Proteomic identification of target proteins following Drosha knockdown in cervical cancer

Abstract

Abstract. The nuclear microRNA (miRNA) processing enzyme Drosha is upregulated in cervical cancer, and its overexpression is related to an invasive tumour phenotype. However, the mechanisms that underlie this effect remain poorly understood. The aim of this study was to identify the potential targets of Drosha in cervical cancer. Here, we demonstrated that Drosha knockdown (Drosha-KD) inhibited proliferation, colony formation and the migration of cervical cancer cells in vitro. A global upregulation of proteins in Drosha-KD cells was revealed by two-dimensional gel electrophoresis (2-DE). Eighteen proteins were identified by liquid chromatography and tandem mass spectrometry technology (LC-MS/MS) from 21 selected protein spots that exhibited significant alterations in Drosha-KD cells. The majority of the identified proteins have been previously associated with tumour formation. The downregulation of tubulin 5β in Drosha-KD cervical cancer cells was further confirmed by western blotting. Our results suggest that Drosha affects the biological activity of cervical cancer cells by regulating the expression of numerous tumourassociated proteins.

Perturbation of DROSHA and DICER expression by human papillomavirus 16 oncoproteins

“…1B). In summary, our data are partially consistent with a previous report (Zhou et al, 2013), and show that DROSHA mRNA levels are significantly increased in all HPV positive cervical cancer lines, whereas DICER mRNA levels were only significantly increased in the HPV16 positive CaSki and SiHa but not in the HPV18 positive HeLa or the HPV negative C33A cervical carcinoma lines.…”

“…1A). These results partially confirm and extend a previous report that documented increased DROSHA mRNA expression in HeLa, SiHa and C33A cervical cancer cell lines (Zhou et al, 2013). …”

“…DROSHA expression is frequently upregulated and 5p gene amplification involving the DROSHA gene has also been reported in cervical cancers. Our experiments partially confirm and extend a previous publication (Zhou et al, 2013) and document higher DROSHA mRNA levels in HPV16 positive SiHa and CaSki, as well as HPV18 positive Hela, but not in the HPV negative C33A cervical cancer line. We show that HPV16 E6/E7 expression in C33A cells causes an increase in DROSHA expression although not to the levels observed in HPV positive cervical cancer lines.…”

“…DROSHA is the most significantly upregulated transcript associated with this chromosomal gain and DROSHA overexpression has been observed in all tumors with 5p gain, suggesting that DROSHA may be a critical 5p target with a potential role in cervical cancer progression (Muralidhar et al, 2007; Scotto et al, 2008). DROSHA mRNA levels were also shown to be increased in the HPV positive SiHa and HeLa cervical cancer cell lines and to a lesser extent in the C33A HPV negative cervical cancer cell line (Muralidhar et al, 2007(Zhou et al, 2013). …”

The Use of Laser Microdissection in the Identification of Suitable Reference Genes for Normalization of Quantitative Real-Time PCR in Human FFPE Epithelial Ovarian Tissue Samples

“…Furthermore, in accordance with our results, Li et al reported that GAPDH was significantly increased in malignant ovarian tissues. The altered expression of GAPDH was also found in other types of neoplasia, such as human glioma, atopic bronchial epithelial cells, and squamous cervical cancer [22] – [24] .…”