A Genetically Encoded FRET Sensor for Intracellular Heme

Song, Yanqun; Yang, Mao; Wegner, Seraphine V.; Zhao, Jingyi; Zhu, Rongfeng; Wu, Yun; He, Chuan; Chen, Peng R.

doi:10.1021/cb5009734

Cited by 70 publications

(100 citation statements)

References 22 publications

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“…For instance, a reevaluation of the heme dissociation constants of the HRM-containing transcription factor Rev-erbβ found that heme binds 100-fold tighter than previously estimated, with K d III and K d II of ∼20 nM vs. 2-6 μM (20). Our observation that subcellular [LH] is heterogeneous stands in striking contrast to a recent report by He and coworkers (19) that used a heme chaperone-based FRET sensor (CISDY-9) for heme in human cell lines. In that work, cytosolic, mitochondrial, and nuclear heme were all reported to be ∼25 nM.…”

Section: Discussioncontrasting

confidence: 95%

“…One variant, HS1-M7A, which was generated by replacing the heme axial Met 7 II value of HS1-M7A is similar in magnitude to previous estimates of the "regulatory" heme pool, 10-100 nM (12,19), as well as the affinities of proteins that may respond to this pool, including the heme-dependent transcription factor Rev-erbβ (20) and constitutive heme oxygenase-2 (HO-2) (21) (SI Appendix, Fig. S3G).…”

Section: Resultsmentioning

confidence: 84%

“…For quantitative heme monitoring, an in situ method to calibrate the sensor was developed (19). The concentration of [heme] accessible to the sensor is governed by the following expression (23):…”

Section: Resultsmentioning

confidence: 99%

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Heme dynamics and trafficking factors revealed by genetically encoded fluorescent heme sensors

Hanna

Harvey

Martinez-Guzman

et al. 2016

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

168

405

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Heme is an essential cofactor and signaling molecule. Heme acquisition by proteins and heme signaling are ultimately reliant on the ability to mobilize labile heme (LH). However, the properties of LH pools, including concentration, oxidation state, distribution, speciation, and dynamics, are poorly understood. Herein, we elucidate the nature and dynamics of LH using genetically encoded ratiometric fluorescent heme sensors in the unicellular eukaryote Saccharomyces cerevisiae. We find that the subcellular distribution of LH is heterogeneous; the cytosol maintains LH at ∼20–40 nM, whereas the mitochondria and nucleus maintain it at concentrations below 2.5 nM. Further, we find that the signaling molecule nitric oxide can initiate the rapid mobilization of heme in the cytosol and nucleus from certain thiol-containing factors. We also find that the glycolytic enzyme glyceraldehyde phosphate dehydrogenase constitutes a major cellular heme buffer, and is responsible for maintaining the activity of the heme-dependent nuclear transcription factor heme activator protein (Hap1p). Altogether, we demonstrate that the heme sensors can be used to reveal fundamental aspects of heme trafficking and dynamics and can be used across multiple organisms, including Escherichia coli, yeast, and human cell lines.

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Section: Discussioncontrasting

confidence: 95%

Section: Resultsmentioning

confidence: 84%

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Heme dynamics and trafficking factors revealed by genetically encoded fluorescent heme sensors

Hanna

Harvey

Martinez-Guzman

et al. 2016

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

168

405

View full text Add to dashboard Cite

show abstract

“…20,21 Another strategy for heme imaging in live cells is based on the principle that association of heme either enhances or attenuates the fluorescence resonance energy transfer (FRET) process or quenches the fluorophore. 22–25 Typically, these sensors recognize only labile heme. 22,23,25 Moreover, the FRET sensors could perturb normal heme homeostasis by sequestering heme and distorting labile heme.…”

Section: ■ Introductionmentioning

confidence: 99%

Label-Free Imaging of Heme Dynamics in Living Organisms by Transient Absorption Microscopy

Chen

Yuan

et al. 2018

Anal. Chem.

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Heme, a hydrophobic and cytotoxic macrocycle, is an essential cofactor in a large number of proteins and is important for cell signaling. This must mean that heme is mobilized from its place of synthesis or entry into the cell to other parts of the cell where hemoproteins reside. However, the cellular dynamics of heme movement is not well understood, in large part due to the inability to image heme noninvasively in live biological systems. Here, using high-resolution transient absorption microscopy, we showed that heme storage and distribution is dynamic in Caenorhabditis elegans. Intracellular heme exists in concentrated granular puncta which localizes to lysosomal-related organelles. These granules are dynamic, and their breaking down into smaller granules provides a mechanism by which heme stores can be mobilized. Collectively, these direct and noninvasive dynamic imaging techniques provide new insights into heme storage and transport and open a new avenue for label-free investigation of heme function and regulation in living systems.

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“…Dual-color photoswitchable fluorescent (DCPF) nanoparticles represent a new class of materials for exploitation of fluorescent technique in biological [1][2][3]. These fluorescent nanoparticles, based on fluorescence resonance energy transfer (FRET) phenomenon, have received many attentions because of some advantages over conventional fluorescent dyes as distinguishing fake positive signals by aberrant fluorescent biomolecules and targeted locations [4,5].…”

Section: Introductionmentioning

confidence: 99%

FRET-based acrylic nanoparticles with dual-color photoswitchable properties in DU145 human prostate cancer cell line labeling

Rad

Mahdavian

Khoei

et al. 2016

Polymer

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A Genetically Encoded FRET Sensor for Intracellular Heme

Cited by 70 publications

References 22 publications

Heme dynamics and trafficking factors revealed by genetically encoded fluorescent heme sensors

Heme dynamics and trafficking factors revealed by genetically encoded fluorescent heme sensors

Label-Free Imaging of Heme Dynamics in Living Organisms by Transient Absorption Microscopy

FRET-based acrylic nanoparticles with dual-color photoswitchable properties in DU145 human prostate cancer cell line labeling

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