Glutathione and the intracellular labile heme pool

O'Keeffe, Rosemary; Latunde-Dada, Gladys O.; Chen, Yulin; Kong, Xiaole; Cilibrizzi, Agostino; Hider, Robert C.

doi:10.1007/s10534-020-00274-w

Cited by 14 publications

(9 citation statements)

References 12 publications

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“…In the current report, we exploited HO-2 selectivity for ferric heme over ferrous heme (86) and perturbations to its expression by silencing or over-expression to infer that LH is largely oxidized (Figure 6). These findings suggest that the glutathione redox buffer does not equilibrate with LH to keep it reduced and supports prior studies that found glutathione binds oxidized heme too weakly (KD III = 20 M) to represent a physiologically relevant buffering factor (90). LH being oxidized is consistent with the observation that DNA and RNA guanine quadruplexes, which have nM affinities for ferric heme, also appear to bind and regulate heme availability in human cell lines (67,71).…”

Section: Discussionsupporting

confidence: 89%

Heme oxygenase-2 (HO-2) binds and buffers labile heme, which is largely oxidized, in human embryonic kidney cells

Da¹,

Moore²,

Liu

et al. 2021

Preprint

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Heme oxygenases (HO) detoxify heme by oxidatively degrading it into carbon monoxide, iron, and biliverdin, which is reduced to bilirubin and excreted. Humans express two isoforms: inducible HO-1, which is up-regulated in response to various stressors, including excess heme, and constitutive HO-2. While much is known about the regulation and physiological function of HO-1, comparatively little is known about the role of HO-2 in regulating heme homeostasis. The biochemical necessity for expressing constitutive HO-2 is largely dependent on whether heme is sufficiently abundant and accessible as a substrate under conditions in which HO-1 is not induced. By measuring labile heme, total heme, and bilirubin in human embryonic kidney HEK293 cells with silenced or over-expressed HO-2, and various HO-2 mutant alleles, we found that endogenous heme is too limiting to support HO-2 catalyzed heme degradation. Rather, we discovered that a novel role for HO-2 is to bind and buffer labile heme. Taken together, in the absence of excess heme, we propose that HO-2 regulates heme homeostasis by acting as a heme buffering factor in control of heme bioavailability. When heme is in excess, HO-1 is induced and both HO-2 and HO-1 can provide protection from heme toxicity by enzymatically degrading it. Our results explain why catalytically inactive mutants of HO-2 are cytoprotective against oxidative stress. Moreover, the change in bioavailable heme due to HO-2 overexpression, which selectively binds ferric over ferrous heme, is consistent with the labile heme pool being oxidized, thereby providing new insights into heme trafficking and signaling.

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Section: Discussionsupporting

confidence: 89%

Heme oxygenase-2 (HO-2) binds and buffers labile heme, which is largely oxidized, in human embryonic kidney cells

Da¹,

Moore²,

Liu

et al. 2021

Preprint

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“…In recent years the term “labile heme” has appeared. ,,,,− The use of this term allows a distinction to be made between the proportion of the total heme content that is available for mobilization, and the proportion that is unavailable (or, more precisely, inert for exchange) because it is bound with a high affinity, and therefore irreversibly, to proteins. Ideas on what the concepts of labile heme actually means mimic the early ideas on free heme. , Labile heme is envisaged as being continuously engaged in transient binding to intracellular proteins that exist to actively buffer heme concentrations in the cell.…”

Section: Terminologymentioning

confidence: 99%

Understanding the Logistics for the Distribution of Heme in Cells

Gallio

Fung

Cammack-Najera

et al. 2021

JACS Au

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Heme is essential for the survival of virtually all living systems—from bacteria, fungi, and yeast, through plants to animals. No eukaryote has been identified that can survive without heme. There are thousands of different proteins that require heme in order to function properly, and these are responsible for processes such as oxygen transport, electron transfer, oxidative stress response, respiration, and catalysis. Further to this, in the past few years, heme has been shown to have an important regulatory role in cells, in processes such as transcription, regulation of the circadian clock, and the gating of ion channels. To act in a regulatory capacity, heme needs to move from its place of synthesis (in mitochondria) to other locations in cells. But while there is detailed information on how the heme lifecycle begins (heme synthesis), and how it ends (heme degradation), what happens in between is largely a mystery. Here we summarize recent information on the quantification of heme in cells, and we present a discussion of a mechanistic framework that could meet the logistical challenge of heme distribution.

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“…However, while the machinery for heme synthesis and degradation is well known, a decades-old question has been to establish precisely how heme is transported between its place of synthesis and subsequently made available to other regions of the cell where heme is in demand. Recent published work has hypothesized that membrane structures ( 3 ) and membrane contacts ( 4 ) are involved in the heme trafficking mechanism. Nevertheless, the scarcity of information in this area stands in stark contrast to the extensive efforts that have been directed toward understanding the structures and reactivities of many different heme proteins (e.g., refs.…”

mentioning

confidence: 99%

Unravelling the mechanisms controlling heme supply and demand

Leung

Fung

Gallio

et al. 2021

Proc. Natl. Acad. Sci. U.S.A.

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In addition to heme’s role as the prosthetic group buried inside many different proteins that are ubiquitous in biology, there is new evidence that heme has substantive roles in cellular signaling and regulation. This means that heme must be available in locations distant from its place of synthesis (mitochondria) in response to transient cellular demands. A longstanding question has been to establish the mechanisms that control the supply and demand for cellular heme. By fusing a monomeric heme-binding peroxidase (ascorbate peroxidase, mAPX) to a monomeric form of green-fluorescent protein (mEGFP), we have developed a heme sensor (mAPXmEGFP) that can respond to heme availability. By means of fluorescence lifetime imaging, this heme sensor can be used to quantify heme concentrations; values of the mean fluorescence lifetime (τMean) for mAPX-mEGFP are shown to be responsive to changes in free (unbound) heme concentration in cells. The results demonstrate that concentrations are typically limited to one molecule or less within cellular compartments. These miniscule amounts of free heme are consistent with a system that sequesters the heme and is able to buffer changes in heme availability while retaining the capability to mobilize heme when and where it is needed. We propose that this exchangeable supply of heme can operate using mechanisms for heme transfer that are analogous to classical ligand-exchange mechanisms. This exquisite control, in which heme is made available for transfer one molecule at a time, protects the cell against the toxic effect of excess heme and offers a simple mechanism for heme-dependent regulation in single-molecule steps.

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Glutathione and the intracellular labile heme pool

Cited by 14 publications

References 12 publications

Heme oxygenase-2 (HO-2) binds and buffers labile heme, which is largely oxidized, in human embryonic kidney cells

Heme oxygenase-2 (HO-2) binds and buffers labile heme, which is largely oxidized, in human embryonic kidney cells

Understanding the Logistics for the Distribution of Heme in Cells

Unravelling the mechanisms controlling heme supply and demand

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