Phosphorylation of the novel mTOR substrate Unkempt regulates cellular morphogenesis

Baskaran, Pranetha, Mihaylov, Simeon R., Vinsland, Elin, Shah, Kriti, Granat, Lucy, Ultanir, Sila K., Tee, Andrew R. ORCID: https://orcid.org/0000-0002-5577-4631, Murn, Jernej and Bateman, Joseph M. 2023. Phosphorylation of the novel mTOR substrate Unkempt regulates cellular morphogenesis. Journal of Biological Chemistry 299 (1) , 102788. 10.1016/j.jbc.2022.102788

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Abstract

Mechanistic target of rapamycin (mTOR) is a protein kinase that integrates multiple inputs to regulate anabolic cellular processes. For example, mTOR complex 1 (mTORC1) has key functions in growth control, autophagy, and metabolism. However, much less is known about the signaling components that act downstream of mTORC1 to regulate cellular morphogenesis. Here, we show that the RNA-binding protein Unkempt, a key regulator of cellular morphogenesis, is a novel substrate of mTORC1. We show that Unkempt phosphorylation is regulated by nutrient levels and growth factors via mTORC1. To analyze Unkempt phosphorylation, we immunoprecipitated Unkempt from cells in the presence or the absence of the mTORC1 inhibitor rapamycin and used mass spectrometry to identify mTORC1-dependent phosphorylated residues. This analysis showed that mTORC1-dependent phosphorylation is concentrated in a serine-rich intrinsically disordered region in the C-terminal half of Unkempt. We also found that Unkempt physically interacts with and is directly phosphorylated by mTORC1 through binding to the regulatory-associated protein of mTOR, Raptor. Furthermore, analysis in the developing brain of mice lacking TSC1 expression showed that phosphorylation of Unkempt is mTORC1 dependent in vivo. Finally, mutation analysis of key serine/threonine residues in the serine-rich region indicates that phosphorylation inhibits the ability of Unkempt to induce a bipolar morphology. Phosphorylation within this serine-rich region thus profoundly affects the ability of Unkempt to regulate cellular morphogenesis. Taken together, our findings reveal a novel molecular link between mTORC1 signaling and cellular morphogenesis.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Medicine
Publisher:	Elsevier
ISSN:	0021-9258
Date of First Compliant Deposit:	15 February 2023
Date of Acceptance:	24 November 2022
Last Modified:	13 Jun 2023 15:43
URI:	https://orca.cardiff.ac.uk/id/eprint/157028

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