RNA-seq analysis for VCPWT and VCPT76A breast tumor

Proper centrosome movement to the bipolar position is a prerequisite for bipolar spindle orientation and genome stability. Polo like kinase 1 (Plk1) is a key mitotic kinase which controls centrosome separation. Here we found that Plk1 phosphorylated residue Thr76 in VCP/p97, an AAA-ATPase mainly involved in protein homeostasis, at centrosome from prophase to anaphase. This phosphorylation recruited VCP to centrosome to regulate centrosome orientation. VCP also exhibited strong co-localization with Eg5, a mitotic kinesin motor for centrosome movement, at mitotic spindle, and dephosphorylation of Thr76 in VCP was required for the enrichment of VCP and Eg5 to the spindle, thus, ensuring proper spindle orientation and chromosome segregation. Since genome instability is the hallmark of cancer and the phosphorylation of Thr76 in VCP is important for proper centrosome movement, spindle orientation, and chromosome segregation, we, thus, knocked down VCP in MDA-MB-231cells by its specific shRNA, and reconstituted the expression of VCP by infecting the cells with lentivirus encoding shRNA-resistant Flag-VCPWT or Flag-VCPT76A. We then implanted VCPWT- or VCPT76A-expressing MDA-MB-231 cells into nude mice and found that the tumor growth in mice implanted with VCPT76A-expressing cells was significantly slower when compared with the mice implanted with the Flag-VCPWT-expressing cells. To better understand the decreased tumor formation ability of Flag-VCPT76A-expressing cells, we isolated the primary breast tumor tissues for the RNA-Seq analysis. Of the 57,905 mapped whole genome genes in the isolated tumors, we identi?ed 234 differentially expressed genes (DEGs; false discovery rate (FDR) < 0.1). Overall design: Study the genome wide difference between VCPWT and VCPT76A derived breast tumors