Mutant B-RAF regulates a Rac-dependent cadherin switch in melanoma

The ability of cells to invade into the dermis is a critical event in the development of cutaneous melanoma and ultimately an indicator of poor prognosis. However, the molecular events surrounding the acquisition of this invasive phenotype remain incompletely understood. Mutations in B-RAF are frequent in melanoma and are known to regulate the invasive phenotype. In this study, we sought to determine the molecular mechanisms controlling melanoma invasion. We found that mutant B-RAF signaling regulates a cadherin switch. In melanoma cells expressing mutant B-RAF we observed high levels of N-cadherin and low levels of E-cadherin. Depletion of mutant B-RAF, by siRNA, caused a decrease in the levels of N-cadherin and an increase in the levels of E-cadherin. Mechanistically, we found that this cadherin switch required the activity of Rac1 and its GEF, Tiam1, both of which show suppressed activity in the presence of mutant B-RAF. Consistent with the work of others, we found that depletion of mutant B-RAF decreased the invasive capacity of the melanoma cells. However, simultaneous depletion of B-RAF and Rac or Tiam1 resulted in invasive capacity similar to that of control cells. Taken together, our results suggest that mutant B-RAF signaling downregulates Tiam1/Rac activity resulting in an increase in N-cadherin levels and a decrease in E-cadherin levels and ultimately enhanced invasion

A Rnd3/p190RhoGAP pathway regulates RhoA activity in idiopathic pulmonary fibrosis fibroblasts

Idiopathic pulmonary fibrosis (IPF) is an incurable disease of the lung that is characterized by excessive deposition of extracellular matrix (ECM), resulting in disruption of normal lung function. The signals regulating fibrosis include both transforming growth factor beta (TGF-β) and tissue rigidity and a major signaling pathway implicated in fibrosis involves activation of the GTPase RhoA. During studies exploring how elevated RhoA activity is sustained in IPF, we discovered that not only is RhoA activated by profibrotic stimuli but also that the expression of Rnd3, a major antagonist of RhoA activity, and the activity of p190RhoGAP (p190), a Rnd3 effector, are both suppressed in IPF fibroblasts. Restoration of Rnd3 levels in IPF fibroblasts results in an increase in p190 activity, a decrease in RhoA activity and a decrease in the overall fibrotic phenotype. We also find that treatment with IPF drugs nintedanib and pirfenidone decreases the fibrotic phenotype and RhoA activity through up-regulation of Rnd3 expression and p190 activity. These data provide evidence for a pathway in IPF where fibroblasts down-regulate Rnd3 levels and p190 activity to enhance RhoA activity and drive the fibrotic phenotype

The role of endothelial MERTK during the inflammatory response in lungs

As a key homeostasis regulator in mammals, the MERTK receptor tyrosine kinase is crucial for efferocytosis, a process that requires remodeling of the cell membrane and adjacent actin cytoskeleton. Membrane and cytoskeletal reorganization also occur in endothelial cells during inflammation, particularly during neutrophil transendothelial migration (TEM) and during changes in permeability. However, MERTK's function in endothelial cells remains unclear. This study evaluated the contribution of endothelial MERTK to neutrophil TEM and endothelial barrier function. In vitro experiments using primary human pulmonary microvascular endothelial cells found that neutrophil TEM across the endothelial monolayers was enhanced when MERTK expression in endothelial cells was reduced by siRNA knockdown. Examination of endothelial barrier function revealed increased passage of dextran across the MERTK-depleted monolayers, suggesting that MERTK helps maintain endothelial barrier function. MERTK knockdown also altered adherens junction structure, decreased junction protein levels, and reduced basal Rac1 activity in endothelial cells, providing potential mechanisms of how MERTK regulates endothelial barrier function. To study MERTK's function in vivo, inflammation in the lungs of global Mertk-/- mice was examined during acute pneumonia. In response to P. aeruginosa, more neutrophils were recruited to the lungs of Mertk-/- than wildtype mice. Vascular leakage of Evans blue dye into the lung tissue was also greater in Mertk-/- mice. To analyze endothelial MERTK's involvement in these processes, we generated inducible endothelial cell-specific (iEC) Mertk-/- mice. When similarly challenged with P. aeruginosa, iEC Mertk-/- mice demonstrated no difference in neutrophil TEM into the inflamed lungs or in vascular permeability compared to control mice. These results suggest that deletion of MERTK in human pulmonary microvascular endothelial cells in vitro and in all cells in vivo aggravates the inflammatory response. However, selective MERTK deletion in endothelial cells in vivo failed to replicate this response

