title: Constitutively active NDR1-PIF kinase functions independent of MST1 and hMOB1 signalling.
abstract: The human MST1/hMOB1/NDR1 tumour suppressor cascade regulates important cellular processes, such as centrosome duplication. hMOB1/NDR1 complex formation appears to be essential for NDR1 activation by autophosphorylation on Ser281 and hydrophobic motif (HM) phosphorylation at Thr444 by MST1. To dissect these mechanistic relationships in MST1/hMOB1/NDR signalling, we designed NDR1 variants carrying modifications that mimic HM phosphorylation and/or abolish hMOB1/NDR1 interactions. Significantly, the analyses of these variants revealed that NDR1-PIF, an NDR1 variant containing the PRK2 hydrophobic motif, remains hyperactive independent of hMOB1/NDR1-PIF complex formation. In contrast, as reported for the T444A phospho-acceptor mutant, NDR1 versions carrying single phospho-mimicking mutations at the HM phosphorylation site, namely T444D or T444E, do not display increased kinase activities. Collectively, these observations suggest that in cells Thr444 phosphorylation by MST1 depends on the hMOB1/NDR1 association, while Ser281 autophosphorylation of NDR1 can occur independently. By testing centrosome-targeted NDR1 variants in NDR1- or MST1-depleted cells, we further observed that centrosome-enriched NDR1-PIF requires neither hMOB1 binding nor MST1 signalling to function in centrosome overduplication. Taken together, our biochemical and cell biological characterisation of NDR1 versions provides novel unexpected insights into the regulatory mechanisms of NDR1 and NDR1's role in centrosome duplication.
authors.affiliation: Tumour Suppressor Signalling Networks laboratory, UCL Cancer Institute, University College London, WC1E 6BT London, United Kingdom.
authors.forename: Dorthe
authors.initials: D
authors.lastname: Cook
authors.affiliation: Tumour Suppressor Signalling Networks laboratory, UCL Cancer Institute, University College London, WC1E 6BT London, United Kingdom.
authors.forename: Lily Y
authors.initials: LY
authors.lastname: Hoa
authors.affiliation: Tumour Suppressor Signalling Networks laboratory, UCL Cancer Institute, University College London, WC1E 6BT London, United Kingdom.
authors.forename: Valenti
authors.initials: V
authors.lastname: Gomez
authors.affiliation: Tumour Suppressor Signalling Networks laboratory, UCL Cancer Institute, University College London, WC1E 6BT London, United Kingdom.
authors.forename: Marta
authors.initials: M
authors.lastname: Gomez
authors.affiliation: Tumour Suppressor Signalling Networks laboratory, UCL Cancer Institute, University College London, WC1E 6BT London, United Kingdom. Electronic address: a.hergovich@ucl.ac.uk.
authors.forename: Alexander
authors.initials: A
authors.lastname: Hergovich
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D018385:Centrosome / Q000378:metabolism; D002522:Chlorocebus aethiops; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011509:Proteoglycans / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction; D000099259:N-myc Downstream-Regulated Gene 1 Protein; D000099317:Hippo Kinases
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D047908:Intracellular Signaling Peptides and Proteins; D017346:Protein Serine-Threonine Kinases; D011509:Proteoglycans; D034741:RNA, Small Interfering; D000099259:N-myc Downstream-Regulated Gene 1 Protein; C042057:proteolysis-inducing peptide; C095566:STK4 protein, human; D000099317:Hippo Kinases
keywords: Centrosome duplication; Hydrophobic motif phosphorylation; Intracellular kinase signalling; MOB proteins; MST1/STK4; NDR1/STK38
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