Cloning of murine translation initiation factor 6 and functional analysis of the homologous sequence YPR016c in Saccharomyces cerevisiae.
The cDNA sequence of a murine gene whose expression was up-regulated after epidermal injury was cloned utilizing differential display. The full-length cDNA was isolated by 3' and 5' rapid amplification of cDNA ends from mouse liver. The predicted protein is >97% identical to the human sequence for eukaryotic translation initiation factor (eIF) 6, thus identifying the gene as murine eIF6. Functional studies of the yeast eIF6 homolog, YPR016c, were initiated in Saccharomyces cerevisiae to determine the cellular role(s) of eIF6. Complete deletion of the YPR016c coding sequence was lethal. Viability was restored in the presence of either YPR016c or murine eIF6, when either was expressed as amino-terminal green fluorescent protein fusion protein. Moreover, both fusion proteins localized to nuclear/perinuclear compartments in their respective yeast strains. When the expression of YPR016c-green fluorescent protein was repressed, there was a dramatic reduction in the 60 S ribosomal subunit and polysome content and decreased 80S monosome content. Additionally, the YPR016c-depleted cells arrested in G1. These studies show that YPR016c, which encodes yeast eIF6, is necessary for maximal polysome formation and plays an important role in determining free 60 S ribosomal subunit content.
Wood
L C
LC
Dermatology and Medical Services, Department of Veterans Affairs Medical Center and Departments of Dermatology and Medicine, University of California, San Francisco, California 94121, USA.
Ashby
M N
MN
Grunfeld
C
C
Feingold
K R
KR
D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D010448:Peptide Initiation Factors / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D017931:DNA Primers; D018076:DNA, Complementary; D005656:Fungal Proteins; D008164:Luminescent Proteins; D010448:Peptide Initiation Factors; D011993:Recombinant Fusion Proteins; C021019:eIF-6; D049452:Green Fluorescent Proteins