The RhoA Guanine Nucleotide Exchange Factor, LARG, Mediates ICAM-1-Dependent Mechanotransduction in Endothelial Cells To Stimulate Transendothelial Migration

RhoA-mediated cytoskeletal rearrangements in endothelial cells (ECs) play an active role in leukocyte transendothelial cell migration (TEM), a normal physiological process in which leukocytes cross the endothelium to enter the underlying tissue. Although much has been learned about RhoA signaling pathways downstream from ICAM-1 in ECs, little is known about the consequences of the tractional forces that leukocytes generate on ECs as they migrate over the surface before TEM. We have found that after applying mechanical forces to ICAM-1 clusters, there is an increase in cellular stiffening and enhanced RhoA signaling compared with ICAM-1 clustering alone. We have identified that leukemia-associated Rho guanine nucleotide exchange factor (LARG), also known as Rho GEF 12 (ARHGEF12) acts downstream of clustered ICAM-1 to increase RhoA activity, and that this pathway is further enhanced by mechanical force on ICAM-1. Depletion of LARG decreases leukocyte crawling and inhibits TEM. To our knowledge, this is the first report of endothelial LARG regulating leukocyte behavior and EC stiffening in response to tractional forces generated by leukocytes

Class A Orphans in GtoPdb v.2023.1

Table 1 lists a number of putative GPCRs identified by NC-IUPHAR [161], for which preliminary evidence for an endogenous ligand has been published, or for which there exists a potential link to a disease, or disorder. These GPCRs have recently been reviewed in detail [121]. The GPCRs in Table 1 are all Class A, rhodopsin-like GPCRs. Class A orphan GPCRs not listed in Table 1 are putative GPCRs with as-yet unidentified endogenous ligands.Table 1: Class A orphan GPCRs with putative endogenous ligands GPR3GPR4GPR6GPR12GPR15GPR17GPR20 GPR22GPR26GPR31GPR34GPR35GPR37GPR39 GPR50GPR63GPR65GPR68GPR75GPR84GPR87 GPR88GPR132GPR149GPR161GPR183LGR4LGR5 LGR6MAS1MRGPRDMRGPRX1MRGPRX2P2RY10TAAR2 In addition the orphan receptors GPR18, GPR55 and GPR119 which are reported to respond to endogenous agents analogous to the endogenous cannabinoid ligands have been grouped together (GPR18, GPR55 and GPR119)

Class A Orphans (version 2020.5) in the IUPHAR/BPS Guide to Pharmacology Database

Table 1 lists a number of putative GPCRs identified by NC-IUPHAR [194], for which preliminary evidence for an endogenous ligand has been published, or for which there exists a potential link to a disease, or disorder. These GPCRs have recently been reviewed in detail [150]. The GPCRs in Table 1 are all Class A, rhodopsin-like GPCRs. Class A orphan GPCRs not listed in Table 1 are putative GPCRs with as-yet unidentified endogenous ligands.Table 1: Class A orphan GPCRs with putative endogenous ligands GPR3 GPR4 GPR6 GPR12 GPR15 GPR17 GPR20 GPR22 GPR26 GPR31 GPR34 GPR35 GPR37 GPR39 GPR50 GPR63 GRP65 GPR68 GPR75 GPR84 GPR87 GPR88 GPR132 GPR149 GPR161 GPR183 LGR4 LGR5 LGR6 MAS1 MRGPRD MRGPRX1 MRGPRX2 P2RY10 TAAR2 In addition the orphan receptors GPR18, GPR55 and GPR119 which are reported to respond to endogenous agents analogous to the endogenous cannabinoid ligands have been grouped together (GPR18, GPR55 and GPR119)

Class A Orphans in GtoPdb v.2022.3

Table 1 lists a number of putative GPCRs identified by NC-IUPHAR [161], for which preliminary evidence for an endogenous ligand has been published, or for which there exists a potential link to a disease, or disorder. These GPCRs have recently been reviewed in detail [121]. The GPCRs in Table 1 are all Class A, rhodopsin-like GPCRs. Class A orphan GPCRs not listed in Table 1 are putative GPCRs with as-yet unidentified endogenous ligands.Table 1: Class A orphan GPCRs with putative endogenous ligands GPR3GPR4GPR6GPR12GPR15GPR17GPR20 GPR22GPR26GPR31GPR34GPR35GPR37GPR39 GPR50GPR63GPR65GPR68GPR75GPR84GPR87 GPR88GPR132GPR149GPR161GPR183LGR4LGR5 LGR6MAS1MRGPRDMRGPRX1MRGPRX2P2RY10TAAR2 In addition the orphan receptors GPR18, GPR55 and GPR119 which are reported to respond to endogenous agents analogous to the endogenous cannabinoid ligands have been grouped together (GPR18, GPR55 and GPR119)

