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Down-regulation of receptor-mediated endocytosis is responsible for senescence-associated hyporesponsiveness
Sang Chul Park1, Jeong-Soo Park, Woong-Yang Park
1Department of Biochemistry and Molecular Biology, Seoul National University College of Medicine, Seoul, South Korea. scpark@snu.ac.kr
Abstract:
Human diploid fibroblasts (HDF) do not divide indefinitely and eventually lead to an arrest of cell division by a process termed cellular or replicative senescence. Irreversible growth arrest of senescent cells is strongly related to the attenuated response to growth factors. Recently, we reported that up-regulation of caveolin in the senescent cells is responsible for the attenuated response to growth factors. Senescent cells did not phosphorylate Erk-1/2 after EGF stimulation, whereas young cells did. In those senescent cells, we found an increased level of caveolin proteins and strong interactions between caveolin-1 and EGFR. When we overexpressed caveolin-1 in young HDF, the activation of Erk-1/2 on EGF stimulation was significantly suppressed. These results suggest that the hyporesponsiveness of senescent fibroblasts to EGF stimulation might be due to the overexpression of caveolin. In addition, the clathrin-dependent endocytosis system plays the more active and dominant role over the caveolae system. Therefore, we monitored the efficiency of clathrin-dependent receptor-mediated endocytosis in the senescent cells in order to elucidate the exact mode of the attenuated response to growth factors in the senescent cells. Using a transferrin-uptake assay and Western blot analysis of endocytosis-related proteins, we found a significant decrease of amphiphysin-1 in human diploid fibroblasts of multipassages. By adjusting the level of amphiphysin, we could modulate the efficiency of receptor-mediated endocytosis either in young or old cells toward growth factors: that is, a dominant negative mutant of amphiphysin-1 blocked the endocytosis in the young cells, while microinjection of the gene resumed its activity in the old cells. Taken together, we conclude that the loss of endocytotic activity of senescent cells is directly related to the down-regulation of amphiphysin-1 and/or up-regulation of caveolins. This opens a new field of functional recovery of the senescent cells simply through adjusting the receptor-mediated endocytosis capacity.
Insights
Cellular senescence in human diploid fibroblasts involves decreased growth factor response due to increased caveolins and reduced amphiphysin-1, impacting endocytosis. Adjusting these proteins may restore cellular function.
Area of Science:
- Cell Biology
- Molecular Biology
- Gerontology
Background:
- Cellular senescence, or replicative senescence, is a state of irreversible growth arrest in human diploid fibroblasts (HDF).
- Senescent cells exhibit an attenuated response to growth factors, a key characteristic impacting their function.
- Previous research indicated a role for caveolin in this hyporesponsiveness.
Purpose of the Study:
- To investigate the molecular mechanisms underlying the attenuated growth factor response in senescent fibroblasts.
- To determine the specific roles of caveolin and endocytosis in cellular senescence.
- To explore potential strategies for functional recovery of senescent cells.
Main Methods:
- Overexpression of caveolin-1 in young HDF to assess its effect on Erk-1/2 phosphorylation after EGF stimulation.
- Analysis of clathrin-dependent endocytosis efficiency in senescent cells using transferrin-uptake assays.
- Western blot analysis of endocytosis-related proteins, specifically amphiphysin-1.
- Modulation of amphiphysin-1 levels in young and senescent cells.
Main Results:
- Senescent cells showed suppressed Erk-1/2 phosphorylation upon EGF stimulation, unlike young cells.
- Increased caveolin-1 levels and interaction with EGFR were observed in senescent cells.
- Overexpressing caveolin-1 in young HDF mimicked the suppressed response.
- A significant decrease in amphiphysin-1 was found in senescent HDF.
- Modulating amphiphysin-1 levels affected receptor-mediated endocytosis efficiency in both young and senescent cells.
Conclusions:
- The hyporesponsiveness of senescent fibroblasts to EGF stimulation is linked to caveolin overexpression.
- Loss of endocytotic activity in senescent cells is directly related to amphiphysin-1 down-regulation and/or caveolin up-regulation.
- Adjusting receptor-mediated endocytosis capacity offers a potential avenue for the functional recovery of senescent cells.