Biology subjects

Clough, R.

Publications and source records attributed to Clough, R..

3 recordsLinked to original sources

bioRxiv2023

A Ciliary SMOOTHENED-GRK2-PKA Signaling Pathway Initiates Hedgehog Signal Transduction

During Hedgehog (Hh) signal transduction in development and disease, the atypical G protein-coupled receptor (GPCR) SMOOTHENED (SMO) communicates with GLI transcription factors by binding the protein kinase A catalytic subunit (PKA-C) and physically blocking its enzymatic activity. Here we show that GPCR kinase 2 (GRK2) orchestrates this process during endogenous Hh pathway activation in the vertebrate primary cilium. Upon SMO activation, GRK2 rapidly relocalizes from the ciliary base to the shaft, triggering SMO phosphorylation and PKA-C interaction. Reconstitution studies reveal that GRK2 phosphorylation enables active SMO to bind PKA-C directly. Lastly, the SMO-GRK2-PKA pathway underlies Hh signal transduction in a range of cellular and in vivo models. Thus, GRK2 phosphorylation of ciliary SMO, and the ensuing PKA-C binding and inactivation, are critical initiating events for the intracellular steps in Hh signaling. More broadly, our study suggests an expanded role for GRKs in enabling direct GPCR interactions with diverse intracellular effectors.

cell biology↗

bioRxiv2019

The Role of the Ovarian Cancer G -Coupled Receptor (OGR1) in Idiopathic Pulmonary Fibrosis

Idiopathic pulmonary fibrosis (IPF) is a disease characterized by irreversible scarring of the lung that is associated with significant mortality and morbidity. The pathophysiology is incompletely understood but it is well-established that fibroblast to myofibroblast differentiation is a key feature of pulmonary fibrosis. Our lab has established that a reduction in extracellular pH is one of several important pathways responsible for the activation of latent TGF-{beta} in the extracellular space. TGF-{beta} activation further decreases extracellular pH and creates a feed-forward mechanism that stimulates myofibroblast differentiation and activation of additional TGF-{beta}. Given the importance of TGF-{beta} and extracellular acidification to the progression of pulmonary fibrosis, we sought to identify novel mechanisms that are involved in pH-dependent fibrotic signaling. The proton sensing G-Protein Coupled family of receptors are activated in acidic environments, but their role in fibrotic signaling has not been studied. Here we report that the Ovarian Cancer G-Protein Coupled Receptor1 (OGR1 or GPR68), a member of the family of proton sensing G-Protein Coupled Receptors, negatively regulates pro-fibrotic signaling. We demonstrate that OGR1 expression is significantly reduced in lung tissue from patients with IPF and TGF-{beta} decreases OGR1 expression. In fibroblasts, a reduction in expression of OGR1 (OGR knockout lung fibroblasts) and knockdown (OGR siRNA), promotes in vitro myofibroblast differentiation. In contrast, OGR1 overexpression inhibits myofibroblast differentiation. Finally, we demonstrate that OGR1 negatively regulates TGF-{beta} stimulation through inhibition of focal adhesion kinase (FAK) phosphorylation. Our results suggest that preserving OGR1 expression may represent a novel therapeutic strategy in pulmonary fibrosis.

cell biology↗

bioRxiv2019

RUNX1/RUNX1T1 controls alternative splicing in the t(8;21)-positive acute myeloid leukemia cells

The fusion oncogene RUNX1/RUNX1T1 encodes an aberrant transcription factor, which plays a key role in the initiation and maintenance of the t(8;21)-positive acute myeloid leukemia. Here we show that this oncogene is a regulator of the alternative RNA splicing for a sub-set of genes in the leukemia cells. We found two primary mechanisms underlying changes in the production of RNA isoforms: (i) RUNX1/RUNX1T1-mediated regulation of alternative transcription start sites selection in target genes, and (ii) direct or indirect control of the expression of the genes encoding splicing factors. The first mechanism leads to the expression of RNA isoforms with alternative structure of the 5-UTR regions. The second mechanism generates alternative transcripts with new junctions between internal cassettes and constitutive exons. We also show that RUNX1/RUNX1T1-mediated differential splicing affects several functional groups of genes and produces proteins with unique conserved domain structures. In summary, this study reveals a novel layer of RUNX1/RUNX1T1-dependent transcriptome organization in t(8;21)-positive acute myeloid leukemia.

cancer biology↗