Normal human fibroblasts have a finite lifespan in culture termed replicative senescence which has long been interpreted as a model of aging at the cellular level. One pathway responsible for initiating cellular replication is known as the extracellular-regulated kinase (ERK) pathway which utilizes a number of mitogen-activated protein kinases (MAPKs) in order to transduce signals to the nucleus that are necessary for cellular replication. A lynchpin in this process is phosphorylation of ERK (a MAPK) and its translocation to the nucleus. Senescent cells have been found to have lower levels of active phospho-ERK in the nucleus. ERK’ s corresponding phosphatase, MAPK-phosphatase-2 (MKP-2), was found to be at higher levels in senescent cells. MKP-2 is degraded via the proteasome, a large proteolytic molecule. Using low and high passage human fibroblasts we investigated MKP-2 disposal and found that it is ubiquitinated prior to its degradation by the proteasome. Using immunofluorescence we found that the cellular location of ubiquitin does not vary with the age of the cells, but that the total amount of ubiquitinated protein does vary with age. Also, the important ubiquitin conjugating enzyme (E2) was found to be decreased in old cells. All of this evidence indicates that much of the phenomenon of aging at the cellular level may owe to important differences in the efficiency of protein disposal, and continued research in this area may lead to therapeutic targets to lessen the effects of aging.
Phillip Bilderback, ’06 Gig Harbor, WA
Major: Biochemistry and Molecular Biology
Sponsor: Craig Tepper