Heat shock protein 70 inhibits hydrogen peroxide-induced nucleolar fragmentation via suppressing cleavage and down-regulation of nucleolin

It has been reported that nucleolar fragmentation is a part of the overall apoptotic morphology, however, it is currently obscure whether and how nucleolar fragmentation can be induced by hydrogen peroxide (H₂O₂) and heat shock protein 70 (Hsp70) can prevent nucleolar fragmentation. To dissect these two questions, C₂C₁₂ myogenic cells and immortalized mouse embryonic fibroblasts (MEFs) with heat shock transcriptional factor 1 (HSF1) null mutation were treated with heat shock response (HS) (42.5 ± 0.5°C for 1 h and recovery at 37°C for 24 h) and then were insulted with 0.5 mmol/L H₂O₂. Morphological changes of nucleoli were observed under contrast microscope or electronic microscope. It was found that (1) stimulation with H₂O₂-induced nucleolar fragmentation by mediating cleavage and down-regulation of nucleolar protein, nucleolin in C₂C₁₂ myocytes and MEFs; (2) HS suppressed nucleolar fragmentation by inducing the expression of Hsp70 in an HSF1-dependent manner as indicated by assays of transfection with Hsp70 antisense oligonucleotides (AS-ONs) or recombinant plasmids of full-length Hsp70 cDNA; (3) protection of Hsp70 against nucleolar fragmentation was related to its accumulation in nucleolus mediated by nuclear localization sequence and its inhibition against cleavage and down-regulation of nucleolin. These results suggested that H₂O₂-induced nucleolar fragmentation and HS or Hsp70 inhibit H₂O₂-induced nucleolar fragmentation through the translocation of Hsp70 into nucleolar and its protection against impairment of nucleolin.