The transport of proteins between the nucleus and the cytoplasm is signal-mediated: nuclear localization and export signals (NLSs and NESs) direct proteins in and out of the nucleus, respectively. The NPC allows ions, small molecules and many small proteins (<40 kDa) to diffuse between the nucleus and cytoplasm, but larger proteins require the assistance of nuclear transport factors to traverse the NPC. 19 Karyopherin-β (Kapβ; also known as Importin and Exportin) proteins in humans recognize NLS/NES, each functioning as distinct nuclear import, export or bidirectional transport factors ( [1
]. Kapβs share similar molecular weights (90–150 kDa) and isoelectric points (pI = 4.0–5.0), low sequence identity (10–20%) and all contain 19–20 helical HEAT repeats arranged into superhelical or ring-like structures.
The Kapβ family of nuclear transport factors.
Each Kapβ recognizes a unique set of proteins or RNA, thus creating multiple transport pathways across the nuclear pore complex (NPC). Kapβs also bind nucleoporins (proteins in the NPC), targeting Kapβ-cargo complexes to the NPC for translocation. Kapβ-cargo interactions and transport directionality are regulated by the Ran GTPase nucleotide cycle [3
]. RanGTP is concentrated in the nucleus, while RanGDP is concentrated in the cytoplasm. In import pathways, RanGTP and cargos bind Kapβs competitively, allowing cargo binding in the cytoplasm and RanGTP-mediated release in the nucleus. In contrast, in export pathways, RanGTP, cargos and Kapβs bind cooperatively for cargo binding in the nucleus and release in the cytoplasm as Ran-bound GTP is hydrolyzed.
The NLS (by extension, also the NES) was defined as a portion of nuclear transport cargo that is 1) necessary for import/export, 2) sufficient to import/export an unrelated protein, 3) able to bind transport factor directly and that 4) transport is abolished when the transport factor is disabled [4
]. These criteria are generally useful until complications arise due to complexities in the cell, such as when transport is mediated by multiple signals and/or transport factors, signals are functional only when appropriately modified or if protein localization is also dictated by nuclear/cytoplasmic retention [5
]. Many transport cargos are known for Importin-β (Impβ; also known as Karyopherin-β1), CRM1 (also known as Exportin1 or Xpo1) and Karyopherin-β2 (Kapβ2; also known as Transportin), but many fewer are known for the other 16 human Kapβs [6
]. Correspondingly, classes of NLS recognized by Impβ, Kapβ2 and an NES for CRM1 have been identified but common properties of signals for the other Kapβs remain unknown. The first characterized nuclear transport signal is the short lysine-rich classical-NLS, which was discovered in the early 1980s and is recognized by Importin-α (Impα; also known as Karyopherin-α), which is an adaptor for Impβ [6
]. More recently, structural analysis of Kapβ2-cargo complexes led to the discovery of a new class of NLS termed PY-NLS [8
]. Finally, many export cargos contain leucine-rich NESs, which are recognized by export-Kapβ CRM1. Structures of the first CRM1-NES complex were reported in 2009 [11••