The eIF2 alpha Kinases GCN2 and PERK Regulate Phosphorylation of TOR Target Proteins and Differential Translation of mRNAs

Open Access
Dang Do, An Ngoc
Graduate Program:
Integrative Biosciences
Doctor of Philosophy
Document Type:
Date of Defense:
June 15, 2006
Committee Members:
  • Leonard Shelton Jefferson Jr., Committee Chair
  • Scot R Kimball, Committee Member
  • Ralph Lauren Keil, Committee Member
  • Douglas Cavener, Committee Member
  • Robert A Gabbay, Committee Member
  • 2
  • cycloheximide
  • 4-di-tert-butylhydroquinone
  • translation initiation
  • translation elongation
  • eIF2
  • eIF2B
  • 4E-BP1
  • GCN2
  • p70S6K1
  • PERK
  • TOR
  • methionine deprivation
  • leucine deprivation
  • polysome
  • microarray
  • histidinol
  • leucinol
  • sucrose gradient
  • atf4
  • atf5
  • slc3a2
  • slc3a3
  • trfr
  • ldlr
  • stch
  • hspa5
  • igfbp-2
In eukaryotes, regulation of mRNA translation enables a fast, localized and finely tuned expression of gene products. Within the translation process, the first stage of translation initiation is most rigorously modulated by the actions of eukaryotic initiation factors (eIFs) and their associated proteins. These 11 eIFs catalyze the joining of the tRNA, mRNA and rRNA into a functional translation complex. Their activity is influenced by a wide variety of extra- and intracellular signals, ranging from global, such as hormone signaling and unfolded proteins, to specific, such as single amino acid imbalance and iron deficiency. Their action is correspondingly comprehensive, in increasing or decreasing recruitment and translation of most cellular mRNAs, and specialized, in targeting translation of mRNAs with regulatory features such as a 5’ terminal oligopyrimidine tract (TOP), upstream open reading frames (uORFs), or an internal ribosomal entry site (IRES). In mammals, two major pathways are linked to targeted mRNA translation. The target of rapamycin (TOR) kinase induces translation of TOP and perhaps other subsets of mRNAs, whereas a family of eIF2 kinases does so with mRNAs containing uORFs or an IRES. TOR targets translation of mRNAs that code for proteins involved in translation, an action compatible with its widely accepted role in regulating cellular growth. The four members of the eIF2 kinase family increase translation of mRNAs coding for stress response proteins such as transcription factors and chaperones. Though all four kinases act on one main substrate, eIF2, published literature demonstrates both common and unique effects by each kinase in response to its specific activating stress. This suggests that the activated eIF2 kinases regulate the translation of both a global and a specific set of mRNAs. Up to now, few studies have attempted to test such a hypothesis; none has been done in mammals. Also unexplored is the interaction between stress-induced inhibition of translation initiation by the eIF2 kinases and its effect on TOR signaling. This is particularly relevant given that both pathways are regulated by some of the same environmental signals, such as nutrient availability. The work presented in this thesis addresses these two topics. The resulting data demonstrate a dependency on translation initiation status for the regulation of downstream targets of TOR, and identify sets of mRNAs whose translation is commonly or uniquely regulated by two of the eIF2 kinases.