Importin is a type ofkaryopherin^[1] that transportsprotein molecules from thecell'scytoplasm to thenucleus. It does so by binding to specificrecognition sequences, callednuclear localization sequences (NLS).

Importin has two subunits, importin α and importin β. Members of the importin-β family can bind and transport cargo by themselves, or can formheterodimers with importin-α. As part of aheterodimer, importin-β mediates interactions with thepore complex, while importin-α acts as an adaptor protein to bind thenuclear localization signal (NLS) on the cargo. The NLS-Importin α-Importin βtrimer dissociates after binding toRanGTP inside thenucleus,^[2] with the two importin proteins being recycled to thecytoplasm for further use.

Importin can exist as either aheterodimer of importin-α/β or as amonomer of Importin-β. Importin-α was first isolated in 1994 by a group includingEnno Hartmann, based at theMax Delbrück Center for Molecular Medicine.^[1] The process of nuclear protein import had already been characterised in previous reviews,^[3] but the key proteins involved had not been elucidated up until that point. A 60 kDacytosolic protein, essential for protein import into the nucleus, and with a 44%sequence identity toSRP1p, was purified fromXenopus eggs. It was cloned, sequenced and expressed inE.coli and in order to completely reconstitute signal dependent transport, had to be combined withRan(TC4). Other key stimulatory factors were also found in the study.^[1]

Importin-β, unlike importin-α, has no directhomologues in yeast, but was purified as a 90-95 kDa protein and found to form aheterodimer with importin-α in a number of different cases. These included a study led by Michael Rexach^[4]and further studies byDirk Görlich.^[5] These groups found that importin-α requires another protein, importin-β to function, and that together they form a receptor fornuclear localization signals (NLS), thus allowing transport into thenucleus. Since these initial discoveries in 1994 and 1995, a host of Importin genes, such asIPO4 andIPO7, have been found that facilitate the import of slightly different cargo proteins, due to their differing structure and locality.

A large proportion of the importin-αadaptor protein is made up of severalarmadillo repeats (ARM) arranged intandem. These repeats can stack together to form a curved-shaped structure, which facilitates binding to theNLS of specific cargo proteins. The major NLS binding site is found towards theN-terminus, with a minor site being found at theC-terminus. As well as theARM structures, Importin-α also contains a 90amino acidN-terminal region, responsible for binding to Importin-β, known as the Importin-β binding (IBB)domain.^[6] This is also a site of autoinhibition,^[7] and is implicated in the release of cargo once importin-α reaches thenucleus.^[8]

Importin-β is the typical structure of a largersuperfamily ofkaryopherins. The basis of their structure is 18-20 tandem repeats of theHEAT motif. Each one of these repeats contains two antiparallelalpha helices linked by aturn, which stack together to form the overall structure of theprotein.^[9]

In order to transport cargo into thenucleus, importin-β must associate with the nuclear pore complexes. It does this by forming weak, transientbonds withnucleoporins at their variousFG (Phe-Gly) motifs.Crystallographic analysis has shown that thesemotifs bind to importin-β at shallowhydrophobic pockets found on its surface.^[10]

The primary function of importin is to mediate the translocation ofproteins withnuclear localization signals into thenucleus, throughnuclear pore complexes (NPC), in a process known as the nuclear protein import cycle.

The first step of this cycle is the binding of cargo. Importin can perform this function as amonomeric importin-βprotein, but usually requires the presence of importin-α, which acts as anadaptor to cargo proteins (via interactions with theNLS). TheNLS is a sequence of basicamino acids that tags theprotein as cargo destined for thenucleus. A cargoprotein can contain either one or two of thesemotifs, which will bind to the major and/or minor binding sites on importin-α.^[11]

Once the cargo protein is bound, importin-β interacts with theNPC, and the complex diffuses into thenucleus from thecytoplasm. The rate ofdiffusion depends on both the concentration of importin-α present in the cytoplasm and also thebinding affinity of importin-α to the cargo. Once inside thenucleus, the complex interacts with theRas-family GTPase,Ran-GTP. This leads to the dissociation of the complex by altering theconformation of importin-β. Importin-β is left bound toRan-GTP, ready to be recycled.^[11]

Now that the importin-α/cargo complex is free of importin-β, the cargo protein can be released into thenucleus. TheN-terminal importin-β-binding (IBB) domain of importin-α contains an auto-regulatory region that mimics theNLS motif.^[7] The release of importin-β frees this region and allows it to loop back and compete for binding with the cargo protein at the majorNLS-binding site. This competition leads to the release of theprotein. In some cases, specific release factors such asNup2 andNup50 can be employed to help release the cargo as well.^[11]

Finally, in order to return to thecytoplasm, importin-α must associate with aRan-GTP/CAS (nuclear export factor) complex which facilitates its exit from thenucleus.CAS (cellular apoptosis susceptibility protein) is part of the importin-β superfamily ofkaryopherins and is defined as a nuclear export factor. Importin-β returns to thecytoplasm, still bound toRan-GTP. Once in thecytoplasm,Ran-GTP ishydrolysed byRanGAP, formingRan-GDP, and releasing the two importins for further activity. It is this hydrolysis ofGTP that provides the energy for the cycle as a whole. In thenucleus, aGEF will chargeRan with aGTP molecule, which is then hydrolysed by aGAP in thecytoplasm, as stated above. It is this activity ofRan that allows for the unidirectional transport ofproteins.^[11]

There are several disease states and pathologies that are associated withmutations or changes in expression of importin-α and importin-β.

Importins are vital regulatoryproteins during the processes ofgametogenesis andembryogenesis. As a result, a disruption in the expression patterns of importin-α has been shown to cause fertility defects inDrosophila melanogaster.^[12]

There have also been studies that link altered importin-α to some cases ofcancer.Breast cancer studies have implicated atruncated form of importin-α in which theNLS binding domain is missing.^[13] In addition, importin-α has been shown to transport thetumour suppressor gene,BRCA1 (breast cancer type 1 susceptibility protein), into thenucleus. The overexpression of importin-α has also been linked with poor survival rates seen in certainmelanoma patients.^[14]

Importin activity is also associated with someviral pathologies. For instance, in the infection pathway of theEbola virus, a key step is the inhibition of the nuclear import ofPY-STAT1. This is achieved by the virus sequestering importin-α in thecytoplasm, meaning it can no longer bind its cargo at theNLS.^[15] As a result, importin cannot function and the cargo protein stays in the cytoplasm.

Many different cargoproteins can be transported into thenucleus by importin. Often, different proteins will require different combinations of α and β in order to translocate. Some examples of different cargo are listed below.

Although importin-α and importin-β are used to describe importin as a whole, they actually represent largerfamilies ofproteins that share a similar structure and function. Various different genes have been identified for both α and β, with some of them listed below. Note that oftenkaryopherin and importin are used interchangeably.

• Importin:IPO4,IPO5,IPO7,IPO8,IPO9,IPO11,IPO13
• Karyopherin-α:KPNA1,KPNA2,KPNA3,KPNA4,KPNA5,KPNA6
• Karyopherin-β:KPNB1

• Karyopherin
• Nuclear localization sequence
• Nuclear pore complex
• Nuclear transport
• Ran (gene)

• Importins at the U.S. National Library of MedicineMedical Subject Headings (MeSH)
• PDB Molecule of the MonthImportins

• Genes on human chromosome 3
• Genes on human chromosome 17
• Protein families
• Transport proteins

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