Background
Mesencephalic astrocyte-derived neurotrophic factor (MANF) is a new candidate growth factor for dopaminergic neurons against endoplasmic reticulum stress (ER stress). HSP70 family, a chaperon like heat shock protein family, was proved to be involved in the MANF induced survival pathway in 6-OHDA treated SHSY-5Y cells. However, the ER stress relative transcriptome, in MANF signaling cascades is still investigated. The involvement of HSP70, a 70kd member of HSP70 family, need further to be verified.
Conclusion
MANF exerts a protective role against 6-OHDA induced apoptosis in SHSY-5Y cells via up-regulating some ER stress genes, including HSP70 family members. The HSP70 expression level plays a key role in MANF-mediated survival pathway.
Methods
The cell apoptosis was assayed by MTT, TUNEL staining and western blot of cleaved Caspase-3. The differentially expressed genes in SHSY-5Y cells under different conditions (control, 6-OHDA, 6-OHDA + MANF) were investigated by RNA-seq. Expression of HSP70 was further confirmed by real-time PCR. RNAi knockdown for HSP70 was performed to investigate the role of HSP70 in the MANF signaling pathway.
Results
MANF inhibits 6-OHDA-induced apoptosis in SHSY-5Y cells. Six ER stress relative genes (HSP70, GRP78, xbp-1, ATF-4, ATF-6, MAPK) were found enriched in 6-OHDA + MANF treatment group. HSP70 was the most significantly up-regulated gene under 6-OHDA + MANF treatment in SHSY-5Y cells. RNAi knockdown for HSP70 inhibits the protective effects of MANF against 6-OHDA toxicity in SHSY-5Y cells.