Expression of human HSP70 during the synthetic phase of the cell cycle.

Abstract

Expression of the major heat shock and stress-induced protein, HSP70, is under complex regulatory control in human cells. In addition to being induced by physiological stress such as heat shock or transition metals, the HSP70 gene is induced by serum stimulation and immortalizing products of the adenovirus ElA 13S and polyoma large tumor antigen genes. Here we show that expression of the human HSP70 gene is tightly regulated during the cell cycle. Using selective mitotic detachment, a noninductive method to obtain synchronous populations of HeLa cells, we show that levels of HSP7O mRNA rapidly increase 10-to 15-fold upon entry into S phase and decline by late S and G2. A transient increase in HSP70 synthesis is detected during early S phase. The subcellular localization of HSP70 varies throughout the cell cycle; the protein is diffusely distributed in the nucleus and cytoplasm in G1, localized in the nucleus in S, and again diffusely distributed in G2 cells. We suggest that the temporal pattern of HSP7O expression during S phase, the nuclear localization, and activation by trans-acting immortalizing proteins indicate a role for HSP7O in the nucleus of replicating cells.The cellular transition from the resting state to the growing state requires expression of certain genes whose products regulate critical events during the Go and G, phases of the cell cycle. Serum stimulation has been widely used as a method to identify and examine genes that are growth-regulated. The addition of fresh serum to quiescent mammalian cells activates the expression of a family of growth-regulated genes, including c-fos, c-myc, P-interferon, proliferin, p53, and HSP70 (1-10). Many of these genes are activated by specific growth factors such as platelet-derived growth factor or epidermal growth factor, suggesting the possibility that their expression may be regulated by common mechanisms (1, 2, 7).Workers in our laboratory have studied the expression of the major heat shock protein, HSP70, in human cells. An intriguing relationship between the expression of heat shock proteins and cellular transformation has been established. HSP70 is expressed at high levels in transformed mammalian cells and is induced in cells infected with DNA tumor viruses (11-15). Expression from the HSP70 promoter is induced by the ElA 13S product of adenovirus and by large tumor antigen of polyoma (15,16 We have demonstrated (10) that HSP70 gene expression in a transformed human cell line, HeLa, was induced by serum stimulation. To examine whether this corresponds to expression at a specific point during the cell cycle or to activation by specific growth factors, we have used a noninductive method-that of selective mitotic detachment-to obtain synchronous populations of cells. We show that maximal levels of HSP70 mRNA are detected in S phase at the peak of DNA synthesis. HSP70 protein synthesis increases transiently at this point and increased levels of HSP70 are detected in the nucleus of S-phase cells. METHODSCell Culture and Synchronization. HeLa cel...

Selective induction of human heat shock gene transcription by the adenovirus E1A gene products, including the 12S E1A product.

“…Maximal hsp7O expression was observed during the late S-early G2 phase of the cell cycle (18). Similar results have been obtained after serum stimulation (56) and synchronization by selective mitotic detachment (29) of HeLa cells, although the peak of induction was somewhat earlier. Because the hsp89ao and hsp7O genes are both induced by ElA, we determined whether the hsp89a gene is also regulated during the cell cycle.…”

Phase-Specific Polypeptides and Poly(A)+ RNAs during the Cell Cycle in Synchronous Cultures of Catharanthus roseus Cells

“…However, the rates of synthesis of cytoplasmic polypeptides showed no characteristic changes in the G2 phase (Table II). These results suggest that posttranslational processing, including modifications, degradations, reconstructions, and changes in intracellular localization of proteins (28) occurred during the G2 phase.…”

“…In the case of mammalian cells and yeast, there have been, until recently, no reports of proteins whose synthesis is limited to a specific phase of the cell cycle. However, the expression of cellular proteins throughout the cell cycle has recently been studied in more detail, and a few proteins whose synthesis is cell-cycle-dependent, such as 'dividin' (10), HSP70 (28), and a pair of 21 kD proteins (41), have been reported in mammalian cells. Furthermore, proliferating cell nuclear antigen (PCNA, also called cyclin), whose synthesis increases in the S phase (8), has been shown to be identical to a DNA polymerase-6 auxiliary protein (9,32).…”

Interaction of canine distemper virus nucleocapsid variants with 70K heat-shock proteins

“…The 70K and 90K hsps are the major heat-shock protein families expressed by mammalian cells (Welch et al, 1982). The predominant 70K hsps in primate and non-primate cells include a constitutively expressed 73K hsp (Welch et al, 1983) and a 72K hsp (inducible in non-primate cells and inducible and constitutive in primate cells) whose expression is cell cycle-regulated (Milarski & Morimoto, 1986). An important function of 70K hsps is mediation of recovery from stress (Riabowol et al, 1988) thought to occur by facilitation of protein renaturation (Pelham, 1986).…”