Department of Molecular Biosciences, Northwestern University, Evanston, IL, 60208, USA.
Department of Biomedical Engineering, Northwestern University, Evanston, IL, 60208, USA.
Commun Biol. 2023 Oct 18;6(1):1033. doi: 10.1038/s42003-023-05398-6.
Nucleoporins (NUPs) comprise nuclear pore complexes, gateways for nucleocytoplasmic transport. As primary human keratinocytes switch from the progenitor state towards differentiation, most NUPs are strongly downregulated, with NUP93 being the most downregulated NUP in this process. To determine if this NUP downregulation is accompanied by a reduction in nuclear pore numbers, we leveraged Stochastic Optical Reconstruction Microscopy. No significant changes in nuclear pore numbers were detected using three independent NUP antibodies; however, NUP reduction in other subcellular compartments such as the cytoplasm was identified. To investigate how NUP reduction influences keratinocyte differentiation, we knocked down NUP93 in keratinocytes in the progenitor-state culture condition. NUP93 knockdown diminished keratinocytes' clonogenicity and epidermal regenerative capacity, without drastically affecting nuclear pore numbers or permeability. Using transcriptome profiling, we identified that NUP93 knockdown induces differentiation genes related to both mechanical and immune barrier functions, including the activation of known NF-κB target genes. Consistently, keratinocytes with NUP93 knockdown exhibited increased nuclear localization of the NF-κB p65/p50 transcription factors, and increased NF-κB reporter activity. Taken together, these findings highlight the gene regulatory roles contributed by differential NUP expression levels in keratinocyte differentiation, independent of nuclear pore numbers.
核孔蛋白（NUPs）构成核孔复合体，是核质转运的门户。在原代人角质形成细胞从祖细胞状态向分化状态转变的过程中，大多数 NUPs 被强烈下调，其中 NUP93 是下调最明显的 NUP。为了确定 NUP 下调是否伴随着核孔数量的减少，我们利用随机光学重建显微镜（Stochastic Optical Reconstruction Microscopy）进行了检测。使用三种独立的 NUP 抗体均未检测到核孔数量的显著变化；然而，在细胞质等其他亚细胞区室中发现了 NUP 的减少。为了研究 NUP 减少如何影响角质形成细胞分化，我们在祖细胞培养条件下敲低了角质形成细胞中的 NUP93。NUP93 敲低降低了角质形成细胞的集落形成能力和表皮再生能力，而对核孔数量或通透性没有明显影响。通过转录组谱分析，我们发现 NUP93 敲低诱导了与机械和免疫屏障功能相关的分化基因，包括已知 NF-κB 靶基因的激活。一致地，NUP93 敲低的角质形成细胞显示 NF-κB p65/p50 转录因子的核定位增加，以及 NF-κB 报告基因活性增加。总之，这些发现强调了 NUP 表达水平的差异在角质形成细胞分化中的基因调控作用，而与核孔数量无关。
