Reaction-based fluorescent sensor for investigating mobile Zn ²⁺ in mitochondria of healthy versus cancerous prostate cells

Abstract

Chelatable, mobile forms of divalent zinc, Zn(II), play essential signaling roles in mammalian biology. A complex network of zinc import and transport proteins has evolved to control zinc concentration and distribution on a subcellular level. Understanding the action of mobile zinc requires tools that can detect changes in Zn(II) concentrations at discrete cellular locales. We present here a zinc-responsive, reaction-based, targetable probe based on the diacetyled form of Zinpyr-1. The compound, (6-amidoethyl)triphenylphosphonium Zinpyr-1 diacetate (DA-ZP1-TPP), is essentially nonfluorescent in the metal-free state; however, exposure to Zn (II) triggers metal-mediated hydrolysis of the acetyl groups to afford a large, rapid, and zinc-induced fluorescence response. DA-ZP1-TPP is insensitive to intracellular esterases over a 2-h period and is impervious to proton-induced turn-on. A TPP unit is appended for targeting mitochondria, as demonstrated by live cell fluorescence imaging studies. The practical utility of DA-ZP1-TPP is demonstrated by experiments revealing that, in contrast to healthy epithelial prostate cells, tumorigenic cells are unable to accumulate mobile zinc within their mitochondria.reaction-based probe | prostate cancer | zinc biology | fluorescence microscopy

Peptide targeting of fluorescein-based sensors to discrete intracellular locales

“…These results mirror recent work with ZP1-TPP, which revealed the propensity for ZP1 conjugates to accumulate within acidic vesicles and lose their ability to respond to zinc. 12 …”

“…12 Our principal findings were (i) that installation of zinc-sensitive acetyl groups onto the phenolic oxygen atoms of a ZP1 derivative locked the fluorescein scaffold into its non-fluorescent lactone isomer; (ii) that the acetyl groups are stable to endogenous esterases but rapidly hydrolyze upon exposure to zinc, resulting in a significantly enhanced dynamic range; 12 and (iii) that acetylation also increased the hydrophobicity of the probe, thus enabling it to avoid endosomal entrapment and, instead, traffic to mitochondria via a triphenyl-phosphonium moiety.…”

Zn(ii)-coordination modulated ligand photophysical processes – the development of fluorescent indicators for imaging biological Zn(ii) ions

“…316 This compound carries the spirolactone form of ZP1 , 317,318 the first member of the ZP series developed in the Lippard laboratory. The two hydroxyl groups on the xanthene ring are acetylated (hence “DA” in the name) to prevent spirolactone ring from opening.…”

“…Zn(II) indicators SZnMito , 313 Mito-ST , 314 DQZn2 , 315 and DA-ZP1-TPP , 316 that contain mitochondrial-targeting alkylTPP group.…”

A conveniently prepared and hypersensitized small molecular fluorescent probe: Rapidly detecting free zinc ion in HepG2 cells and Arabidopsis

“…Meanwhile, zinc ions act a significant role in many biochemical reactions in plants such as maize, sorghum and sugarcane, which show reduced photosynthetic carbon metabolism due to zinc deficiency (Dittmer, etal., 2009;Sinha, etal., 2013;Chyan, etal., 2014) [7][8][9]. Therefore, it is of great significance to develop analytical methods for detecting and monitoring Zn 2+ at the level of cells and tissues in the body including plants and animals.…”