Limits...
Phagosome maturation during endosome interaction revealed by partial rhodopsin processing in retinal pigment epithelium.

Wavre-Shapton ST, Meschede IP, Seabra MC, Futter CE - J. Cell. Sci. (2014)

Bottom Line: Loss of the cytoplasmic rhodopsin epitope was insensitive to pH but sensitive to protease inhibition and coincided with the interaction of phagosomes with endosomes.Thus, during pre-lysosomal maturation of ROS-containing phagosomes, limited rhodopsin processing occurs upon interaction with endosomes.This potentially provides a sensitive readout of phagosome-endosome interactions that is applicable to multiple phagocytes.

View Article: PubMed Central - PubMed

Affiliation: Molecular Medicine Section, National Heart and Lung Institute, Imperial College London, London SW7 2AZ, UK UCL Institute of Ophthalmology, University College London, London EC1V 9EL, UK.

Show MeSH

Related in: MedlinePlus

Synchronised loss of the C-terminal cytoplasmic rhodopsin epitope correlates with phagosome position in the RPE. (A) The positions of the phagosomes analysed in Fig. 1E were scored as apical or basal depending on whether the phagosome was above or below the tight junctions, respectively. Phagosomes from both time-points were pooled together for this analysis, as their position according to the labelling does not vary with time. Data are expressed as a percentage of total double-labelled phagosomes (black bars) or total single-labelled phagosomes (white bars). A total of 313 phagosomes were analysed in eight eyes. Data show the mean±s.e.m. (B) In the same population of phagosomes as in A, the density of PAG associated with 1D4 was calculated in phagosomes and outer segments. 1D4 density in >60 ROS and >140 phagosomes was calculated in total and normalised to outer segment density in each sample. Four eyes were analysed for each time-point. The thin horizontal black lines show the mean in each sample; ***P<0.0001.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4150067&req=5

f02: Synchronised loss of the C-terminal cytoplasmic rhodopsin epitope correlates with phagosome position in the RPE. (A) The positions of the phagosomes analysed in Fig. 1E were scored as apical or basal depending on whether the phagosome was above or below the tight junctions, respectively. Phagosomes from both time-points were pooled together for this analysis, as their position according to the labelling does not vary with time. Data are expressed as a percentage of total double-labelled phagosomes (black bars) or total single-labelled phagosomes (white bars). A total of 313 phagosomes were analysed in eight eyes. Data show the mean±s.e.m. (B) In the same population of phagosomes as in A, the density of PAG associated with 1D4 was calculated in phagosomes and outer segments. 1D4 density in >60 ROS and >140 phagosomes was calculated in total and normalised to outer segment density in each sample. Four eyes were analysed for each time-point. The thin horizontal black lines show the mean in each sample; ***P<0.0001.

Mentions: To determine whether the loss of the cytoplasmic rhodopsin epitope correlated with phagosome movement from the apical to the basal region of the cell, the position of phagosomes in the RPE in relation to their labelling for both rhodopsin antibodies was analysed. As shown in Fig. 2A, >90% of early phagosomes (1D4 positive, RET-P1 positive) were apical (at or above the tight junctions), whereas late phagosomes (1D4 negative, RET-P1 positive) could be both apical and basal (below the tight junctions), although most (>75%) were basal. This suggests that the loss of the cytoplasmic epitope of rhodopsin occurs early in the maturation process and might precede movement to the basal region.


Phagosome maturation during endosome interaction revealed by partial rhodopsin processing in retinal pigment epithelium.

Wavre-Shapton ST, Meschede IP, Seabra MC, Futter CE - J. Cell. Sci. (2014)

Synchronised loss of the C-terminal cytoplasmic rhodopsin epitope correlates with phagosome position in the RPE. (A) The positions of the phagosomes analysed in Fig. 1E were scored as apical or basal depending on whether the phagosome was above or below the tight junctions, respectively. Phagosomes from both time-points were pooled together for this analysis, as their position according to the labelling does not vary with time. Data are expressed as a percentage of total double-labelled phagosomes (black bars) or total single-labelled phagosomes (white bars). A total of 313 phagosomes were analysed in eight eyes. Data show the mean±s.e.m. (B) In the same population of phagosomes as in A, the density of PAG associated with 1D4 was calculated in phagosomes and outer segments. 1D4 density in >60 ROS and >140 phagosomes was calculated in total and normalised to outer segment density in each sample. Four eyes were analysed for each time-point. The thin horizontal black lines show the mean in each sample; ***P<0.0001.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4150067&req=5

f02: Synchronised loss of the C-terminal cytoplasmic rhodopsin epitope correlates with phagosome position in the RPE. (A) The positions of the phagosomes analysed in Fig. 1E were scored as apical or basal depending on whether the phagosome was above or below the tight junctions, respectively. Phagosomes from both time-points were pooled together for this analysis, as their position according to the labelling does not vary with time. Data are expressed as a percentage of total double-labelled phagosomes (black bars) or total single-labelled phagosomes (white bars). A total of 313 phagosomes were analysed in eight eyes. Data show the mean±s.e.m. (B) In the same population of phagosomes as in A, the density of PAG associated with 1D4 was calculated in phagosomes and outer segments. 1D4 density in >60 ROS and >140 phagosomes was calculated in total and normalised to outer segment density in each sample. Four eyes were analysed for each time-point. The thin horizontal black lines show the mean in each sample; ***P<0.0001.
Mentions: To determine whether the loss of the cytoplasmic rhodopsin epitope correlated with phagosome movement from the apical to the basal region of the cell, the position of phagosomes in the RPE in relation to their labelling for both rhodopsin antibodies was analysed. As shown in Fig. 2A, >90% of early phagosomes (1D4 positive, RET-P1 positive) were apical (at or above the tight junctions), whereas late phagosomes (1D4 negative, RET-P1 positive) could be both apical and basal (below the tight junctions), although most (>75%) were basal. This suggests that the loss of the cytoplasmic epitope of rhodopsin occurs early in the maturation process and might precede movement to the basal region.

Bottom Line: Loss of the cytoplasmic rhodopsin epitope was insensitive to pH but sensitive to protease inhibition and coincided with the interaction of phagosomes with endosomes.Thus, during pre-lysosomal maturation of ROS-containing phagosomes, limited rhodopsin processing occurs upon interaction with endosomes.This potentially provides a sensitive readout of phagosome-endosome interactions that is applicable to multiple phagocytes.

View Article: PubMed Central - PubMed

Affiliation: Molecular Medicine Section, National Heart and Lung Institute, Imperial College London, London SW7 2AZ, UK UCL Institute of Ophthalmology, University College London, London EC1V 9EL, UK.

Show MeSH
Related in: MedlinePlus