New mechanistic insights into the RAS-SIN1 interaction at the membrane

Abstract

Stress-activated MAP kinase-interacting protein 1 (SIN1) is a central member of the mTORC2 complex that contains an N-terminal domain (NTD), a conserved region in the middle (CRIM), a RAS-binding domain (RBD), and a pleckstrin homology domain. Recent studies provided valuable structural and functional insights into the interactions of SIN1 and the RAS-binding domain of RAS proteins. However, the mechanism for a reciprocal interaction of the RBD-PH tandem with RAS proteins and the membrane as an upstream event to spatiotemporal mTORC2 regulation is not clear. The biochemical assays in this study led to the following results: 1) all classical RAS paralogs, including HRAS, KRAS4A, KRAS4B, and NRAS, can bind to SIN1-RBD in biophysical and SIN1 full length (FL) in cell biology experiments; 2) the SIN1-PH domain modulates interactions with various types of membrane phosphoinositides and constantly maintains a pool of SIN1 at the membrane; and 3) a KRAS4A-dependent decrease in membrane binding of the SIN1-RBD-PH tandem was observed, suggesting for the first time a mechanistic influence of KRAS4A on SIN1 membrane association. Our study strengthens the current mechanistic understanding of SIN1-RAS interaction and suggests membrane interaction as a key event in the control of mTORC2-dependent and mTORC2-independent SIN1 function.

The allosteric mechanism of mTOR activation can inform bitopic inhibitor optimization

“…Binding of mSIN1 Ras binding domain (RBD) to H-Ras and K-Ras can dampen MAPK signaling. 66,97,98 DEP domaincontaining mTOR interacting protein (DEPTOR) operates as an endogenous mTOR inhibitor. DEPTOR binding to the mTOR FAT domain induces an inactive conformation, partially inhibiting mTORC1 and mTORC2.…”

“…The obligate cellular localization and substrate recruitment of mTORCs rely on their components. 9,66 X-ray and cryo-EM structures of mTORC1 and mTORC2 provided structural insights into their regulation, 2,4,14,53,67–69 and studies largely focused on mTORCs' ability to sense substrate signals. For both complexes, mTOR is the catalytic core.…”

The mTORC2 signaling network: targets and cross-talks

“…SIN1-PH domain mutations increase cellular oncogenicity by preventing mTOR inhibition, highlighting the role of this domain in mTORC2 activation [ 49 ]. In addition to PIP3 binding, the SIN1-PH also binds to several types of membrane phosphoinositides in vitro [ 132 ]. Furthermore, based on sequence analysis, rictor is also predicted to harbor PH domains and may thus also modulate mTORC2 membrane localization [ 38 ].…”

Unmasking the tumourigenic role of SIN1/MAPKAP1 in the mTOR complex 2

“…In tumoural cells, oncogenic RAS binds to the SIN1‐RBD and this interaction not only promotes the kinase activity of the mTORC2 at the plasma membrane but also activates the pro‐proliferative cell cycle transcription program of RAS 86–88 . Moreover, the crystal structure of the RBD‐SIN1 and RAS interaction has been published allowing the characterization of the amino acids important for this interaction and opening the way to peptide inhibition 29,30,46–47,89 …”

“…[86][87][88] Moreover, the crystal structure of the RBD-SIN1 and RAS interaction has been published allowing the characterization of the amino acids important for this interaction and opening the way to peptide inhibition. 29,30,[46][47]89 Cell-penetrating peptides (CPPs) are short peptides (less than 30 residues) capable of actively or passively crossing cell membranes. They can transport chemical compounds, large proteins and even nucleic acids.…”