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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.

No MeSH data available.


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DT criterion to determine Tf-endosome–mitochondria interactions. (A, a–c) Representative confocal z stack image of MDCK-PTR cells pulsed with AF647-Tf (red) for 2 min, chased, fixed, and immunolabeled with AF555 anti-OMM (green) and AF488 anti-IMM (cyan). (d–f) Corresponding Imaris-based 3D-rendered images depicting OMM and IMM by surface and early Tf-endosomes by spots. Bars, 10 µm. (B, a–c) Magnified ROIs (dotted boxes in A, d–f) where the Tf-endosome under consideration (yellow spots) was in close proximity to the OMM (a) and subsequently validated by DT = 0. Upon deselecting the OMM and selecting the IMM surface render, the Tf-endosome showed failed interaction with the IMM of the same mitochondrion (b), also confirmed by a DT > 0 value with respect to the IMM surface; merged image (c). (d–f) Magnified ROIs (solid boxes in A, d–f) where another Tf-endosome (yellow spots) was identified to be in close proximity to both OMM (d) and IMM (e). Bars, 1 µm. The brightness and contrast of images have been enhanced equally across each panel to aid visualization. (C) Percentage of Tf-endosomes detected in close proximity to OMM or IMM per z stack of an image field (four to seven cells per field).
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fig2: DT criterion to determine Tf-endosome–mitochondria interactions. (A, a–c) Representative confocal z stack image of MDCK-PTR cells pulsed with AF647-Tf (red) for 2 min, chased, fixed, and immunolabeled with AF555 anti-OMM (green) and AF488 anti-IMM (cyan). (d–f) Corresponding Imaris-based 3D-rendered images depicting OMM and IMM by surface and early Tf-endosomes by spots. Bars, 10 µm. (B, a–c) Magnified ROIs (dotted boxes in A, d–f) where the Tf-endosome under consideration (yellow spots) was in close proximity to the OMM (a) and subsequently validated by DT = 0. Upon deselecting the OMM and selecting the IMM surface render, the Tf-endosome showed failed interaction with the IMM of the same mitochondrion (b), also confirmed by a DT > 0 value with respect to the IMM surface; merged image (c). (d–f) Magnified ROIs (solid boxes in A, d–f) where another Tf-endosome (yellow spots) was identified to be in close proximity to both OMM (d) and IMM (e). Bars, 1 µm. The brightness and contrast of images have been enhanced equally across each panel to aid visualization. (C) Percentage of Tf-endosomes detected in close proximity to OMM or IMM per z stack of an image field (four to seven cells per field).

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
DT criterion to determine Tf-endosome–mitochondria interactions. (A, a–c) Representative confocal z stack image of MDCK-PTR cells pulsed with AF647-Tf (red) for 2 min, chased, fixed, and immunolabeled with AF555 anti-OMM (green) and AF488 anti-IMM (cyan). (d–f) Corresponding Imaris-based 3D-rendered images depicting OMM and IMM by surface and early Tf-endosomes by spots. Bars, 10 µm. (B, a–c) Magnified ROIs (dotted boxes in A, d–f) where the Tf-endosome under consideration (yellow spots) was in close proximity to the OMM (a) and subsequently validated by DT = 0. Upon deselecting the OMM and selecting the IMM surface render, the Tf-endosome showed failed interaction with the IMM of the same mitochondrion (b), also confirmed by a DT > 0 value with respect to the IMM surface; merged image (c). (d–f) Magnified ROIs (solid boxes in A, d–f) where another Tf-endosome (yellow spots) was identified to be in close proximity to both OMM (d) and IMM (e). Bars, 1 µm. The brightness and contrast of images have been enhanced equally across each panel to aid visualization. (C) Percentage of Tf-endosomes detected in close proximity to OMM or IMM per z stack of an image field (four to seven cells per field).
© Copyright Policy - openaccess
Related In: Results  -  Collection

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fig2: DT criterion to determine Tf-endosome–mitochondria interactions. (A, a–c) Representative confocal z stack image of MDCK-PTR cells pulsed with AF647-Tf (red) for 2 min, chased, fixed, and immunolabeled with AF555 anti-OMM (green) and AF488 anti-IMM (cyan). (d–f) Corresponding Imaris-based 3D-rendered images depicting OMM and IMM by surface and early Tf-endosomes by spots. Bars, 10 µm. (B, a–c) Magnified ROIs (dotted boxes in A, d–f) where the Tf-endosome under consideration (yellow spots) was in close proximity to the OMM (a) and subsequently validated by DT = 0. Upon deselecting the OMM and selecting the IMM surface render, the Tf-endosome showed failed interaction with the IMM of the same mitochondrion (b), also confirmed by a DT > 0 value with respect to the IMM surface; merged image (c). (d–f) Magnified ROIs (solid boxes in A, d–f) where another Tf-endosome (yellow spots) was identified to be in close proximity to both OMM (d) and IMM (e). Bars, 1 µm. The brightness and contrast of images have been enhanced equally across each panel to aid visualization. (C) Percentage of Tf-endosomes detected in close proximity to OMM or IMM per z stack of an image field (four to seven cells per field).
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–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.

No MeSH data available.


Related in: MedlinePlus