Scribble knockdown induced metastasis, identification of its associated novel molecular candidates through proteome...

Metastasis is the primary cause of cancer associated deaths globally. Loss of function of Scribble, a cell polarity regulator and tumor suppressor gene, is associated with many forms of human cancers but its role in cell proliferation and metastasis remains unknown. We generated metastatic cancer condition in Drosophila using UAS^RNAi-GAL4 system by knockdown of Scribble in the wing imaginal discs and tracked metastasis events from early to late pupae (0hr-84 h s) using fluorescence microscopy. Here, we report, for the first time, that the knockdown of Scribble alone could lead to the development of primary tumor in the wing imaginal discs, which is capable of establishing metastasis, apparently leading to secondary tumor formation in pupae at early stage, eventually resulting in absolute pupal lethality without organ development. MMP1, a metastasis biomarker, levels were assessed during pre-and post-metastatic phases in pupae using qRT-PCR and Western blot analysis. Further, we analyzed the proteome of Scribble knockdown induced tumor-bearing pupae by 2-D gel electrophoresis followed by MALDI-TOF MS to identify some novel proteins possibly involved in the progression of tumorigenesis and metastasis events. Six differentially expressed proteins, Obp 99b, Fer2LCH,CG13492, Hsp23, Ubiquitin and Colt, were identified in Scrib knockdown pupae and validated their expression using qRT-PCR. Thus, our results suggested that loss of Scrib alone capable of causing metastasis, without the need for cooperative interaction with oncogenic Ras. The newly identified proteins could be important candidates for biomarker/therapeutic target against Scrib associated metastatic cancers.