Actin-binding and cell proliferation activities of angiomotin family members are regulated by Hippo pathway-mediated phosphorylation.
Whether the Hippo pathway has downstream targets other than YAP and TAZ is unknown. In this report, we have identified angiomotin (Amot) family members as novel substrates of Hippo core kinases. The N-terminal regions of Amot proteins contain a conserved HXRXXS consensus site for LATS1/2-mediated phosphorylation. Phospho-specific antibodies showed that Hippo core kinases could mediate phosphorylation of endogenous as well as exogenous Amot family members. Knockdown of LATS1 and LATS2 endogenously reduced the phosphorylation of Amots detected by the phospho-specific antibodies. Mutation of the serine to alanine within this HXRXXS site in Amot and AmotL2 established that this site was essential for Hippo core kinase-mediated phosphorylation. Wild-type and non-phosphorylated Amot (Amot-S175A) were targeted to actin filaments, whereas phospho-mimic Amot (Amot-S175D) failed to be localized with actin. Overexpression of LATS2 caused dissociation of Amot from actin but not Amot-S175A. Mapping of the actin-binding site of Amot showed that serine 175 of Amot was important for the actin-binding activity. Amot-S175A promoted, whereas Amot and Amot-S175D inhibited, cell proliferation. These results collectively suggest that the Hippo pathway negatively regulates the actin-binding activity of Amot family members through direct phosphorylation.
From the Institute of Molecular and Cell Biology, 61 Biopolis Drive, Singapore 138673 and.
Siew Wee
SW
Chan
Chun Jye
CJ
Lim
Fusheng
F
Guo
Ivan
I
Tan
Thomas
T
Leung
Wanjin
W
Hong
D000199:Actins / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution; D000091082:Angiomotins; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation; D004285:Dogs; D057809:HEK293 Cells; D000090683:Hippo Signaling Pathway; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D061985:Madin Darby Canine Kidney Cells; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D020125:Mutation, Missense; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*
C423985:AMOT protein, human; C508566:AMOTL2 protein, human; D000199:Actins; D000091082:Angiomotins; D002352:Carrier Proteins; D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D025521:Tumor Suppressor Proteins; C092745:LATS1 protein, human; C406449:LATS2 protein, human; D017346:Protein Serine-Threonine Kinases
Actin; Angiomotin; Cell Proliferation; Hippo Pathway; LATS Kinase; Phosphorylation; Protein Kinases; Signal Transduction