Hydrophobic resorufamine derivatives: Potent and selective red fluorescent probes of the endoplasmic reticulum of mammalian cells

Abstract

The endoplasmic reticulum (ER) of eukaryotic cells plays critical roles in the processing of secreted and transmembrane proteins. Defects in these functions are associated with a wide range of pathologies. To image this organelle, cells are often treated with fluorescent ER-Tracker dyes. Although these compounds are selective, existing red fluorescent probes of the ER are costly glibenclamide derivatives that inhibit ER-associated sulphonylurea receptors. To provide simpler and more cost-effective red fluorescent probes of the ER, we synthesized amino analogues of the fluorophore resorufin. By varying the polarity of linked substituents, we identified hexyl resorufamine (HRA) as a novel hydrophobic (cLogD (pH 7.4) = 3.8) red fluorescent (Ex. 565 nm; Em. 614 nm in ethanol) molecular probe. HRA is exceptionally bright in organic solvents (quantum yield = 0.70), it exclusively localizes to the ER of living HeLa cells as imaged by confocal microscopy, it is effective at concentrations as low as 100 nM, and it is non-toxic under these conditions. To examine its utility, we used HRA to facilitate visualization of small molecule-mediated release of a GFP-GPI fusion protein from the ER into the secretory pathway. HRA represents a potent, selective, and cost-effective probe for imaging and labeling the ER.

Fluorescent probes for targeting endoplasmic reticulum: design strategies and their applications

“…Sulfonylurea is known for its inhibitory action towards the ATP-dependent K + channels, which are abundant on the cytosolic face of the ER membrane and the lumen of ER has sulphonamide receptors that aid in binding. 27,28 Therefore, glyburide or glibenclamide analogues of sulfonylurea are utilized as targeting groups for ER. Apart from the sulfonylurea, probes containing halogens, amphiphilic cationic groups, or substituents with optimum hydrophobicity are also used to target the ER.…”

An aggregation-induced emission platform for efficient Golgi apparatus and endoplasmic reticulum specific imaging

“…Phenylsulfonamide unit is utilized as the Golgi targeting moiety due to its strong binding ability with cyclooxygenase-2 (COX-2), which is an inducing enzyme that expresses and accumulates in the Golgi apparatus. 33,34 Additionally, glibenclamide moiety that can bind to ATP-sensitive potassium channel (K ATP ) on the ER membrane, 35,36 is employed to achieve ER-targeting. TTBS, TTVBS, TTANBS and TANBS are designed as GA probes (Fig.…”

Endoplasmic reticulum targeting fluorescent probes to image mobile Zn2+

“…Glibenclamide, an antidiabetic drug, is employed in the commercial dyes ER tracker red and ER tracker green because it binds and inhibits ATP-sensitive K + channels that are abundant in the ER membrane. 50,51 We were further encouraged by a report of the use of glibenclamide as a targeting group in a small molecule chemical probe which was shown to selectively accumulate in the ER and image glutathione. 52 We therefore sought to develop a route to a 'clickable' variant of glibenclamide that we could incorporate into our modular synthetic route and prepared 'click' glibenclamide 2 through six steps in moderate yield (see Scheme S1, ESI †).…”