Upregulation of clusterin/apolipoprotein J in lactacystin-treated SH-SY5Y cells.
Clusterin (apolipoprotein J) is a highly conserved, multifunctional, vertebrate glycoprotein. Several isoforms of clusterin have been described including the predominant secreted isoform (sCLU) and several nuclear isoforms (nCLU) associated with cell death. sCLU has been shown to bind a variety of partly unfolded, stressed proteins including those associated with Lewy bodies (LBs) in patients with Parkinson's disease (PD). The development of familial and sporadic PD has been associated with the ubiquitin-proteasome system (UPS) dysfunction and aberrant protein degradation. This suggests that failure of the UPS to degrade abnormal proteins may underlie nigral degeneration and LB formation in PD. The effects of toxin-mediated proteasomal impairment on changes in gene expression and cell viability were studied in differentiated SH-SY5Y cells. Clusterin expression was increased in cells exposed for 24 hr to the proteasomal inhibitor lactacystin (10 microM) as determined by gene microarray analysis. RT-PCR showed that sCLU, not nCLU, was the major clusterin isoform expressed in both control and lactacystin-treated cells. Western blot analysis identified statistically significant increases in sCLU in total cell lysates after 24 hr of lactacystin exposure and showed that sCLU fractionates with the endoplasmic reticulum. Time-course studies demonstrated that maximal decreases in proteasome activity (4 hr) preceded maximal increases in clusterin expression (24 hr). Together these data suggest that proteasome impairment results in the upregulation of sCLU in SH-SY5Y cells, supporting the hypothesis that the association of clusterin with LBs in PD may be related to UPS failure.
Carreras
Isabel
I
Department of Veterans Affairs, VA Medical Center, Bedford, Massachusetts 01730, USA.
Garrett-Young
Rosemary
R
Ullman
M David
MD
Eisenhauer
Patricia B
PB
Fine
Richard E
RE
Wells
John M
JM
Conn
Kelly J
KJ
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D000704:Analysis of Variance; D015153:Blotting, Western / Q000379:methods; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D051152:Clusterin; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects*; D006023:Glycoproteins / Q000378:metabolism*; D006801:Humans; D046228:Microarray Analysis / Q000379:methods; D008954:Models, Biological; D018832:Molecular Chaperones / Q000378:metabolism*; D009447:Neuroblastoma; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism; D013997:Time Factors; D015854:Up-Regulation / Q000187:drug effects
C495242:CLU protein, human; D051152:Clusterin; D015853:Cysteine Proteinase Inhibitors; D006023:Glycoproteins; D018832:Molecular Chaperones; D012333:RNA, Messenger; C067713:lactacystin; D046988:Proteasome Endopeptidase Complex; D000111:Acetylcysteine