Structural Basis of DEPTOR to Recognize Phosphatidic Acid Using its Tandem DEP Domains

Click chemistry and optogenetic approaches to visualize and manipulate phosphatidic acid signaling

“…In parallel to studies that harness the ability of IMPACT to precisely track PLD signaling dynamics, the application of optoPLD to other PA-dependent signaling pathways beyond Hippo signaling could be a powerful approach to determine the localization of functionally relevant pools of PA. For instance, the involvement of PA in mTOR signaling has long been debated (59,60). PA binds to and allosterically regulates mTOR and its components for their activation (61)(62)(63), but the precise location(s) where this activation takes place and which of the biological functions of mTOR are controlled by PA (i.e., using stringent tests for necessity and sufficiency) remain open questions in many contexts (63,64). We envision that the panel of optoPLDs with varying catalytic efficiencies will serve as useful tools to address these questions, as well as questions regarding the role of PA at other biological sites, including mitochondrial dynamics (65,66) and nuclear envelope maintenance (67).…”

“…PA binds to and allosterically regulates mTOR and its components for their activation ( 61 , 62 , 63 ), but the precise location(s) where this activation takes place and which of the biological functions of mTOR are controlled by PA ( i.e. , using stringent tests for necessity and sufficiency) remain open questions in many contexts ( 63 , 64 ). We envision that the panel of optoPLDs with varying catalytic efficiencies will serve as useful tools to address these questions, as well as questions regarding the role of PA at other biological sites, including mitochondrial dynamics ( 65 , 66 ) and nuclear envelope maintenance ( 67 ).…”

Loop modelling provides insights into regulation of mTORC1 activity via DEPTOR dimerisation

“…Further adding to the puzzle is the state of DEPTOR’s oligomerisation — DEP1–DEP2 has been characterised and crystallised as both a monomer 8,9 and a dimer 8,10 . The crystal structure of monomeric DEP1– DEP2 shows a dumbbell conformation 9 , while that of dimeric DEP1–DEP2, notably, harbours a domain-swapped DEP1 which is stabilised by an intermolecular disulphide bond 8 .…”

“…Further adding to the puzzle is the state of DEPTOR’s oligomerisation — DEP1–DEP2 has been characterised and crystallised as both a monomer 8,9 and a dimer 8,10 . The crystal structure of monomeric DEP1– DEP2 shows a dumbbell conformation 9 , while that of dimeric DEP1–DEP2, notably, harbours a domain-swapped DEP1 which is stabilised by an intermolecular disulphide bond 8 . DEP domains are known to dimerise by domain swapping 11,12 — the DEP1 domain of the chemotaxis-mediating P-Rex1, which likewise contains two DEP domains and two PDZ domains, forms a highly similar domain-swapped dimer 12 .…”

“…DEP domains are known to dimerise by domain swapping 11,12 — the DEP1 domain of the chemotaxis-mediating P-Rex1, which likewise contains two DEP domains and two PDZ domains, forms a highly similar domain-swapped dimer 12 . DEPTOR dimerisation, however, is not taken into account in many studies, which have purified the protein and its variants in the presence of reducing agents such as TCEP 3,8 or DTT 9 .…”