Instituto de Ciencias Biomedicas, Departamento de Biologia Celular e do Desenvolvimento, Universidade de Sao Paulo, Av. Prof. Lineu Prestes, 1524, São Paulo, SP, 05508-000, Brazil.
Laboratório de Ciclo Celular, Center of Toxins, Immune Response and Cell Signaling-CeTICS, Instituto Butantan, Av. Vital Brasil, 1500, São Paulo, SP, 05503-900, Brazil.
Cell Commun Signal. 2021 Aug 14;19(1):86. doi: 10.1186/s12964-021-00758-3.
Maspin (SERPINB5) is a potential tumor suppressor gene with pleiotropic biological activities, including regulation of cell proliferation, death, adhesion, migration and gene expression. Several studies indicate that nuclear localization is essential for maspin tumor suppression activity. We have previously shown that the EGFR activation leads to maspin nuclear localization in MCF-10A cells. The present study investigated which EGFR downstream signaling molecules are involved in maspin nuclear localization and explored a possible role of cell-cell contact in this process.
MCF-10A cells were treated with pharmacological inhibitors against EGFR downstream pathways followed by EGF treatment. Maspin subcellular localization was determined by immunofluorescence. Proteomic and interactome analyses were conducted to identify maspin-binding proteins in EGF-treated cells only. To investigate the role of cell-cell contact these cells were either treated with chelating agents or plated on different cell densities. Maspin and E-cadherin subcellular localization was determined by immunofluorescence.
We found that PI3K-Akt and JAK2-STAT3, but not MAP kinase pathway, regulate EGF-induced maspin nuclear accumulation in MCF-10A cells. We observed that maspin is predominantly nuclear in sparse cell culture, but it is redistributed to the cytoplasm in confluent cells even in the presence of EGF. Proteomic and interactome results suggest a role of maspin on post-transcriptional and translation regulation, protein folding and cell-cell adhesion.
Maspin nuclear accumulation is determined by an interplay between EGFR (via PI3K-Akt and JAK2-STAT3 pathways) and cell-cell contact. Video Abstract.
Maspin（丝氨酸蛋白酶抑制剂 B5）是一种潜在的肿瘤抑制基因，具有多种生物学活性，包括调节细胞增殖、死亡、黏附、迁移和基因表达。多项研究表明，核定位对于 Maspin 的肿瘤抑制活性至关重要。我们之前已经表明，EGFR 的激活导致 MCF-10A 细胞中 Maspin 的核定位。本研究调查了 EGFR 下游信号分子中哪些参与了 Maspin 的核定位，并探讨了细胞-细胞接触在此过程中的可能作用。
用针对 EGFR 下游途径的药理学抑制剂处理 MCF-10A 细胞，然后用 EGF 处理。通过免疫荧光法确定 Maspin 的亚细胞定位。进行蛋白质组学和相互作用组学分析，以鉴定仅在 EGF 处理的细胞中与 Maspin 结合的蛋白质。为了研究细胞-细胞接触的作用，这些细胞要么用螯合剂处理，要么在不同的细胞密度下培养。通过免疫荧光法确定 Maspin 和 E-钙黏蛋白的亚细胞定位。
我们发现，PI3K-Akt 和 JAK2-STAT3 途径，但不是 MAP 激酶途径，调节 MCF-10A 细胞中 EGF 诱导的 Maspin 核积累。我们观察到，在稀疏细胞培养中，Maspin 主要位于核内，但即使存在 EGF，在细胞汇合时也会重新分布到细胞质中。蛋白质组学和相互作用组学结果表明 Maspin 在转录后和翻译调控、蛋白质折叠和细胞-细胞黏附中发挥作用。
Maspin 的核积累是由 EGFR（通过 PI3K-Akt 和 JAK2-STAT3 途径）和细胞-细胞接触之间的相互作用决定的。视频摘要。
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