Rab1 GTPase promotes expression of β-adrenergic receptors in rat pulmonary microvascular endothelial cells

Abstract

It is known that Rab1 regulates the expression and function of beta-adrenoceptors (β-ARs) in many cells. However, the effect of these changes in rat pulmonary microvascular endothelial cells (RPMVECs) is not known. In the present study, we investigated the role of Rab1, a Ras-like GTPase that coordinates protein transport from the endoplasmic reticulum (ER) to the Golgi body and regulates the cell-surface targeting and function of endogenous β-ARs in RPMVECs in the presence of lipopolysaccharide (LPS). We found that lentivirus-driven expression of wild-type Rab1 (Rab1WT) in RPMVECs strongly enhanced the amount of β-ARs on the cell surface, whereas the dominant-negative mutant Rab1N124I significantly attenuated β-ARs expression on the cell surface. In addition, LPS stimulation significantly reduced β-ARs expression on the cell surface in RPMVECs; however, this effect was reversed by over-expression of wild-type Rab1WT. Fluorescent microscopy analysis demonstrated that expression of Rab1N124I and Rab1 small interfering RNA (siRNA) significantly induced the accumulation of green fluorescent protein (GFP)-tagged β2-AR in the ER. Consistent with their effects on β-ARs export, Rab1WT and Rab1N124I differentially modified the β-AR-mediated activation of extracellular signal-regulated kinase1/2 (ERK1/2). Importantly, over-expression of Rab1WT markedly reduced LPS-induced hyper-permeability of RPMVECs by increasing the expression of β2-AR on the cell surface. These data reveal that β-ARs function in RPMVECs could be modulated by manipulating β-ARs traffic from the ER to the Golgi body. We propose the ER-to-Golgi transport as a regulatory site for control of permeability of RPMVECs.

Regulation on Toll-like Receptor 4 and Cell Barrier Function by Rab26 siRNA-loaded DNA Nanovector in Pulmonary Microvascular Endothelial Cells

“…Inactivation of Rab GTPases induces the mistrafficking of GPCRs, which leads to the dysfunction of the receptors that is directly linked to disorder of cell function 3, 4, 44. Consistent with other studies 3, 4, 44, our study showed that Rab26 silencing with siRab26-DYM in LPS stimulated cells increased the surface levels of TLR4 and subsequently activated TLR4 signal pathway in primary HPMVECs. Up-regulation of Rab26 expression with an adenovirus partially reversed the LPS induced activation of TLR4 signal pathway.…”

“…Previous studies have demonstrated that Rab26 plays an important role in regulating the transport of α2-ARs from the Golgi to the membrane 9, lysosome trafficking and mitochondrial redistribution 10 and induces the autophagy of neuronal somata 11. As shown in our previous study, the GTPases Rab1 and Rab5a control the permeability of HPMVECs by regulating the trafficking and localization of the β-AR 3, 4. However, whether Rab26 regulates the permeability of HPMVECs remains unclear.…”

Rab1 GTPase regulates phenotypic modulation of pulmonary artery smooth muscle cells by mediating the transport of angiotensin II type 1 receptor under hypoxia

“…Wild type Rab1-pcDNA3 (Rab1WT), pcDNA3 plasmid, pDsRed2-ER, an ER marker, AT1R-pCDM8 tagged with green fluorescent protein (GFP-AR1T) and AT1R tagged with hemagglutinin (HA-AT1R) were kind gifts of Dr. Wu (Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Sciences Center, New Orleans, Louisiana). All other materials were described elsewhere (Wang et al, 2008; Li et al, 2010). …”

Small GTPase regulation of GPCR anterograde trafficking

“…The Rab1 effects on the cell-surface export of β 2 -AR and AT1R are more complicated than the effects on other GPCRs. Overexpression of Rab1 enhanced the cell-surface transport of endogenous β 2 -AR in endothelial cells [16] but had no effect on exogenous β 2 -AR in HEK293 cells [15] or endogenous β 2 -AR in cardiomyocytes [14]. Rab1 expression increased the cell-surface expression of endogenous AT1R in cardiomyocytes [18] and endothelial cells [19] without influencing exogenously expressed AT1R in HEK293 cells [15].…”