Discovery of new Longin and Roadblock domains that form platforms for small GTPases in Ragulator and TRAPP-II

Levine, Tim P.; Daniels, Rachel D.; Wong, Louise H.; Gatta, Alberto T.; Gerondopoulos, Andreas; Barr, Francis

doi:10.4161/sgtp.24262

Cited by 86 publications

(110 citation statements)

References 53 publications

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“…Profile‐profile tools such as HHsearch (Box 3) have previously detected many new domains,49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59 so we asked if HHsearch can improve on PSI‐BLAST for PH‐like domains. We first studied how HHsearch detects PH‐like domains in PDB, where we are certain if hits are true or false.…”

Section: Figurementioning

confidence: 99%

Using HHsearch to tackle proteins of unknown function: A pilot study with PH domains

Fidler¹,

Murphy²,

Courtis³

et al. 2016

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Advances in membrane cell biology are hampered by the relatively high proportion of proteins with no known function. Such proteins are largely or entirely devoid of structurally significant domain annotations. Structural bioinformaticians have developed profile‐profile tools such as HHsearch (online version called HHpred), which can detect remote homologies that are missed by tools used to annotate databases. Here we have applied HHsearch to study a single structural fold in a single model organism as proof of principle. In the entire clan of protein domains sharing the pleckstrin homology domain fold in yeast, systematic application of HHsearch accurately identified known PH‐like domains. It also predicted 16 new domains in 13 yeast proteins many of which are implicated in intracellular traffic. One of these was Vps13p, where we confirmed the functional importance of the predicted PH‐like domain. Even though such predictions require considerable work to be corroborated, they are useful first steps. HHsearch should be applied more widely, particularly across entire proteomes of model organisms, to significantly improve database annotations.

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Section: Figurementioning

confidence: 99%

Using HHsearch to tackle proteins of unknown function: A pilot study with PH domains

Fidler¹,

Murphy²,

Courtis³

et al. 2016

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“…The observations made by Levine et al [23] indicated that the uncharacterized yeast proteins Ycr075w-a/Ego2 and/or Ynr034w-a/Ego4 might, like LAMTOR4-LAM-TOR5, be positive regulators of the Gtr GTPases and TORC1. To investigate the potential roles of these two proteins in TORC1 signaling, we assessed the ability of cells lacking either Ego2 or Ego4 to grow in the presence of increasing concentrations of the TORC1 inhibitor rapamycin.…”

Section: Ego2 Is a Positive Regulator Of Torc1 Signalingmentioning

confidence: 99%

“…Moreover, no obvious counterparts of mammalian LAMTOR4 and LAMTOR5, shown to be essential for the GEF activity of the Ragulator, appeared to exist in yeast. However, a recent study that employed HHpred, a highly sensitive method for structural homology detection, identified two uncharacterized yeast proteins Ycr075w-a and Ynr034w-a as hypothetical LAMTOR4 and LAMTOR5 homologs [23]. While very little is known about either protein, the NMR structure of Ynr034w-a reveals the presence of a Roadblock domain-like fold, a common feature of many GTPase-interacting proteins.…”

Section: Introductionmentioning

confidence: 99%

“…In this system, the bait protein is artificially localized to membranes. Since Ego2 and Ego4 are predicted to have highly similar structures [23], it is possible that Ego4 interacts with Gtr1 and Gtr2 only when it is forced to localize to membranes, but not necessarily under physiological conditions. Consistent with such an explanation, Ego2-HA 3 was found to co-precipitate with both Ego1-GFP and Ego3-GFP, whereas Ego4-HA 3 showed no interaction with either protein ( Figure 3B).…”

Section: Ego2 Is a Subunit Of The Egocmentioning

confidence: 99%

“…While very little is known about either protein, the NMR structure of Ynr034w-a reveals the presence of a Roadblock domain-like fold, a common feature of many GTPase-interacting proteins. Curiously, Roadblock domains are predicted to be present not only in Ycr075w-a and the LAMTOR4-LAMTOR5 heterodimer, but also to exist in the LAMTOR2-LAMTOR3 heterodimer, the Ego3 homodimer, as well as within the C-terminal dimerization domains of the Rag/Gtr GTPases [23].…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Crystal structure of the Ego1-Ego2-Ego3 complex and its role in promoting Rag GTPase-dependent TORC1 signaling

et al. 2015

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The target of rapamycin complex 1 (TORC1) integrates various hormonal and nutrient signals to regulate cell growth, proliferation, and differentiation. Amino acid-dependent activation of TORC1 is mediated via the yeast EGO complex (EGOC) consisting of Gtr1, Gtr2, Ego1, and Ego3. Here, we identify the previously uncharacterized Ycr075w-a/Ego2 protein as an additional EGOC component that is required for the integrity and localization of the heterodimeric Gtr1-Gtr2 GTPases, equivalent to mammalian Rag GTPases. We also report the crystal structure of the Ego1-Ego2-Ego3 ternary complex (EGO-TC) at 2.4 Å resolution, in which Ego2 and Ego3 form a heterodimer flanked along one side by Ego1. Structural data also reveal the structural conservation of protein components between the yeast EGO-TC and the human Ragulator, which acts as a GEF for Rag GTPases. Interestingly, however, artificial tethering of Gtr1-Gtr2 to the vacuolar membrane is sufficient to activate TORC1 in response to amino acids even in the absence of the EGO-TC. Our structural and functional data therefore support a model in which the EGO-TC acts as a scaffold for Rag GTPases in TORC1 signaling.

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The TRAPP complexes: oligomeric exchange factors that activate the small GTPases Rab1 and Rab11

Galindo

Munro

2022

FEBS Letters

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The Transport Protein Particle (TRAPP) complexes are highly conserved multisubunit complexes that act as nucleotide exchange factors (GEFs) for Rab GTPases. They act in both protein secretion and autophagy and have also been proposed to have a role in other processes such as cytokinesis and ciliogenesis. There are two TRAPP complexes in metazoans: TRAPPII, which activates Rab11; and TRAPPIII, which activates Rab1. Both complexes share a core of small subunits that form the active site for the exchange of GDP for GTP. In addition, each TRAPP complex has distinct large subunits that determine the specificity of each complex towards its substrate Rab and are essential for activity in vivo. Crystal structures have revealed the organisation of the TRAPP core and the mechanism of Rab1 activation, whilst recent cryo‐EM structures have unveiled the arrangement of the specific subunits around the core to form each complex. Combining these findings with functional experiments has allowed the proposal of mechanisms for how the specificity of each complex towards their cognate Rab is determined and for the arrangement of these large complexes on the membrane.

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Discovery of new Longin and Roadblock domains that form platforms for small GTPases in Ragulator and TRAPP-II

Cited by 86 publications

References 53 publications

Using HHsearch to tackle proteins of unknown function: A pilot study with PH domains

Using HHsearch to tackle proteins of unknown function: A pilot study with PH domains

Crystal structure of the Ego1-Ego2-Ego3 complex and its role in promoting Rag GTPase-dependent TORC1 signaling

The TRAPP complexes: oligomeric exchange factors that activate the small GTPases Rab1 and Rab11

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