PLAGL1 gene function during hepatoma cell proliferation.

VEGA BENEDETTI, A F.; VANNI, R; SAUCEDO, CN; PARADA, LA; ZAVATTARI, P

Congreso; LXI Annual Meeting of the Argentinian Society of Clinic Research; 2016

PLAGL1 gene encodes a homonymous Zinc-finger protein that regulates cell cycle arrest and apoptosis. Such regulation occurs through pathways that include p53 and PPARγ, which induce p21Cip1, a cell cycle regulator. To elucidate its role in tumor growth, we studied the transcript and protein levels of PLAGL1, p53, PPARγ and p21 in hepatoma proliferating cells.Hepatoma cell lines HepG2, Huh7, PLC and SkHep1, and normal fibroblasts (control) were cultured according to standard protocols for proliferation studies. Then, cell count, flow cytometry, Western blot and RT-qPCR analyses were performed at 48, 72 and 96hs. The transcript level was quantified by the 2-ΔΔCt method, using PPIA as a reference gene. Protein level was quantified by the ImageJ program, using Actin as a loading control. Statistic analyses were performed by ANOVA.We determined that fibroblasts have lower proliferation rates than cancer cell lines. In general, the PLAGL1 mRNA level was significantly higher in fibroblasts than in tumor cell lines, which exhibited distinct patterns of transcription and expression. In fibroblasts, the PLAGL1 transcript and expression levels decreased significantly after 48hs post release from serum starvation, and then gradually increased until 96hs, while p53 and p21 followed this PLAGL1 pattern. In contrast, the tumor cell lines showed uniform low transcription and expression levels of PLAGL1 during the experimental period, except for SkHep1 cells. Despite of this, the transcript and expression levels of p53 and p21 presented different dynamics among tumor cell lines. Regarding PPARγ, our experiments demonstrated that its transcriptional level was significantly low during proliferation in normal and and tumor cell lines, except only for Huh7.Our results show that there is not a straight forward functional relationship between PLAGL1 and p53, PPARγ and p21 in the cell-cycle control of hepatoma cells.