Identification of proteins differentially expressed in quiescent and proliferatively senescent fibroblast cultures.
We have examined the differential expression of proteins in quiescent (nongrowing) early versus late population doubling level (PDL) WI-38 human diploid fibroblasts. Age-associated changes in gene expression at the level of both secreted and total cellular protein were identified using high-resolution, two-dimensional gel electrophoresis followed by N-terminal sequencing or Western blot analysis. A comparison of secreted proteins revealed the senescence-specific absence of all forms of EPC-1, a protein associated with early PDL cells in the G0 state. This comparison also revealed the absence of a processed (cleaved) form of alpha I (type III) procollagen in senescent cells, suggesting a distinctive remodeling of the extracellular matrix. A comparison of total cellular protein between early passage and proliferatively aged fibroblasts identified an altered form of the enzyme ubiquitin carboxyl-terminal hydrolase in senescent cells, a result that could explain the increase in ubiquitinated proteins in these cells. These findings further elucidate the phenotype of fibroblasts aged in vitro and support the concept that senescent cells are arrested in a state fundamentally different from that of G0 early PDL cells.
DiPaolo
B R
BR
Center for Gerontological Research, Medical College of Pennsylvania, Philadelphia 19129, USA.
Pignolo
R J
RJ
Cristofalo
V J
VJ
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002453:Cell Cycle / Q000502:physiology*; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000502:physiology*; D015180:Electrophoresis, Gel, Two-Dimensional; D005136:Eye Proteins; D005347:Fibroblasts / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D009414:Nerve Growth Factors; D011347:Procollagen / Q000032:analysis; D011506:Proteins / Q000032:analysis* / Q000378:metabolism; D015843:Serpins / Q000032:analysis; D013869:Thiolester Hydrolases / Q000032:analysis; D043222:Ubiquitin Thiolesterase; D014452:Ubiquitins / Q000378:metabolism; D000099174:Pigment Epithelium-Derived Factor
D005136:Eye Proteins; D009414:Nerve Growth Factors; D011347:Procollagen; D011506:Proteins; D015843:Serpins; D013869:Thiolester Hydrolases; D043222:Ubiquitin Thiolesterase; D014452:Ubiquitins; D000099174:Pigment Epithelium-Derived Factor