Biology subjects

Yeshaw, W. M.

Publications and source records attributed to Yeshaw, W. M..

3 recordsLinked to original sources

bioRxiv2025

PPM1M, a LRRK2-counteracting, phosphoRab12-preferring phosphatase with potential link to Parkinsons disease

Leucine-rich repeat kinase 2 (LRRK2) phosphorylates a subset of Rab GTPases that regulate receptor trafficking; activating mutations in LRRK2 are linked to Parkinsons disease. Rab phosphorylation is a transient event that can be reversed by phosphatases, including PPM1H, that acts on phosphoRab8A and phosphoRab10. Here we report a phosphatome-wide siRNA screen that identified PPM1M as a phosphoRab12-preferring phosphatase that also acts on phosphoRab8A and phosphoRab10. Upon knockout from cells or mice, PPM1M displays selectivity for phosphoRab12. As shown previously for mice harboring LRRK2 pathway mutations, knockout of Ppm1m leads to primary cilia loss in striatal cholinergic interneurons. We have also identified a rare PPM1M mutation in patients with Parkinsons disease that is catalytically inactive when tested in vitro and in cells. These findings identify PPM1M as a key player in the LRRK2 signaling pathway and provide a new therapeutic target for the possible benefit of patients with Parkinsons disease. TeaserParkinsons linked Rab phosphorylation is reversed by PPM1M; the inactive D440N variant is implicated in rare patient cases.

cell biology↗

bioRxiv2023

Localization of PPM1H phosphatase tunes Parkinsons disease-linked LRRK2 kinase-mediated Rab GTPase phosphorylation and ciliogenesis

PPM1H phosphatase reverses Parkinsons disease-associated, LRRK2-mediated Rab GTPase phosphorylation. We show here that PPM1H relies on an N-terminal amphipathic helix for Golgi localization. The amphipathic helix enables PPM1H to bind to liposomes in vitro, and small, highly curved liposomes stimulate PPM1H activity. We artificially anchored PPM1H to the Golgi, mitochondria, or mother centriole. Our data show that regulation of Rab10 GTPase phosphorylation requires PPM1H access to Rab10 at or near the mother centriole. Moreover, poor co-localization of Rab12 explains in part why it is a poor substrate for PPM1H in cells but not in vitro. These data support a model in which localization drives PPM1H substrate selection and centriolar PPM1H is critical for regulation of Rab GTPase-regulated ciliogenesis. Moreover, Golgi localized PPM1H maintains active Rab GTPases on the Golgi to carry out their non-ciliogenesis-related functions in membrane trafficking. Significance StatementPathogenic, hyperactive LRRK2 kinase is strongly linked to Parkinsons disease and LRRK2 phosphorylates a subset of Rab GTPases that are master regulators of membrane trafficking. PPM1H phosphatase specifically dephosphorylates Rab8A and Rab10, the major LRRK2 substrates. Here we provide novel cell biological and biochemical insight related to the localization and activation of PPM1H phosphatase. Understanding how PPM1H modulates LRRK2 activity is of fundamental interest and also important, as activators of PPM1H may eventually benefit Parkinsons disease patients.

cell biology↗

bioRxiv2023

Genome-wide screen reveals Rab12 GTPase as a critical activator of pathogenic LRRK2 kinase

Activating mutations in the Leucine Rich Repeat Kinase 2 (LRRK2) cause Parkinsons disease. LRRK2 phosphorylates a subset of Rab GTPases, particularly Rab10 and Rab8A, and we showed previously that phosphoRabs play an important role in LRRK2 membrane recruitment and activation (Vides et al., 2022). To learn more about LRRK2 pathway regulation, we carried out an unbiased, CRISPR-based genome-wide screen to identify modifiers of cellular phosphoRab10 levels. A flow cytometry assay was developed to detect changes in phosphoRab10 levels in pools of mouse NIH-3T3 cells harboring unique CRISPR guide sequences. Multiple negative and positive regulators were identified; surprisingly, knockout of the Rab12 gene was especially effective in decreasing phosphoRab10 levels in multiple cell types and knockout mouse tissues. Rab-driven increases in phosphoRab10 were specific for Rab12, LRRK2 dependent and PPM1H phosphatase reversible; they were seen with wild type and pathogenic G2019S and R1441C LRRK2. AlphaFold modeling revealed a novel Rab12 binding site in the LRRK2 Armadillo domain and we show that residues predicted to be essential for Rab12 interaction at this site influence overall phosphoRab levels in a manner distinct from Rab29 activation of LRRK2. Our data support a model in which Rab12 binding to a new site in the LRRK2 Armadillo domain activates LRRK2 kinase for Rab phosphorylation and could serve as a new therapeutic target for a novel class of LRRK2 inhibitors that do not target the kinase domain.

biochemistry↗