背景:炎性肌纤维母细胞瘤(IMT)的特征是肌纤维母细胞增殖和炎性细胞浸润。我们先前对IMT的研究表明,破坏NMD途径会降低触发免疫细胞浸润的阈值,从而导致不适当的免疫激活。然而,肌成纤维细胞的分化和增殖尚不清楚。
方法:RT-PCR,RT-qPCR,DNA序列,西方螺栓,在这项研究中使用了种族分析和位点特异性诱变。
结果:这里,在2个胰腺IMT中发现了选择性剪接(ALS)UPF2mRNA跳过外显子2和3,并对应于截短的UPF2蛋白。我们表明,存在于UPF2mRNA的5'UTR中的uORF负责翻译抑制,而ALSUPF2更容易翻译成截短的UPF2蛋白。在IMT样品中,NMD的几个mRNA靶标被上调,表示截断的UPF2函数受到强烈扰动,导致IMT中NMD通路中断。这些上调的NMD目标包括cdkn1a表达和高水平p21(waf1/cip1)的产生,这可能有助于触发IMT。
结论:UPFs/NMD通路的破坏可能与IMT的分化和增殖相关的分子改变有关。
Inflammatory myofibroblastic tumors (IMTs) are characterized by myofibroblast proliferation and an inflammatory cell infiltrate. Our previous study on IMTs reveals that disrupt NMD pathway causes to lower the threshold for triggering the immune cell infiltration, thereby resulting in inappropriate immune activation. However, myofibroblast differentiation and proliferation is not yet known.
RT-PCR, RT-qPCR, DNA sequence, western bolt, 5\'race analysis and site-specific mutagenesis were used in this study.
Here, an alternative spliced (ALS) UPF2 mRNA skipping exon 2 and 3 and corresponding to the truncated UPF2 protein were found in 2 pancreatic IMTs. We showed that the uORF present in the 5\'UTR of UPF2 mRNA is responsible for the translation inhibition, whiles ALS UPF2 is more facilitated to be translated into the truncated UPF2 protein. Several mRNA targets of the NMD were upregulated in IMT samples, indicating that the truncated UPF2 function is strongly perturbed, resulted in disrupted NMD pathway in IMTs. These upregulated NMD targets included cdkn1a expression and the generation of high levels of p21 (waf1/cip1), which may contribute to triggering IMTs.
The disrupt UPFs/NMD pathway may link to molecular alteration associated with differentiation and proliferation for IMTs.