Dynamic behavior of Arabidopsis eIF4A-III, putative core protein of exon junction complex: fast relocation to nucleolus and splicing speckles under hypoxia.
Here, we identify the Arabidopsis thaliana ortholog of the mammalian DEAD box helicase, eIF4A-III, the putative anchor protein of exon junction complex (EJC) on mRNA. Arabidopsis eIF4A-III interacts with an ortholog of the core EJC component, ALY/Ref, and colocalizes with other EJC components, such as Mago, Y14, and RNPS1, suggesting a similar function in EJC assembly to animal eIF4A-III. A green fluorescent protein (GFP)-eIF4A-III fusion protein showed localization to several subnuclear domains: to the nucleoplasm during normal growth and to the nucleolus and splicing speckles in response to hypoxia. Treatment with the respiratory inhibitor sodium azide produced an identical response to the hypoxia stress. Treatment with the proteasome inhibitor MG132 led to accumulation of GFP-eIF4A-III mainly in the nucleolus, suggesting that transition of eIF4A-III between subnuclear domains and/or accumulation in nuclear speckles is controlled by proteolysis-labile factors. As revealed by fluorescence recovery after photobleaching analysis, the nucleoplasmic fraction was highly mobile, while the speckles were the least mobile fractions, and the nucleolar fraction had an intermediate mobility. Sequestration of eIF4A-III into nuclear pools with different mobility is likely to reflect the transcriptional and mRNA processing state of the cell.
Department of Cell Biology, John Ines Centre, Colney, Norwich NR4 7UH, United Kingdom.
O A
OA
Koroleva
G
G
Calder
A F
AF
Pendle
S H
SH
Kim
D
D
Lewandowska
C G
CG
Simpson
I M
IM
Jones
J W S
JW
Brown
P J
PJ
Shaw
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015687:Cell Hypoxia; D002466:Cell Nucleolus / Q000378:metabolism*; D017124:Conserved Sequence; D004791:Enzyme Inhibitors / Q000494:pharmacology; D039601:Eukaryotic Initiation Factor-4A / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000032:analysis; D007976:Leupeptins / Q000494:pharmacology; D008969:Molecular Sequence Data; D021381:Protein Transport / Q000187:drug effects; D011993:Recombinant Fusion Proteins / Q000032:analysis; D016415:Sequence Alignment; D019810:Sodium Azide / Q000494:pharmacology; D020798:Two-Hybrid System Techniques
D029681:Arabidopsis Proteins; D004791:Enzyme Inhibitors; D007976:Leupeptins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D019810:Sodium Azide; D039601:Eukaryotic Initiation Factor-4A; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde