Tài liệu Báo cáo khoa học: Association of mammalian sterile twenty kinases, Mst1 and Mst2, with

Association of mammalian sterile twenty kinases, Mst1

and Mst2, with hSalvador via C-terminal coiled-coil

domains, leads to its stabilization and phosphorylation

Bernard A. Callus1,*, Anne M. Verhagen1 and David L. Vaux1,*

1 The Walter and Eliza Hall Institute, Parkville, VC, Australia

The mammalian serine ⁄threonine kinases, Mst1 and

Mst2, were originally identified by their similarity to

yeast Sterile Twenty (Ste20) kinase [1,2]. Mst3 and

Mst4 were subsequently identified the same way [3–5].

The four Mst kinases belong to a subfamily of Ste20-

like germinal center kinases (GCKs) that is character￾ized by an N-terminal kinase domain (reviewed in [6]).

Based on their similarity with each other, the Mst kin￾ases can be further subdivided into two groups, Mst1

and Mst2 (GCKII) and Mst3 and Mst4 (GCKIII).

Mst1 has been widely studied and is the best charac￾terized member of the family. In addition to its kinase

domain, Mst1 contains an inhibitory domain, deletion

of which results in increased kinase activity, and a pre￾dicted coiled-coil domain at the C-terminus that is

essential for the formation of Mst1 dimers (multimers)

[7]. Full-length Mst1 is mainly cytoplasmic, but can

shuttle continuously between the cytoplasm and nuc￾leus in a phosphorylation dependent manner [8–10].

Ectopic expression of Mst1 and Mst2 in certain cells

types has been reported to induce cell death in a stress

activated protein kinase (SAPK) dependent pathway

[11–13]. In apoptotic cells, activated caspases can

cleave Mst1 and Mst2 C-terminal to the kinase domain

Keywords

coiled-coil domain; dimerization; Mst kinase;

phosphorylation; Salvador

Correspondence

B. A. Callus, Department of Biochemistry,

La Trobe University, Plenty Road, Bundoora,

VIC 3086, Australia

Fax: +613 9479 2467

Tel: +613 9479 1669

E-mail: [email protected]

*Present address

Department of Biochemistry, La Trobe Uni￾versity, Plenty Road, Bundoora, VIC 3086,

Australia

(Received 21 May 2006, accepted 18 July

2006)

doi:10.1111/j.1742-4658.2006.05427.x

Genetic screens in Drosophila have revealed that the serine ⁄threonine kinase

Hippo (Hpo) and the scaffold protein Salvador participate in a pathway

that controls cell proliferation and apoptosis. Hpo most closely resembles

the pro-apoptotic mammalian sterile20 kinases 1 and 2 (Mst1 and 2), and

Salvador (Sav) has a human orthologue hSav (also called hWW45). Here

we show that Mst and hSav heterodimerize in an interaction requiring the

conserved C-terminal coiled-coil domains of both proteins. hSav was also

able to homodimerize, but this did not require its coiled-coil domain. Coex￾pression of Mst and hSav led to phosphorylation of hSav and also increased

its abundance. In vitro phosphorylation experiments indicate that the phos￾phorylation of Sav by Mst is direct. The stabilizing effect of Mst was much

greater on N-terminally truncated hSav mutants, as long as they retained

the ability to bind Mst. Mst mutants that lacked the C-terminal coiled-coil

domain and were unable to bind to hSav, also failed to stabilize or phos￾phorylate hSav, whereas catalytically inactive Mst mutants that retained the

ability to bind to hSav were still able to increase its abundance, although

they were no longer able to phosphorylate hSav. Together these results

show that hSav can bind to, and be phosphorylated by, Mst, and that the

stabilizing effect of Mst on hSav requires its interaction with hSav but is

probably not due to phosphorylation of hSav by Mst.

Abbreviations

dIAP1, Drosophila inhibitor of apoptosis I; GCK, germinal center kinase; HA, hemagglutinin; Hpo, serine/threonine kinase Hippo; IL-2,

interleukin-2; Lats, large tumour suppressor; MBP, myelin basic protein; Mst, mammalian sterile20 kinase; Nore1, novel Ras effector 1;

PBST, NaCl ⁄ Pi-Tween 20; PPIA, peptidyl-prolyl cis-trans isomerase A; Rassf1, Ras suppressor factor 1; Sav, Salvador; SAPK, stress

activated protein kinase; Ste20, Sterile Twenty; Wts, warts kinase; Yki, Yorkie.

4264 FEBS Journal 273 (2006) 4264–4276 ª 2006 The Authors Journal compilation ª 2006 FEBS