Xiao Michael, Dhungel Sajina, Azad Roksana, Favaro Denize C, Rajesh Rajaian Pushpabai, Gardner Kevin H, Kikani Chintan K
Department of Biology, University of Kentucky, Lexington, KY 40502, USA.
Structural Biology Initiative, CUNY Advanced Science Research Center, New York, NY 10031, USA.
bioRxiv. 2023 Nov 2:2023.09.06.556462. doi: 10.1101/2023.09.06.556462.
The ligand-regulated PAS domains are one of the most diverse signal-integrating domains found in proteins from prokaryotes to humans. By biochemically connecting cellular processes with their environment, PAS domains facilitate an appropriate cellular response. PAS domain-containing Kinase (PASK) is an evolutionarily conserved protein kinase that plays important signaling roles in mammalian stem cells to establish stem cell fate. We have shown that the nuclear translocation of PASK is stimulated by differentiation signaling cues in muscle stem cells. However, the mechanistic basis of the regulation of PASK nucleo-cytoplasmic translocation remains unknown. Here, we show that the PAS-A domain of PASK contains a putative monopartite nuclear localization sequence (NLS) motif. This NLS is inhibited in cells via intramolecular association with a short linear motif, termed the PAS Interacting Motif (PIM), found upstream of the kinase domain. The interaction between the PAS-A domain and PIM is evolutionarily conserved and serves to retain PASK in the cytosol in the absence of signaling cues. Consistent with that, we show that metabolic inputs induce PASK nuclear import, likely by disrupting the PAS-A: PIM association. We suggest that a route for such linkage may occur through the PAS-A ligand binding cavity. We show that PIM recruitment and artificial ligand binding to the PAS-A domain occur at neighboring locations that could facilitate metabolic control of the PAS-PIM interaction. Thus, the PAS-A domain of PASK integrates metabolic signaling cues for nuclear translocation and could be targeted to control the balance between self-renewal and differentiation in stem cells.
配体调节的PAS结构域是从原核生物到人类的蛋白质中发现的最多样化的信号整合结构域之一。通过将细胞过程与其环境进行生化连接，PAS结构域促进了适当的细胞反应。含PAS结构域的激酶（PASK）是一种进化保守的蛋白激酶，在哺乳动物干细胞中发挥重要的信号作用以确立干细胞命运。我们已经表明，PASK的核转位受到肌肉干细胞中分化信号线索的刺激。然而，PASK核质转位调控的机制基础仍然未知。在这里，我们表明PASK的PAS-A结构域包含一个推定的单部分核定位序列（NLS）基序。该NLS在细胞中通过与激酶结构域上游发现的一个短线性基序（称为PAS相互作用基序，PIM）的分子内结合而受到抑制。PAS-A结构域与PIM之间的相互作用在进化上是保守的，并且在没有信号线索的情况下用于将PASK保留在细胞质中。与此一致，我们表明代谢输入可能通过破坏PAS-A：PIM结合诱导PASK核输入。我们认为这种联系的一条途径可能通过PAS-A配体结合腔发生。我们表明PIM募集和人工配体与PAS-A结构域的结合发生在相邻位置，这可能有助于对PAS-PIM相互作用进行代谢控制。因此，PASK的PAS-A结构域整合了用于核转位的代谢信号线索，并且可以作为靶点来控制干细胞自我更新和分化之间的平衡。
