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Endosome – mitochondria interactions are modulated by iron release from transferrin

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ABSTRACT

Using superresolution and quantitative fluorescence microscopy, Das et al. have revealed that iron-transferrin–containing endosomes directly interact with mitochondria, facilitating iron transfer in epithelial cells. Their findings further enrich the repertoire of organelle–organelle direct interactions to accomplish a functional significance.

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Tf-endosome–mitochondria interactions facilitate functional iron transfer. MDCK-PTR cells with iron sensor RDA-labeled mitochondria were pulsed with AF647-Tf for 2 min at 37°C and chased, and time-lapse images of 15-s duration (107-ms interval) were acquired at 5 min after the Tf pulse to capture the early time points of Tf-endosomal trafficking. (A and B) Videomicrographs of distinct kiss interactions between 3D-rendered Tf-endosomes (white spots) and surface-rendered mitochondria (asterisks), proceeding from left to right at 107-ms intervals. The rendered mitochondrial color, indicated by the color bars, is based on the mitochondria’s mean fluorescence intensity. See Videos 3 and 4. Graphs represent Tf-endosomal interaction in terms of its distance from the mitochondrion (blue lines) and corresponding change in mean intensity of interacting (red lines) and noninteracting mitochondria (green lines). The dotted lines mark the beginning of the kiss interaction, after which the interacting mitochondrial mean intensity gradually quenches, whereas that of the noninteracting mitochondria remains unchanged. Bars, 0.5 µm. (C) Normalized mean intensity of interacting mitochondria (n = 12) is significantly reduced after the kiss interaction. (D) Mean intensity of noninteracting mitochondria (n = 31) remains unchanged over time. Error bars: 95% confidence interval; **, P < 0.001, Student’s t test. a.u., arbitrary units.
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fig3: Tf-endosome–mitochondria interactions facilitate functional iron transfer. MDCK-PTR cells with iron sensor RDA-labeled mitochondria were pulsed with AF647-Tf for 2 min at 37°C and chased, and time-lapse images of 15-s duration (107-ms interval) were acquired at 5 min after the Tf pulse to capture the early time points of Tf-endosomal trafficking. (A and B) Videomicrographs of distinct kiss interactions between 3D-rendered Tf-endosomes (white spots) and surface-rendered mitochondria (asterisks), proceeding from left to right at 107-ms intervals. The rendered mitochondrial color, indicated by the color bars, is based on the mitochondria’s mean fluorescence intensity. See Videos 3 and 4. Graphs represent Tf-endosomal interaction in terms of its distance from the mitochondrion (blue lines) and corresponding change in mean intensity of interacting (red lines) and noninteracting mitochondria (green lines). The dotted lines mark the beginning of the kiss interaction, after which the interacting mitochondrial mean intensity gradually quenches, whereas that of the noninteracting mitochondria remains unchanged. Bars, 0.5 µm. (C) Normalized mean intensity of interacting mitochondria (n = 12) is significantly reduced after the kiss interaction. (D) Mean intensity of noninteracting mitochondria (n = 31) remains unchanged over time. Error bars: 95% confidence interval; **, P < 0.001, Student’s t test. a.u., arbitrary units.

Mentions: To investigate the prevalence of endosome–mitochondria interactions within the early endocytic pathway, we established a criterion to quantitatively measure interactions in an unbiased and semiautomated manner. Previously, quantitative analyses of cell-to-cell (Malide et al., 2012; McKee et al., 2013) or organelle–organelle interactions (Whalen et al., 2012; Bouvet et al., 2013; Wang et al., 2015) were performed using the distance transformation (DT) algorithm. DT is a representation of the digital image in terms of pixels with assigned values based on their respective distances from the boundary of a specific object. Therefore, 3D rendering of the object in question is a prerequisite for carrying out the DT operation. Here, we used the DT algorithm, built in the image analysis software Imaris, for quantitative determination of the distance of Tf-endosomes from the boundary of 3D-rendered mitochondrial surfaces, which are considered to be the anchor organelle, as they are less dynamic than endosomes. The DT criterion was evaluated for its consistency and reliability on immunofluorescent fixed cells (Fig. 2) before it was used in the analysis of time-lapse videos to study the dynamics of endosome–mitochondria interactions in live cells (see Figs. 3, 4, and 5).