Class A Orphans in GtoPdb v.2021.3

Table 1 lists a number of putative GPCRs identified by NC-IUPHAR [161], for which preliminary evidence for an endogenous ligand has been published, or for which there exists a potential link to a disease, or disorder. These GPCRs have recently been reviewed in detail [121]. The GPCRs in Table 1 are all Class A, rhodopsin-like GPCRs. Class A orphan GPCRs not listed in Table 1 are putative GPCRs with as-yet unidentified endogenous ligands.Table 1: Class A orphan GPCRs with putative endogenous ligands GPR3GPR4GPR6GPR12GPR15GPR17GPR20 GPR22GPR26GPR31GPR34GPR35GPR37GPR39 GPR50GPR63GRP65GPR68GPR75GPR84GPR87 GPR88GPR132GPR149GPR161GPR183LGR4LGR5 LGR6MAS1MRGPRDMRGPRX1MRGPRX2P2RY10TAAR2 In addition the orphan receptors GPR18, GPR55 and GPR119 which are reported to respond to endogenous agents analogous to the endogenous cannabinoid ligands have been grouped together (GPR18, GPR55 and GPR119)

Class A Orphans (version 2019.4) in the IUPHAR/BPS Guide to Pharmacology Database

Table 1 lists a number of putative GPCRs identified by NC-IUPHAR [191], for which preliminary evidence for an endogenous ligand has been published, or for which there exists a potential link to a disease, or disorder. These GPCRs have recently been reviewed in detail [148]. The GPCRs in Table 1 are all Class A, rhodopsin-like GPCRs. Class A orphan GPCRs not listed in Table 1 are putative GPCRs with as-yet unidentified endogenous ligands.Table 1: Class A orphan GPCRs with putative endogenous ligands GPR3GPR4GPR6GPR12GPR15GPR17GPR20 GPR22GPR26GPR31GPR34GPR35GPR37GPR39 GPR50GPR63GRP65GPR68GPR75GPR84GPR87 GPR88GPR132GPR149GPR161GPR183LGR4LGR5 LGR6MAS1MRGPRDMRGPRX1MRGPRX2P2RY10TAAR2 In addition the orphan receptors GPR18, GPR55 and GPR119 which are reported to respond to endogenous agents analogous to the endogenous cannabinoid ligands have been grouped together (GPR18, GPR55 and GPR119)

Nuclear SUN1 stabilizes endothelial cell junctions via microtubules to regulate blood vessel formation

Endothelial cells line all blood vessels, where they coordinate blood vessel formation and the blood-tissue barrier via regulation of cell-cell junctions. The nucleus also regulates endothelial cell behaviors, but it is unclear how the nucleus contributes to endothelial cell activities at the cell periphery. Here, we show that the nuclear-localized linker of the nucleoskeleton and cyto-skeleton (LINC) complex protein SUN1 regulates vascular sprouting and endothelial cell-cell junction morphology and function. Loss of murine endothelial Sun1 impaired blood vessel formation and destabilized junctions, angiogenic sprouts formed but retracted in SUN1-depleted sprouts, and zebrafish vessels lacking Sun1b had aberrant junctions and defective cell-cell connections. At the cellular level, SUN1 stabilized endothelial cell-cell junctions, promoted junction function, and regulated contractility. Mechanistically, SUN1 depletion altered cell behaviors via the cytoskeleton without changing transcriptional profiles. Reduced peripheral microtubule density, fewer junction contacts, and increased catastrophes accompanied SUN1 loss, and microtubule depolymerization phenocopied effects on junctions. Depletion of GEF-H1, a microtubule-regulated Rho activator, or the LINC complex protein nesprin-1 rescued defective junctions of SUN1-depleted endothelial cells. Thus, endothelial SUN1 regulates peripheral cell-cell junctions from the nucleus via LINC complex-based microtubule interactions that affect peripheral microtubule dynamics and Rho-regulated contractility, and this long-range regulation is important for proper blood vessel sprouting and junction integrity