TY - JOUR
T1 - Eukaryotic translation initiation factor eIF5 promotes the accuracy of start codon recognition by regulating Pi release and conformational transitions of the preinitiation complex
AU - Saini, Adesh K.
AU - Nanda, Jagpreet S.
AU - Martin-Marcos, Pilar
AU - Dong, Jinsheng
AU - Zhang, Fan
AU - Bhardwaj, Monika
AU - Lorsch, Jon R.
AU - Hinnebusch, Alan G.
N1 - Funding Information:
Intramural Program of the National Institutes of Health [in part to A.G.H. and J.R.L.]; National Institutes of Health Grant [GM62128] [formerly to J.R.L.]; Department of Science and Technology, Government of India Grant [Int/NZ/P-2/13] [to A.K.S.]. Funding for open access charge: Intramural Program of the National Institutes of Health [to A.G.H. and J.R.L.]; National Institutes of Health Grant [GM62128] [formerly to J.R.L.]; Department of Science and Technology, Government of India Grant [Int/NZ/P-2/13] [to A.K.S.].
Funding Information:
Intramural Program of the National Institutes of Health [in part to A.G.H. and J.R.L.]; National Institutes of Health Grant [GM62128] [formerly to J.R.L.]; Department of Science and Technology, Government of India Grant [Int/NZ/P-2/13] [to A.K.S.]. Funding for open access charge: Intramural Program of the National Institutes of Health [ to A.G.H. and J.R.L.]; National Institutes of Health Grant [GM62128] [formerly to J.R.L.]; Department of Science and Technology, Government of India Grant [Int/NZ/P-2/13] [to A.K.S.]. Conflict of interest statement. None declared.
PY - 2014/9/2
Y1 - 2014/9/2
N2 - eIF5 is the GTPase activating protein (GAP) for the eIF2·GTP·Met-tRNAiMet ternary complex with a critical role in initiation codon selection. Previous work suggested that the eIF5 mutation G31R/SUI5 elevates initiation at UUG codons by increasing GAP function. Subsequent work implicated eIF5 in rearrangement of the preinitiation complex (PIC) from an open, scanning conformation to a closed state at AUG codons, from which Pi is released from eIF2·GDP·Pi. To identify eIF5 functions crucial for accurate initiation, we investigated the consequences of G31Ron GTP hydrolysis and Pi release, and the effects of intragenic G31Rsuppressors on these reactions, and on the partitioning of PICs between open and closed states. eIF5-G31R altered regulation of Pi release, accelerating it at UUG while decreasing it at AUG codons, consistent with its ability to stabilize the closed complex at UUG. Suppressor G62S mitigates both defects of G31R, accounting for its efficient suppression of UUG initiation in G31R, G62S cells; however suppressor M18V impairs GTP hydrolysis with little effect on PIC conformation. The strong defect in GTP hydrolysis conferred by M18V likely explains its broad suppression of Sui- mutations in numerous factors. We conclude that both of eIF5's functions, regulating Pi release and stabilizing the closed PIC conformation, contribute to stringent AUG selection in vivo.
AB - eIF5 is the GTPase activating protein (GAP) for the eIF2·GTP·Met-tRNAiMet ternary complex with a critical role in initiation codon selection. Previous work suggested that the eIF5 mutation G31R/SUI5 elevates initiation at UUG codons by increasing GAP function. Subsequent work implicated eIF5 in rearrangement of the preinitiation complex (PIC) from an open, scanning conformation to a closed state at AUG codons, from which Pi is released from eIF2·GDP·Pi. To identify eIF5 functions crucial for accurate initiation, we investigated the consequences of G31Ron GTP hydrolysis and Pi release, and the effects of intragenic G31Rsuppressors on these reactions, and on the partitioning of PICs between open and closed states. eIF5-G31R altered regulation of Pi release, accelerating it at UUG while decreasing it at AUG codons, consistent with its ability to stabilize the closed complex at UUG. Suppressor G62S mitigates both defects of G31R, accounting for its efficient suppression of UUG initiation in G31R, G62S cells; however suppressor M18V impairs GTP hydrolysis with little effect on PIC conformation. The strong defect in GTP hydrolysis conferred by M18V likely explains its broad suppression of Sui- mutations in numerous factors. We conclude that both of eIF5's functions, regulating Pi release and stabilizing the closed PIC conformation, contribute to stringent AUG selection in vivo.
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U2 - 10.1093/nar/gku653
DO - 10.1093/nar/gku653
M3 - Article
C2 - 25114053
AN - SCOPUS:84960528165
SN - 0305-1048
VL - 42
SP - 9623
EP - 9640
JO - Nucleic acids research
JF - Nucleic acids research
IS - 15
ER -