Initial characterization of heat‐induced excess nuclear proteins in HeLa cells

Abstract

Exposure of mammalian cells to hyperthermia is known to cause protein aggregation in the nucleus. The presence of such aggregates has been detected as the relative increase in the protein mass that is associated with nuclei isolated from heated cells. We have characterized these excess nuclear proteins from the nuclei of heated HeLa cells by two-dimensional gel electrophoresis. The abundance of cytoskeletal elements which co-purify with the nuclei did not increase with exposure to hyperthermia, indicating that these proteins are not part of the excess nuclear proteins. In contrast, several specific polypeptides become newly bound or increase in abundance in nuclei isolated from heated cells. Members of the hsp 70 family were identified as a major component of the excess nuclear proteins. Among the other excess nuclear proteins we identified ten that had apparent molecular weights of 130, 95, 75, 58, 53, 48, 46, 37, 28, and 26 kilodaltons. Since hsp 70 is mainly cytoplasmic in non-heated cells, its association with nuclei in heated cells indicates that one mechanism accounting for the heat-induced excess nuclear proteins is the movement of cytoplasmic proteins to the nucleus. We also obtained evidence that increased binding of nuclear proteins is another mechanism for this effect. No overall increase or decrease in the phosphorylation of nuclear proteins was found to be associated with such altered binding or movement from the cytoplasm to the nucleus.

Modulation of Cellular Thermoresistance and Actin Filament Stability Accompanies Phosphorylation-Induced Changes in the Oligomeric Structure of Heat Shock Protein 27

“…It is conceivable that the insolubilization and translocation of HSP27 during heat shock simply reflect the protein denaturing effect of heat. A large number of other proteins are similarly insolubilized and associate with nuclear structures during heat shock (13,23,24,33,45,54).…”

Alterations in nuclear matrix ultrastructure of G1 mammalian cells following heat shock: Resinless section electron microscopy, biochemical, and immunofluorescence studies

“…For example, the proteins belonging to the 70 kD heat shock protein group, HSP 70, which are synthesized as a result of heat shock, become translocated from cytoplasm to nucleoli following heat shock (Welch and Suhan, 1986). In a recent two-dimensional electrophoresis study, Laszlo et al (1992) (Ornelles and Penman, 1990) as well as mammalian cells (Reiter and Penman, 1983). In particular, Ornelles and Penman (1990) reported a correlation between alterations in nuclear matrix morphology and the translocation of prompt heat shock proteins into the nuclear matrix-IF fraction in Drosophila cells.…”

Lamin B is a prompt heat shock protein

“…Recent evidence suggests that the nuclear matrix represents a critical target for heat cytotoxicity and that nuclear matrix and nuclear matrix-associated proteins figure prominently in the response to hyperthermia (Warters et al, 1986;Laszlo et al, 1992;Roti Roti and Turkel, 1994;VanderWaal et al, 1996;Roti Roti et al, 1997). Hyperthermia causes changes in nuclear matrix architecture (Wachsberger and Coss, 1993), and these structural changes, as well as changes in the abundance and conformation of specific nuclear matrix proteins, could be critical for the continuation or resumption of nuclear metabolism after heat shock (Henle and Leeper, 1979;Wong and Dewey, 1982).…”