Endosome – mitochondria interactions are modulated by iron release from transferrin
Tf-endosome–mitochondria interactions facilitate functional iron transfer. MDCK-PTR cells with iron sensor RDA-labeled mitochondria were pulsed with AF647-Tf for 2 min at 37°C and chased, and time-lapse images of 15-s duration (107-ms interval) were acquired at 5 min after the Tf pulse to capture the early time points of Tf-endosomal trafficking. (A and B) Videomicrographs of distinct kiss interactions between 3D-rendered Tf-endosomes (white spots) and surface-rendered mitochondria (asterisks), proceeding from left to right at 107-ms intervals. The rendered mitochondrial color, indicated by the color bars, is based on the mitochondria’s mean fluorescence intensity. See Videos 3 and 4. Graphs represent Tf-endosomal interaction in terms of its distance from the mitochondrion (blue lines) and corresponding change in mean intensity of interacting (red lines) and noninteracting mitochondria (green lines). The dotted lines mark the beginning of the kiss interaction, after which the interacting mitochondrial mean intensity gradually quenches, whereas that of the noninteracting mitochondria remains unchanged. Bars, 0.5 µm. (C) Normalized mean intensity of interacting mitochondria (n = 12) is significantly reduced after the kiss interaction. (D) Mean intensity of noninteracting mitochondria (n = 31) remains unchanged over time. Error bars: 95% confidence interval; **, P < 0.001, Student’s t test. a.u., arbitrary units.
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fig3: Tf-endosome–mitochondria interactions facilitate functional iron transfer. MDCK-PTR cells with iron sensor RDA-labeled mitochondria were pulsed with AF647-Tf for 2 min at 37°C and chased, and time-lapse images of 15-s duration (107-ms interval) were acquired at 5 min after the Tf pulse to capture the early time points of Tf-endosomal trafficking. (A and B) Videomicrographs of distinct kiss interactions between 3D-rendered Tf-endosomes (white spots) and surface-rendered mitochondria (asterisks), proceeding from left to right at 107-ms intervals. The rendered mitochondrial color, indicated by the color bars, is based on the mitochondria’s mean fluorescence intensity. See Videos 3 and 4. Graphs represent Tf-endosomal interaction in terms of its distance from the mitochondrion (blue lines) and corresponding change in mean intensity of interacting (red lines) and noninteracting mitochondria (green lines). The dotted lines mark the beginning of the kiss interaction, after which the interacting mitochondrial mean intensity gradually quenches, whereas that of the noninteracting mitochondria remains unchanged. Bars, 0.5 µm. (C) Normalized mean intensity of interacting mitochondria (n = 12) is significantly reduced after the kiss interaction. (D) Mean intensity of noninteracting mitochondria (n = 31) remains unchanged over time. Error bars: 95% confidence interval; **, P < 0.001, Student’s t test. a.u., arbitrary units.
Mentions: To investigate the prevalence of endosome–mitochondria interactions within the early endocytic pathway, we established a criterion to quantitatively measure interactions in an unbiased and semiautomated manner. Previously, quantitative analyses of cell-to-cell (Malide et al., 2012; McKee et al., 2013) or organelle–organelle interactions (Whalen et al., 2012; Bouvet et al., 2013; Wang et al., 2015) were performed using the distance transformation (DT) algorithm. DT is a representation of the digital image in terms of pixels with assigned values based on their respective distances from the boundary of a specific object. Therefore, 3D rendering of the object in question is a prerequisite for carrying out the DT operation. Here, we used the DT algorithm, built in the image analysis software Imaris, for quantitative determination of the distance of Tf-endosomes from the boundary of 3D-rendered mitochondrial surfaces, which are considered to be the anchor organelle, as they are less dynamic than endosomes. The DT criterion was evaluated for its consistency and reliability on immunofluorescent fixed cells (Fig. 2) before it was used in the analysis of time-lapse videos to study the dynamics of endosome–mitochondria interactions in live cells (see Figs. 3, 4, and 5).

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Using superresolution and quantitative fluorescence microscopy, Das et al. have revealed that iron-transferrin&ndash;containing endosomes directly interact with mitochondria, facilitating iron transfer in epithelial cells. Their findings further enrich the repertoire of organelle&ndash;organelle direct interactions to accomplish a functional significance.

No MeSH data available.


Related in: MedlinePlus