The auto-inhibitory function of importin alpha is essential in vivo.
Proteins that contain a classical nuclear localization signal (NLS) are recognized in the cytoplasm by a heterodimeric import receptor composed of importin/karyopherin alpha and beta. The importin alpha subunit recognizes classical NLS sequences, and the importin beta subunit directs the complex to the nuclear pore. Recent work shows that the N-terminal importin beta binding (IBB) domain of importin alpha regulates NLS-cargo binding in the absence of importin beta in vitro. To analyze the in vivo functions of the IBB domain, we created a series of mutants in the Saccharomyces cerevisiae importin alpha protein. These mutants dissect the two functions of the N-terminal IBB domain, importin beta binding and auto-inhibition. One of these importin alpha mutations, A3, decreases auto-inhibitory function without impacting binding to importin beta or the importin alpha export receptor, Cse1p. We used this mutant to show that the auto-inhibitory function is essential in vivo and to provide evidence that this auto-inhibitory-defective importin alpha remains bound to NLS-cargo within the nucleus. We propose a model where the auto-inhibitory activity of importin alpha is required for NLS-cargo release and the subsequent Cse1p-dependent recycling of importin alpha to the cytoplasm.
Harreman
Michelle T
MT
Department of Biochemistry, School of Medicine and the Graduate Program in Biochemistry, Cell and Developmental Biology, Emory University, Atlanta, Georgia 30322, USA.
Hodel
Mary R
MR
Fanara
Patrizia
P
Hodel
Alec E
AE
Corbett
Anita H
AH
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D002467:Cell Nucleus / Q000502:physiology; D006801:Humans; D008954:Models, Biological; D008969:Molecular Sequence Data; D021122:Protein Subunits / Q000502:physiology; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D016415:Sequence Alignment; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D028901:alpha Karyopherins / Q000037:antagonists & inhibitors / Q000502:physiology*; D028961:beta Karyopherins / Q000502:physiology*
D021122:Protein Subunits; D011994:Recombinant Proteins; D028901:alpha Karyopherins; D028961:beta Karyopherins