Li Bingsheng, Li Pan, Xia Weiping, You Baiyang, Yu Qingfeng, Zhang Bo, Huang Ru, Wang Ruixiao, Liu Yuhan, Chen Zhi, Gan Yu, He Yao, Hennenberg Martin, Stief Christian G, Chen Xiang
Department of Urology, Xiangya Hospital, Central South University, Changsha, China.
Department of Urology, University Hospital Munich, LMU Munich, Munich, Germany.
Ann Transl Med. 2021 Sep;9(17):1380. doi: 10.21037/atm-21-3194.
Integrin α2β1 inhibitor BTT-3033 (1-(4-fluorophenyl)-N-methyl-N-[4[[(phenylamino)carbonyl]amino]phenyl]-1H-pyrazole-4-sulfonamide) was recently reported to inhibit neurogenic and thromboxane A2-induced human prostate smooth muscle contraction, and thus represents a target with a different inhibition spectrum than that of α1-blockers in benign prostate hyperplasia (BPH) treatments. Clarifying the underlying mechanisms of the inhibition effects will provide insights into the role of integrin α2β1 in prostate contraction and enable new intracellular targets for smooth muscle contraction to be explored.
ProteomeHD was used to predict and enrich the top co-regulated proteins of integrin α2 (ITGA2). A phosphoproteomic analysis was conducted on human prostate stromal cells (WPMY-1) treated with 1 or 10 µM of BTT-3033 or solvent for controls. A clustering analysis was conducted to identify the intracellular targets that were inhibited in a dose-dependent manner. Gene ontology (GO) and annotation enrichments were conducted to examine any functional alterations and identify possible downstream targets. A Kinase-substrate enrichment analysis (KSEA) was conducted to identify kinases-substrate relationships.
Enrichments of the actin cytoskeleton and guanosine triphosphatases (GTPases) signaling were predicted from the co-regulated proteins with ITGA2. LIM domain kinases, including LIM domain and actin-binding 1 (LIMA1), zyxin (ZYX), and thyroid receptor-interacting protein 6 (TRIP6), which are functionally associated with focal adhesions and the cytoskeleton, were present in the clusters with dose-dependent phosphorylation inhibition pattern. 15 substrates were dose-dependently inhibited according to the KSEA, including polo-like kinase 1 (PLK1), and GTPases signaling proteins, such as disheveled segment polarity protein 2 (DVL2).
In this study, we proposed that the mechanisms underlying the contractile and proliferative effects of integrin α2β1 are the LIM domain kinases, including the ZYX family, and substrates, including PLK1 and DVL2.
整合素α2β1抑制剂BTT - 3033（1 - （4 - 氟苯基）-N - 甲基 - N - [4[[（苯基氨基）羰基]氨基]苯基]-1H - 吡唑 - 4 - 磺酰胺）最近被报道可抑制神经源性和血栓素A2诱导的人前列腺平滑肌收缩，因此在良性前列腺增生（BPH）治疗中代表了一种与α1阻滞剂具有不同抑制谱的靶点。阐明其抑制作用的潜在机制将有助于深入了解整合素α2β1在前列腺收缩中的作用，并能够探索平滑肌收缩的新细胞内靶点。
使用ProteomeHD预测并富集整合素α2（ITGA2）的顶级共调节蛋白。对用1或10 μM BTT - 3033处理的人前列腺基质细胞（WPMY - 1）或作为对照的溶剂进行磷酸化蛋白质组分析。进行聚类分析以鉴定以剂量依赖性方式被抑制的细胞内靶点。进行基因本体（GO）和注释富集以检查任何功能改变并确定可能的下游靶点。进行激酶 - 底物富集分析（KSEA）以鉴定激酶 - 底物关系。
从与ITGA2共调节的蛋白质中预测到肌动蛋白细胞骨架和鸟苷三磷酸酶（GTPases）信号的富集。与粘着斑和细胞骨架功能相关的LIM结构域激酶，包括LIM结构域和肌动蛋白结合1（LIMA1）、斑联蛋白（ZYX）和甲状腺受体相互作用蛋白6（TRIP6），存在于具有剂量依赖性磷酸化抑制模式的聚类中。根据KSEA，15种底物被剂量依赖性抑制，包括polo样激酶1（PLK1）和GTPases信号蛋白，如无序段极性蛋白2（DVL2）。
在本研究中，我们提出整合素α2β1的收缩和增殖作用的潜在机制是LIM结构域激酶，包括ZYX家族，以及底物，包括PLK1和DVL2。
