Figure 6.

HSP101 enhancement is specific to Ω-containing mRNAs and is not dependent on a poly(A) tail. (A) In vitro-synthesized luc mRNA constructs terminating with or without a poly(A) tail or with the TMV 3′UTR (indicated in each panel) were electroporated into SL304A (containing either pGAL1–NtHSP101 or pYES2 as indicated at left) and expression measured following the completion of translation. The expression ratio is shown at right. (B) Ω-luc-A₅₀ and luc-A₅₀ mRNAs were introduced into SGM-grown, SL304A(pGAL1–NtHSP101), SL304A(pGAL1–TaHSP101), or SL304A(pYES2) by electroporation and expression measured following the completion of translation. The expression ratio is shown at right. (C) NtHSP101 increases the rate of translation from an Ω-containing mRNA. Ω-luc-A₅₀ and luc-A₅₀ mRNAs were introduced into SGM-grown, SL304A(pGAL1–NtHSP101) cells by electroporation, luciferase expression measured following RNA delivery, and the data plotted as a function of the time of translation. (D) NtHSP101 does not regulate translation from TEV 5′ leader-containing mRNA. TEV-luc-A₅₀, Ω-luc-A₅₀, or luc-A₅₀ mRNAs were electroporated into SL304A containing either pGAL1–NtHSP101 or pYES2. Expression is shown relative to the level obtained from each construct in SL304A(pYES2).