Increase of the incubation temperature of HeLa cells to 40°C for 5 mins followed by recovery of the cells at the normal growth temperature, results in the induction of three particular protein bands (heat-shock protein bands) of molecular weights 100,000, 72,000-74,000 and 37,000 daltons. These protein bands are resolved into more than one polypeptides each. Maximum synthesis of these heat-shock polypeptides takes place 2 hrs after the recovery from the heat-shock treatment and declines thereafter. The induction of the heat-shock polypeptides is blocked by actinomycin D, suggesting that their production may be regulated at the transcriptional level. Heat-shock treatment of HeLa cells results in degradation of pre-existing polysomes. Normal polysomal profile is obtained 1-2 hrs after the heat-shock treatment. mRNA synthesis is slightly affected during the 2 hrs after the heat shock as judged by the incorporation of [³H]-uridine into poly(A)⁺ mRNA molecules. In vitro translation, using a rabbit reticulocyte lysate cell-free protein synthesising system, of polysomal nRNA from HeLa and Friend cells has shown that a number of polypeptides are encoded by both poly (A)⁺ and poly (A)⁻ mRNAs. Using the same method, it was shown that the prominent 72,000-74,000 heat-shock protein band is encoded by seven mRNA species, all of which exist in both poly (A)⁺ and poly(A)⁻ forms. The heat-shock polypeptides encoded by these seven mRNA species are designated as α, α', β, γ, δ, ε, and ζ.