TEIF. Biochem Biophys Res Commun. 2005 Aug 5;333(3):908-16. DOI:10.1016/j.bbrc.2005.05.172 | PubMed ID:15963946 | HubMed [Zhao]
12. Gong Y, Sun Y, McNutt MA, Sun Q, Hou L, Liu H, Shen Q, Ling Y, Chi Y, and Zhang B. Localization of TEIF in the centrosome and its functional association with centrosome amplification in DNA damage, telomere dysfunction and human cancers. Oncogene. 2009 Mar 26;28(12):1549-60. DOI:10.1038/onc.2008.503 | PubMed ID:19198626 | HubMed [Gong]
13. Chafe SC and Mangroo D. Scyl1 facilitates nuclear tRNA export in mammalian cells by acting at the nuclear pore complex. Mol Biol Cell. 2010 Jul 15;21(14):2483-99. DOI:10.1091/mbc.e10-03-0176 | PubMed ID:20505071 | HubMed [Schafe]
14. Sullivan A, Uff CR, Isacke CM, and Thorne RF. PACE-1, a novel protein that interacts with the C-terminal domain of ezrin. Exp Cell Res. 2003 Apr 1;284(2):224-38. DOI:10.1016/s0014-4827(02)00054-x | PubMed ID:12651155 | HubMed [Sullivan]
15. Bettencourt-Dias M, Giet R, Sinka R, Mazumdar A, Lock WG, Balloux F, Zafiropoulos PJ, Yamaguchi S, Winter S, Carthew RW, Cooper M, Jones D, Frenz L, and Glover DM. Genome-wide survey of protein kinases required for cell cycle progression. Nature. 2004 Dec 23;432(7020):980-7. DOI:10.1038/nature03160 | PubMed ID:15616552 | HubMed [Bettencourt-Dias]
16. Scheeff ED, Eswaran J, Bunkoczi G, Knapp S, and Manning G. Structure of the pseudokinase VRK3 reveals a degraded catalytic site, a highly conserved kinase fold, and a putative regulatory binding site. Structure. 2009 Jan 14;17(1):128-38. DOI:10.1016/j.str.2008.10.018 | PubMed ID:19141289 | HubMed [Scheeff]
17. Johnson, AD, Mullen, RT, Mangroo, D. Arabidopsis At2g40730 encodes a cytoplasmic protein involved in nuclear tRNA export. Botany, 2011, 89:(3) 175-190. http://dx.doi.org/doi:10.1139/B10-090 [Johnstone]

1. Bjorklund pmid=16496002