EVIDENCE FOR PHOSPHORYLATION AND OLIGOMERIC ASSEMBLY OF PRESENILIN-1

Pathogenic mutations in presenilin 1 (PS1) are associated with approxi mate to 50% of early-onset familial Alzheimer disease, PS1 is endoprot eolytically cleaved to yield a 30-kDa N-terminal fragment (NTF) and an 18-kDa C-terminal fragment (CTF), Using COS7 cells transfected with h uman PS1, we have found that phorbol 12,13-dibutyrate and forskolin in crease the state of phosphorylation of serine residues of the human CT F, Phosphorylation of the human CTP resulted in a shift in electrophor etic mobility from a single major species of 18 kDa to a doubler of 20 -23 kDa, This mobility shift was also observed with human PS1 that had been transfected into mouse neuroblastoma (N2a) cells, Treatment of t he phosphorylated CTF doublet with phage lambda protein phosphatase el iminated the 20- to 23-kDa doubler while enhancing the 18-kDa species, consistent with the interpretation that the electrophoretic mobility shift was due to the addition of phosphate to the 18-kDa species, The NTF and CTF eluted from a gel filtration column at an estimated mass o f over 100 kDa, suggesting that these fragments exist as an oligomeriz ed species, Upon phosphorylation of the PS1 CTF, the apparent mass of the NTF- or CTF-containing oligomers was unchanged, Thus, the associat ion of PS1 fragments may be maintained during cycles of phosphorylatio n/dephosphorylation of the PS1 CTF.