Intramolecular Feedback Regulation of the LRRK2 Roc G Domain by LRRK2 Kinase-Dependent Autophosphorylation

แนวคิดหลัก

LRRK2 kinase activity negatively regulates the GTPase activity of the Roc domain through an intramolecular feedback mechanism involving autophosphorylation of the Roc P-loop residue T1343.

บทคัดย่อ

This study investigates the interplay between the GTPase and kinase domains of the Parkinson's disease-associated protein LRRK2. The key findings are:

1. LRRK2 has a KM value for GTP hydrolysis that lies within the physiological range of cellular GTP concentrations, suggesting its GTPase activity is sensitive to changes in GTP levels.

2. The Parkinson's disease variants R1441G and G2019S differentially impact the kinetics of LRRK2's GTPase activity, with R1441G showing increased catalytic efficiency and G2019S exhibiting reduced efficiency.

3. LRRK2 kinase activity negatively regulates the GTPase activity of the Roc domain through an intramolecular feedback mechanism. Autophosphorylation of the Roc P-loop residue T1343 is critical for this regulation.

4. The T1343A phospho-null mutant disrupts this negative feedback loop, leading to LRRK2 activity comparable to the hyperactive G2019S variant.

5. This interplay between the GTPase and kinase domains, mediated by autophosphorylation, likely plays an important role in the regulation of LRRK2 signaling and the differential impact of Parkinson's disease mutations.

สถิติ

LRRK2 wild-type has a KM of 554 ± 62 μM and a kcat of 0.36 ± 0.02 min-1 for GTP hydrolysis.
The R1441G variant has a KM of 271 ± 27 μM and an increased kcat compared to wild-type.
The G2019S variant has a significantly increased KM of 867 ± 110 μM compared to wild-type.
The kinase-dead K1906M variant has a lower KM of 181 ± 58 μM.
Incubation of wild-type LRRK2 with ATP increases the KM to 1036 ± 168 μM and decreases the kcat to 0.13 ± 0.01 min-1.

คำพูด

"LRRK2 kinase activity might counteract this mechanism by auto-phosphorylation of the P-loop residue T1343, shifting the equilibrium back to the monomeric state."
"Removing of this regulatory phosphosite leads to an LRRK2 activity comparable to G2019S, and cannot be further increased by introducing the double mutant T1343A/G2019S."

ข้อมูลเชิงลึกที่สำคัญจาก

Intramolecular feedback regulation of the LRRK2 Roc G domain by a LRRK2 kinase dependent mechanism

by Gilsbach,B. ... ที่ www.biorxiv.org 07-31-2023

https://www.biorxiv.org/content/10.1101/2023.07.31.549909v2

สอบถามเพิ่มเติม

How might the differential impact of Parkinson's disease mutations on LRRK2 GTPase kinetics contribute to the variable penetrance and phenotypes observed in patients

The differential impact of Parkinson's disease mutations on LRRK2 GTPase kinetics can contribute to the variable penetrance and phenotypes observed in patients in several ways. Firstly, mutations that increase the catalytic efficiency of the GTPase activity, such as the R1441G variant, may lead to hyperactivation of LRRK2 signaling pathways, resulting in more severe phenotypes. On the other hand, mutations like G2019S, which increase the KM value and reduce catalytic efficiency, may result in a dampened GTPase activity, leading to milder phenotypes. These variations in GTPase kinetics can affect the overall signaling output of LRRK2, influencing downstream pathways involved in Parkinson's disease pathogenesis. The interplay between the kinase domain and the G domain, as well as the feedback regulation mechanism described, can further modulate the impact of these mutations on LRRK2 activity, contributing to the variable penetrance observed in patients.

What other cellular factors or signaling pathways might interact with or modulate this intramolecular feedback regulation of LRRK2 by its kinase domain

Several cellular factors and signaling pathways may interact with or modulate the intramolecular feedback regulation of LRRK2 by its kinase domain. One key factor is the cellular GTP concentration, as changes in global or local GTP levels can directly impact the GTPase activity of LRRK2. Additionally, other kinases or phosphatases within the cell may regulate the phosphorylation status of LRRK2, influencing its feedback mechanism. Signaling pathways involved in protein degradation or turnover, such as the ubiquitin-proteasome system, may also play a role in regulating LRRK2 activity by controlling its stability and degradation. Furthermore, pathways related to cellular stress responses, inflammation, or mitochondrial function, which are known to be dysregulated in Parkinson's disease, could potentially intersect with LRRK2 signaling and modulate its feedback regulation.

Given the conserved phosphorylation of P-loop residues in Rab proteins, could similar regulatory mechanisms involving GTPase activity and monomer-dimer equilibrium be a common theme across the Rab protein family

The conserved phosphorylation of P-loop residues in Rab proteins suggests that similar regulatory mechanisms involving GTPase activity and monomer-dimer equilibrium could be a common theme across the Rab protein family. Phosphorylation of P-loop residues in Rab proteins may serve as a regulatory switch to modulate their GTPase activity, affecting their interactions with effector proteins and downstream signaling pathways. This regulatory mechanism could provide a means for fine-tuning the activity of Rab proteins in response to cellular signals or stress conditions. Additionally, the modulation of the monomer-dimer equilibrium by phosphorylation of P-loop residues could impact the subcellular localization and function of Rab proteins, further diversifying their roles in cellular processes. Overall, the regulatory mechanisms involving P-loop phosphorylation in Rab proteins may contribute to the dynamic control of intracellular trafficking, vesicle transport, and signaling pathways in the cell.